chondroitin-sulfates and Disease-Models--Animal

Mucopolysaccharidosis IVA (MPS IVA) is a rare disorder caused by mutations in the N-acetylgalactosamine-6-sulfate-sulfatase (

Topics: Animals; Chondrocytes; Chondroitin Sulfates; Chondroitinsulfatases; Disease Models, Animal; Humans; Keratan Sulfate; Mice; Mucopolysaccharidosis IV

Heparin-induced thrombocytopenia (HIT), a drug-induced immunohaematological adverse reaction, is a rare but potentially very severe condition. The main problem for this complex syndrome is its recognition and management, which should be as early as possible to avoid the development of life-threatening complications. Most studies have reported heterogeneous populations of patients with other diseases that potentially induce thrombocytopenia. There is no gold standard diagnostic criteria, and we have established a score with anamnestic criteria that allows us to evaluate the likelihood of HIT. In clinical practice, the diagnosis is based on the analysis of clinical features and laboratory tests. Platelet aggregation test (PAT) and an ELISA test (heparin platelet-induced antibodies) are generally performed by expert laboratories to confirm the occurrence of HIT. In our experience, both tests are concordant in the majority of patients. PAT seems to correlate better with the clinical features while ELISA appears more specific. Regarding their limits, both are complementary in the determination of HIT diagnosis coupled to the clinical score system. The treatment often requires a multidisciplinary approach. Danaparoid (Orgaran) or lepirudin (Refludan) are the two alternative treatments for HIT patients with marketing approval. To avoid further exposure to heparin, every HIT patient should carry a written document that confirms the immunoallergy.

Topics: Animals; Anticoagulants; Chondroitin Sulfates; Dermatan Sulfate; Disease Models, Animal; Drug Combinations; Fibrinolytic Agents; Heparin; Heparitin Sulfate; Hirudin Therapy; Hirudins; Humans; Immunoassay; Platelet Activation; Recombinant Proteins; Thrombocytopenia

Ulcerative colitis (UC) is a high incidence disease worldwide and clinically presents as relapsing and incurable inflammation of the colon. Bilirubin (BR), a natural antioxidant with significant anti-colitic effects, is utilized in preclinical studies as an intestinal disease therapy. Due to their water-insolubility, the design of BR-based agents usually involves complicated chemosynthetic processes, introducing various uncertainties in BR development. After screening numerous materials, it is identified that chondroitin sulfate can efficiently mediate the construction of BR self-assembled nanomedicine (BSNM) via intermolecular hydrogen bonds between dense sulfate and carboxyl of chondroitin sulfate and imino groups of BR. BSNM exhibits pH sensitivity and reactive oxygen species responsiveness, enabling targeted delivery to the colon. After oral administration, BSNM significantly inhibits colonic fibrosis and apoptosis of colon and goblet cells; it also reduces the expression of inflammatory cytokines. Moreover, BSNM maintains the normal level of zonula occludens-1 and occludin to sustain the integrity of intestinal barrier, regulates the macrophage polarization from M1 to M2 type, and promotes the ecological recovery of intestinal flora. Collectively, the work provides a colon-targeted and transformable BSNM that is simple to prepare and is useful as an efficient targeted UC therapy.

Topics: Animals; Bilirubin; Chondroitin Sulfates; Colitis; Colitis, Ulcerative; Colon; Disease Models, Animal; Mice; Mice, Inbred C57BL

Previous research studies have shown that sulfated polysaccharides can inhibit food allergy, but the detailed mechanism remains largely unknown. In this study, RBL-2H3 cells were used to compare the anti-allergic activities of four sulfated polysaccharides, and an ovalbumin (OVA)-sensitized allergic mouse experiment was used to explore their desensitization effect, with regard to the alteration in gut microbiota and immune cell differentiation. Compared with the shark, bovine and porcine chondroitin sulfate, sea cucumber chondroitin sulfate (SCCS) significantly inhibited the degranulation of RBL-2H3 cells. SCCS reduced allergic symptoms and protected the jejunum from injury in mice. Furthermore, SCCS increased the relative abundance of

Topics: Animals; Anti-Allergic Agents; Cattle; Cell Differentiation; Chondroitin Sulfates; Disease Models, Animal; Food Hypersensitivity; Gastrointestinal Microbiome; Mice; Mice, Inbred BALB C; Ovalbumin; Polysaccharides; Sea Cucumbers; T-Lymphocytes, Regulatory

Despite the progress in the therapy of ulcerative colitis (UC), long-lasting UC remission can hardly be achieved in the majority of UC patients. The key pathological characteristics of UC include an impaired mucosal barrier and local inflammatory infiltration. Thus, a two-pronged approach aiming at repairing damaged mucosal barrier and scavenging inflammatory mediators simultaneously might hold great potential for long-term remission of UC. A rectal formulation can directly offer preferential and effective drug delivery to inflamed colon. However, regular intestinal peristalsis and frequent diarrhea in UC might cause transient drug retention. Therefore, a bioadhesive hydrogel with strong interaction with intestinal mucosa might be preferable for rectal administration to prolong drug retention. Here, we designed a bioadhesive hydrogel formed by the cross-linking of sulfhydryl chondroitin sulfate and polydopamine (CS-PDA). The presence of PDA would ensure the mucosa-adhesive behavior, and the addition of CS in the hydrogel network was expected to achieve the restoration of the intestinal epithelial barrier. To scavenge the key player (excessive reactive oxygen species, ROS) in inflamed colon, sodium ferulic (SF), a potent ROS inhibitor, was incorporated into the CS-PDA hydrogel. After rectal administration, the SF-loaded CS-PDA hydrogel could adhere to the colonic mucosa to allow prolonged drug retention. Subsequently, sustained SF release could be achieved to persistently scavenge ROS in inflammatory areas. Meanwhile, the presence of CS would promote the restoration of the mucosal barrier. Ultimately, scavenging ROS and restoring the mucosal barrier could be simultaneously achieved via this SF-loaded bioadhesive hydrogel scaffold. Our two-pronged approach might provide new insight for effective UC treatment.

Topics: Animals; Chondroitin Sulfates; Colitis, Ulcerative; Disease Models, Animal; Humans; Hydrogels; Intestinal Mucosa; Reactive Oxygen Species

Necrotizing enterocolitis (NEC) continues to be a devastating disease in preterm neonates and has a paucity of medical management options. Chondroitin sulfate (CS) is a naturally occurring glycosaminoglycan (GAG) in human breast milk (HM) and has been shown to reduce inflammation. We hypothesized that supplementation with CS in an experimental NEC model would alter microbial diversity, favorably alter the cytokine profile, and (like other sulfur compounds) improve outcomes in experimental NEC via the eNOS pathway. NEC was induced in 5-day-old pups. Six groups were studied (n = 9-15/group): (1) WT breastfed and (2) Formula fed controls, (3) WT NEC, (4) WT NEC + CS, (5) eNOS KO (knockout) NEC, and (6) eNOS KO NEC + CS. Pups were monitored for clinical sickness score and weights. On postnatal day 9, the pups were killed. Stool was collected from rectum and microbiome analysis was done with 16 s rRNA sequencing. Intestinal segments were examined histologically using a well-established injury scoring system and segments were homogenized and analyzed for cytokine profile. Data were analyzed using GraphPad Prism with p < 0.05 considered significant. CS supplementation in formula improved experimental NEC outcomes when compared to NEC alone. CS supplementation resulted in similar improvement in NEC in both the WT and eNOS KO mice. CS supplementation did not result in microbial changes when compared to NEC alone. Our data suggest that although CS supplementation improved outcomes in NEC, this protection is not conferred via the eNOS pathway or alteration of microbial diversity. CS therapy in NEC does improve the intestinal cytokine profile and further experiments will explore the mechanistic role of CS in altering immune pathways in this disease.

Topics: Animals; Chondroitin Sulfates; Cytokines; Dietary Supplements; Disease Models, Animal; Enterocolitis, Necrotizing; Female; Fetal Diseases; Humans; Infant, Newborn; Mice

Novel intra-articular nanoreservoirs were implemented employing different cartilage targeting approaches to improve cartilage bioavailability of a chondroprotective drug, cassic acid (CA), for effective amelioration of cartilage deterioration off-targeting CA gastrointestinal disorders. Herein, we compared active cartilage-targeting approach via chondroitin sulfate (CHS) functionalization versus passive targeting using positively charged nanoparticles to target negatively charged cartilage matrix. Firstly, CA integrated nanoreservoirs (CA-NRs) were fabricated based on ionic conjugation between CA and cationic hydrophobic surface modifier octadecylamine (ODA) and were further functionalized with CHS to develop CHS-CA-NRs. Confocal laser microscope was used to visualize the accumulation of nanoparticles into the cartilage tissue. Both targeting approaches promoted CA local cartilage availability and prolonged its residence time. Compared to passive targeted CA-NRs, active targeted CHS-CA-NRs showed higher fluorescence signals in proximity to and inside chondrocytes which lasted for up to 21 days. In MIA-osteoarthritic rats, CHS-CA-NRs showed superior antiosteoarthritic activity, exhibiting highest cartilage repair compared to CA-NRs. Additionally, CHS-CA-NRs significantly inhibited OA inflammatory cytokine, degradation enzyme and oxidative stress and improved cartilage matrix biosynthesis. Conclusively, CHS-CA-NRs improved OA repair showing a superior efficacy for articular cartilage targeting with CHS which could be a potential advance for OA therapy.

Topics: Animals; Anthraquinones; Cartilage, Articular; Chemistry, Pharmaceutical; Chondroitin Sulfates; Cytokines; Disease Models, Animal; Dose-Response Relationship, Drug; Drug Carriers; Drug Liberation; Lipids; Male; Nanoparticles; Osteoarthritis; Oxidative Stress; Particle Size; Random Allocation; Rats; Rats, Wistar; Surface Properties

Chondroitin sulfate E (CS-E), which is characterized by oversulfated disaccharide units, has been shown to regulate neuronal adhesion, neurite outgrowth and exert neuroprotective effects. In view of these findings, here we investigated the anti-Alzheimer's disease (AD) activities of CSE by using transgenic Caenorhabditis elegans model of Alzheimer's disease. The behavioral experiments demonstrated that CSE at the concentration of 1 mg/mL significantly delayed the worm paralysis caused by Aβ aggregation as compared with control group. Western blot analysis revealed that the level of small oligomers in the transgenic C. elegans was significantly reduced upon treatment with CSE. The number of Aβ plaque deposits in transgenic worm was significantly decreased. In addition, CSE also protected the worms from oxidative stress and rescued chemotaxis dysfunction in transgenic strain CL2355. Taken together, these data suggested that CSE could protect against Aβ-induced toxicity in C. elegans. These results offer valuable evidence for the future use of CSE in the development of agents for the treatment of AD.

Topics: Alzheimer Disease; Amyloid beta-Peptides; Animals; Animals, Genetically Modified; Caenorhabditis elegans; Chondroitin Sulfates; Disease Models, Animal

To explore the therapeutic effect of chondroitin sulfate (CS) on Aspergillus fumigatus (A. fumigatus) keratitis.. The nontoxic concentration of CS was determined using the Draize eye test and cell counting kit-8. Cell scratch test and cell proliferation test were evaluated the impact of CS on the proliferation and migration of human corneal epithelial cells (HCECs). Adherence assay and plate counting were used to detect fungal load in vivo and in vitro, respectively. Clinical score and hematoxylin-eosin (HE) staining were applied to assess the therapeutic effects of CS in an A. fumigatus keratitis mice model. The neutrophil infiltration and activity were detected by flow cytometry (FCM), immunofluorescence staining, and myeloperoxidase (MPO) assay. Toll-like receptor 4 (TLR-4) expression in RAW 264.7 cells and mouse cornea was detected by immunofluorescence staining. Real-time PCR (RT-PCR), western blot, and enzyme-linked immunosorbent assay (ELISA) were applied to examine the expression of inflammatory mediators.. CS at 400 μg/mL (non-cytotoxic) significantly promoted proliferation and migration of HCECs. In an A. fumigatus keratitis mice model, CS treatment alleviated fungal keratitis (FK) severity by decreasing corneal fungal load and inhibiting neutrophil infiltration. In RAW 264.7 cells, the mRNA and protein levels of TLR-4, phosphorylated nuclear factor (NF)-κB (p-NF-κB), interleukin-1β (IL-1β), interleukin-6 (IL-6), tumor necrosis factor-a (TNF-α), cyclooxygenase 2 (COX-2), and macrophage inflammatory protein-2 (MIP-2) were remarkably lower in the siTLR-4 treated group, while higher in rTLR-4 treated group than in the corresponding control group. CS treatment suppressed rTLR-4 induced the mRNA and protein expression of TLR-4, p-NF-κB, IL-1β, IL-6, COX-2, TNF-α, and MIP-2 in RAW cells.. CS may ameliorate the prognosis of A. fumigatus keratitis by promoting corneal epithelial proliferation, inhibiting the recruitment and activity of neutrophils, and inhibiting the inflammatory response by down-regulation of the TLR-4/NF-κB signaling.

Topics: Animals; Aspergillosis; Aspergillus fumigatus; Chondroitin Sulfates; Cyclooxygenase 2; Disease Models, Animal; Interleukin-6; Keratitis; Mice; Mice, Inbred C57BL; NF-kappa B; RNA, Messenger; Toll-Like Receptor 4; Tumor Necrosis Factor-alpha

Lignans are plant-derived compounds that act as partial estrogen agonists. Chondroitin sulfate proteoglycans (CSPGs) represent one of the major components of the extracellular matrix. Here we aimed to understand the role of sesamin (SES), a major lignan compound, in the biosynthesis and degradation of CSPGs in the mouse hippocampus because CSPGs play a key role in the regulation of cognitive functions through the promotion of adult neurogenesis. The expression of the pro-inflammatory cytokine interleukin-1β was decreased by SES administration in the hippocampus of lipopolysaccharide (LPS)-treated mice, a model of neuroinflammation-induced cognitive deficits. The expression of genes related to biosynthesis and degradation of CSPGs in the hippocampus of LPS-treated mice was both increased and decreased by SES administration. Further, the diffuse extracellular matrix labeling of CSPGs by Wisteria floribunda agglutinin (WFA) in the hippocampus of LPS-treated mice was increased by SES administration. The densities of neural stem cells, late transit-amplifying cells, and newborn-granule cells in the hippocampus of LPS-treated mice were also increased by SES administration. Moreover, SES-induced alterations in gene expression, WFA labeling, and adult neurogenesis in LPS-treated mice were more evident in the dorsal hippocampus (center of cognition) than in the ventral hippocampus (center of emotion). Neither LPS nor SES administration affected locomotor activity, anxiety-like behavior, and depression-related behavior. However, impairments in contextual memory and sensorimotor gating in LPS-treated mice were recovered by SES administration. Our results show that SES can promote adult hippocampal neurogenesis through the upregulation of CSPGs, which may alleviate cognitive deficits induced by neuroinflammation.

Topics: Animals; Chondroitin Sulfate Proteoglycans; Chondroitin Sulfates; Cognition; Dioxoles; Disease Models, Animal; Hippocampus; Lignans; Lipopolysaccharides; Mice; Mice, Inbred C57BL; Neuroinflammatory Diseases; Up-Regulation

Growing evidence for the importance of the gut-brain axis in Parkinson's disease (PD) has attracted researchers' interest in the possible application of microbiota-based treatment approaches. Using a 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-induced mouse model, we looked into the prospect of treating PD with fucosylated chondroitin sulfate obtained from sea cucumbers

Topics: 1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine; alpha-Synuclein; Animals; Chondroitin Sulfates; Disease Models, Animal; DNA, Bacterial; Dopamine; Dysbiosis; Inflammation; Intestines; Mice; Mice, Inbred C57BL; NF-kappa B; Parkinson Disease; Polysaccharides; RNA, Ribosomal, 16S; Sea Cucumbers

Lack of highly expressed tumor target and ligands limits application of nano-medicine against triple-negative breast cancer (TNBC). Previous study reported that placenta-derived oncofetal chondroitin sulfate glycosaminoglycan chain (CSA) expressed on 90% of stage I-III invasive ductal breast carcinomas. Our study found the CSA anchor protein VAR2CSA derived small peptide plCSA had strong binding activity with TNBC cell lines and tumor tissue. Here, we combined the AIEgens TBZ-DPNA and therapy drug paclitaxel (PTX) to fabricate near-infrared fluorescence-guided nanodrug (plCSA-NP) to investigate its targeting and anti-tumor effect on TNBC.. We synthesized and purified TBZ-DPNA with one step, measured optical properties and photoluminescence (PL) spectra. We prepared nanodrug plCSA-NP by encapsulating TBZ-DPNA and PTX and conjugating them with peptide plCSA. We evaluated plCSA-NP targeting activity by examining AIEdots fluorescence signal on TNBC cell lines and subcutaneous and lung metastatic mouse model. We assessed PTX delivery effect by cytotoxicity assay on TNBC line and tumor growth of subcutaneous and lung metastatic mouse models.. PL spectra and TEM imaging results showed plCSA-NP had maximum emission feature at 718 nm and nearly monodispersed nanosphere with an average diameter of 70 nm. In vitro studies showed plCSA-NPs had high affinity and cytotoxicity with TNBC cell lines. In vivo subcutaneous and lung metastasis mouse studies showed plCSA-NPs accumulated on TNBC tumor tissue, and significantly prevented TNBC subcutaneous and lung metastasis tumor growth.. In conclusion, we provide solid evidence for chondroitin sulfate targeting peptide plCSA guided nanodrug, exhibit good targeting efficiency and therapeutic effect against TNBC primary and lung metastatic tumor growth.

Topics: Animals; Chondroitin Sulfates; Disease Models, Animal; Humans; Lung; Lung Neoplasms; Mice; Nanospheres; Paclitaxel; Triple Negative Breast Neoplasms

Chondroitin sulfate (CS) is a glycosaminoglycan with proven anti-inflammatory, anti-apoptotic, anti-oxidant properties. CS increases type II collagen and proteoglycan synthesis in human joint chondrocytes. CS can reduce the production of pro-inflammatory mediators and proteases to improve the anabolic/catabolic balance of the extracellular cartilage matrix (ECM). Due to these characteristics, it is a natural compound that is considered to be Symptomatic Slow-Acting Drugs for Osteoarthritis (SYSADOA). Microbial chondroitin sulfate (MCS) was produced from two different bacterial sources using biotechnological methods by our team. In this study, we aimed to apply microbially produced CS and bovine-derived commercial CS forms to rabbit knees with osteoarthritis experimentally and to evaluate the results.. In this study, a cruciate ligament cutting model was applied to 40 New Zealand rabbits to induce experimental osteoarthritis. Four weeks after the surgical procedure, rabbits were divided into 4 groups as control, animal-derived MCS, E coli-derived MCS and PaJC-derived MCS group. The standard rabbit diet was fed to the control group, and the other groups were additionally fed 17 mg/kg/day CS/MCS for 12 weeks. The rabbits were sacrificed at the 12th week after surgery and the preparations obtained were evaluated histopathologically.. As a result, it was observed that regeneration tissue was statistically significant in histopathological cartilage tissue compared to the control group of CS developed from different sources given to rabbits with osteoarthritis. It was determined that among the CS groups produced from different sources, the group with the highest chondroprotective effect was MCS originating from E.coli.. This vegan product (MCS), which we obtained as a result of our study, was produced by our team from a microbial source. According to our analysis, it has the potential to be an effective alternative therapy agent in the treatment of osteoarthritis.

Topics: Animals; Arthritis, Experimental; Cattle; Chondroitin Sulfates; Disease Models, Animal; Escherichia coli; Osteoarthritis, Knee; Rabbits

Our previous study found that low molecular weight chondroitin sulfate (LMWCS) had neuroprotective effects against the toxicity of amyloid-β (Aβ) peptides both in vitro and in vivo, and we speculated that the effects might be related with its anti-oxidative activities. In this study, the anti-Alzheimer's disease (AD) activity of LMWCS was further studied in 5XFAD transgenic mice. After 4-month gavage, the levels of Aβ

Topics: Alzheimer Disease; Amyloid beta-Peptides; Amyloid beta-Protein Precursor; Animals; Behavior, Animal; Chondroitin Sulfates; Cytokines; Disease Models, Animal; Mice; Mice, Transgenic; Molecular Weight; Neuroinflammatory Diseases; Neuroprotective Agents; Oxidative Stress; Presenilin-1; tau Proteins

Calcified cartilage is a mineralized osteochondral interface region between the hyaline cartilage and subchondral bone. There are few reported artificial biomaterials that could offer bioactivities for substantial reconstruction of calcified cartilage. Herein we developed new poly(L-lactide-

Topics: Animals; Biocompatible Materials; Bone Diseases; Calcification, Physiologic; Cartilage; Chondroitin Sulfates; Disease Models, Animal; Glass; Membranes, Artificial; Polyesters; Prostheses and Implants; Rabbits; Tensile Strength; X-Ray Microtomography

To determine alterations of chondroitin sulfate (CS) that reflect cartilage damage in an experimental osteoarthritis (OA) model as well as in human OA samples.. Rats were subjected to anterior cruciate ligament transection (ACLT; OA) or a sham procedure and sacrificed 14, 28, or 70 days after ACLT for histopathology and analysis of extracted CS. Cartilage samples from 14 patients undergoing hip or shoulder arthroplasty secondary to OA or fracture (control) were subjected to the same protocol. The CS content (µg/mg dry cartilage) after proteolysis was determined by densitometry, using agarose-gel electrophoresis. Molar mass (MM) and peak MM of CS were determined using high-performance size-exclusion chromatography (HPSEC).. OA and sham joints at 70 d had 24 [22-24] and 3 [1-6] median histopathology scores, respectively (p < 0.001). Relative CS content (77.7 ± 8.3 µg/mg) was significantly increased in OA samples 70 d after ACLT, as compared to sham (53.5 ± 10.0 µg/mg). Peak MM of CS was higher in OA than in sham samples (P < 0.05). Similarly, CS content and peak MM were higher in cartilage from human OA patients, as compared to fracture samples, reproducing experimental data.. Cartilage matrix from experimental and human OA samples has increased in the relative CS content. Increase in the peak MM distinguishes CS of the extracellular matrix of OA from normal cartilage.

Topics: Animals; Anterior Cruciate Ligament; Cartilage, Articular; Chondroitin Sulfates; Disease Models, Animal; Extracellular Matrix; Humans; Osteoarthritis; Rats

Components of the extracellular matrix (ECM) are overexpressed in fibrotic liver. Collagen is the main component of the liver fibrosis stroma. Here we demonstrate that chondroitin sulfate coated multilayered 50-nm nanoparticles encapsulating collagenase and silibinin (COL + SLB-MLPs) break down the dense collagen stroma, while silibinin inhibits activated hepatic stellate cells. The nanoparticles were taken up to a much greater extent by hepatic stellate cells than by normal hepatocytes, and they down-regulated production of type I collagen. In addition, chondroitin sulfate protected the collagenase from premature deactivation. COL + SLB-MLPs were delivered to the cirrhotic liver, and the collagenase and silibinin synergistically inhibited fibrosis in mice. Immunofluorescence staining of liver tissues revealed that CD44, mediated by chondroitin sulfate, delivered the nanoparticles to hepatic stellate cells. This strategy holds promise for degrading extracellular stroma and thereby facilitating drug penetration into fibrotic liver and related diseases such as liver cirrhosis and liver cancer.

Topics: Animals; Capsules; Cell Line; Chondroitin Sulfates; Collagenases; Disease Models, Animal; Hepatic Stellate Cells; Humans; Hyaluronan Receptors; Liver; Liver Cirrhosis; Mice; Nanoparticles; Silybin

Development of scaffold from biopolymers can ease the requirements for donor skin autograft and plays an effective role in the treatment of burn wounds. In the current study, a porous foam based, bilayered hydrogel scaffold was developed using gelatin, hyaluronic acid and chondroitin sulfate (G-HA-CS). The fabricated scaffold was characterized physicochemically for pre- and post-sterilization efficacy by scanning electron microscopy (SEM), Fourier transform infrared (FTIR) spectroscopy, differential scanning calorimetry (DSC) and thermal gravimetric analysis (TGA).

Topics: Animals; Biocompatible Materials; Burns; Chondroitin Sulfates; Disease Models, Animal; Gamma Rays; Gelatin; Hyaluronic Acid; Hydrogels; Male; Porosity; Swine; Tissue Scaffolds; Wound Healing

The role of glycosaminoglycan sulfation patterns, particularly in regard to scar formation and inhibition of neuritogenesis, has been mainly studied in cell culture with a focus on chondroitin 4-sulfate. In this study, we investigated chondroitin 6-sulfate (C6S) and found that it also inhibits neurite outgrowth of mouse cerebellar granule neurons in vitro. To examine whether the inhibitory activity of C6S could be neutralized, seven previously characterized high-affinity C6S-binding peptides were tested, among which three peptides neutralized the inhibitory functions of C6S. We further investigated these peptides in a mouse model of spinal cord injury, since upregulation of C6S expression in the glial scar following injury has been associated with reduced axonal regrowth and functional recovery. We here subjected mice to severe compression injury at thoracic levels T7-T9, immediately followed by inserting gelfoam patches soaked in C6S-binding peptides or in a control peptide. Application of C6S-binding peptides led to functional recovery after injury and prevented fibrotic glial scar formation, as seen by decreased activation of astrocytes and microglia/macrophages. Decreased expression of several lecticans and deposition of fibronectin at the site of injury were also observed. Application of C6S-binding peptides led to axonal regrowth and inhibited the C6S-mediated activation of RhoA/ROCK and decrease of PI3K-Akt-mTOR signaling pathways. Taken together, these results indicate that treatment with C6S-binding peptides improves functional recovery in a mouse model of spinal cord injury.

Topics: Animals; Axons; Cells, Cultured; Chondroitin Sulfate Proteoglycans; Chondroitin Sulfates; Cicatrix; Disease Models, Animal; Gliosis; Glycogen Synthase Kinase 3 beta; Locomotion; Macrophages; Mice, Inbred C57BL; Microglia; Neuronal Outgrowth; Neurons; Peptides; Phosphatidylinositol 3-Kinases; Proto-Oncogene Proteins c-akt; Recovery of Function; Remyelination; rho-Associated Kinases; rhoA GTP-Binding Protein; Spinal Cord Injuries; TOR Serine-Threonine Kinases

Examination of intervertebral disc (IVD) regeneration in an ovine annular lesion model.. Sulfation motifs are important functional determinants in glycosaminoglycans (GAGs). Previous studies have correlated 3-B-3(-) and 7-D-4 chondroitin sulfate (CS) motifs in tissues undergoing morphogenetic transition in development. We hypothesize that these motifs may also be expressed in degenerate IVDs and may represent a reparative response.. Induction of disc degeneration by 5 mm or 6 × 20 mm lesions in the annulus fibrosus (AF) over 6 or 3 to 6 months postoperation (PO). Tissue sections were stained with toluidine blue-fast green, 3-B-3(-) and 7-D-4 CS-sulfation motifs were immunolocalized in 3-month PO 6 × 20 mm lesion IVDs. Sulfated glycosaminoglycan (GAG), 3-B-3(-), and 7-D-4 epitopes were quantitated by ELISIA (enzyme-linked immunosorbent inhibition assay) in extracts of AF (lesion site and contralateral half) and nucleus pulposus (NP) 0, 3, and 6 months PO.. Collagenous overgrowth of lesions occurred in the outer AF. Chondroid metaplasia in ~20% of the 6 × 20 mm affected discs resulted in integration of an outgrowth of NP tissue with the inner AF lamellae preventing propagation of the lesion. 3-B-3(-) and 7-D-4 CS sulfation motifs were immunolocalized in this chondroid tissue. ELISIA quantified CS sulfation motifs demonstrating an increase 3 to 6 months PO in the AF lesion and a reduction in sulfated GAG not evident in the contralateral AF.. (1) Outer annular lesions underwent spontaneous repair. (2) Chondroid metaplasia of the inner 6 × 20 mm defect prevented its propagation suggesting an apparent reparative response.

Topics: Animals; Annulus Fibrosus; Chondroitin Sulfates; Disease Models, Animal; Enzyme-Linked Immunosorbent Assay; Epitopes; Glycosaminoglycans; Intervertebral Disc; Intervertebral Disc Degeneration; Regeneration; Sheep

Glaucoma is one of the most frequent causes of visual impairment worldwide, and involves selective damage to retinal ganglion cells (RGCs) and their axons. We analyzed the effect of enriched environment (EE) housing on the optic nerve, and retinal alterations in an induced model of ocular hypertension. For this purpose, male Wistar rats were weekly injected with vehicle or chondroitin sulfate (CS) into the eye anterior chamber for 10 weeks and housed in standard environment or EE. EE housing prevented the effect of experimental glaucoma on visual evoked potentials, retinal anterograde transport, phosphorylated neurofilament-immunoreactivity, axon number, microglial/macrophage reactivity (ionized calcium binding adaptor molecule 1-immunoreactivity), and astrocytosis (glial fibrillary acidic protein-immunostaining), as well as oligodendrocytes alterations (luxol fast blue staining, and myelin basic protein-immunoreactivity) in the proximal portion of the optic nerve. Moreover EE prevented the increase in ionized calcium binding adaptor molecule-1 levels, and RGC loss (Brn3a-immunoreactivity) in the retina from hypertensive eyes. EE increased retinal brain-derived neurotrophic factor levels. When EE housing started after 6 weeks of ocular hypertension, a preservation of visual evoked potentials amplitude, axon, and Brn3a(+) RGC number was observed. Taken together, these results suggest that EE preserved visual functions, reduced optic nerve axoglial alterations, and protected RGCs against glaucomatous damage.

Topics: Animals; Cholera Toxin; Chondroitin Sulfates; Disease Models, Animal; Environment; Evoked Potentials, Visual; Glaucoma; Housing, Animal; Male; Neuroprotection; Ocular Hypertension; Optic Nerve; Rats; Rats, Wistar; Retinal Ganglion Cells; Vision Disorders

Previous studies have suggested that thrombospondin-1 (TSP-1) regulates the transforming growth factor beta 1 (TGF-β1)/phosphorylated Smad2/3 (pSmad2/3) pathway. Moreover, TSP-1 is closely associated with epilepsy. However, the role of the TSP-1-regulated TGF-β1/pSmad2/3 pathway in seizures remains unclear. In this study, changes in this pathway were assessed following kainic acid (KA)-induced status epilepticus (SE) in rats. The results showed that increases in the TSP-1/TGF-β1/pSmad2/3 levels spatially and temporally matched the increases in glial fibrillary acidic protein (GFAP)/chondroitin sulfate (CS56) levels following KA administration. Inhibition of TSP-1 expression by small interfering RNA or inhibition of TGF-β1 activation with a Leu-Ser-Lys-Leu peptide significantly reduced the severity of KA-induced acute seizures. These anti-seizure effects were accompanied by decreased GFAP/CS56 expression and Smad2/3 phosphorylation. Moreover, inhibiting Smad2/3 phosphorylation with ponatinib or SIS3 also significantly reduced seizure severity, alongside reducing GFAP/CS56 immunoreactivity. These results suggest that the TSP-1-regulated TGF-β1/pSmad2/3 pathway plays a key role in KA-induced SE and astrogliosis, and that inhibiting this pathway may be a potential anti-seizure strategy.

Topics: Animals; Chondroitin Sulfates; Disease Models, Animal; Excitatory Amino Acid Agonists; Glial Fibrillary Acidic Protein; Imidazoles; Isoquinolines; Kainic Acid; Male; Protein Kinase Inhibitors; Pyridazines; Pyridines; Pyrroles; Rats; Rats, Sprague-Dawley; Signal Transduction; Smad2 Protein; Smad3 Protein; Status Epilepticus; Thrombospondin 1; Transforming Growth Factor beta1

Osteoarthritis is a degenerative disease that causes substantial changes in joint tissues, such as cartilage degeneration and subchondral bone sclerosis. Chondroitin sulfate and glucosamine are commonly used products for the symptomatic treatment of osteoarthritis. The aim of the present study was to investigate the effects of these products when used as structure-modifying drugs on the progression of osteoarthritis in the rabbit temporomandibular joint. Thirty-six New Zealand rabbits were divided into 3 groups (n = 12/group): control (no disease); osteoarthritis (disease induction); and treatment (disease induction and administration of chondroitin sulfate and glucosamine). Osteoarthritis was induced by intra-articular injection of monosodium iodoacetate. Animals were killed at 30 and 90 days after initiation of therapy. The treatment was effective in reducing disease severity, with late effects and changes in the concentration of glycosaminoglycans in the articular disc. The results indicate that chondroitin sulfate and glucosamine may have a structure-modifying effect on the tissues of rabbit temporomandibular joints altered by osteoarthritis.

Topics: Animals; Arthritis, Experimental; Cartilage, Articular; Chondroitin Sulfates; Disease Models, Animal; Drug Therapy, Combination; Extracellular Matrix; Glucosamine; Humans; Injections, Intra-Articular; Injections, Subcutaneous; Iodoacetic Acid; Male; Osteoarthritis; Rabbits; Severity of Illness Index; Temporomandibular Joint

Although dual endothelin receptor antagonists (ERAs) show great promise for treating various conditions, their propensity to induce liver injury limits their clinical usage. Inflammation and fibrosis are important processes in liver responses to injury and it has been suggested that they and dual ERA-induced liver injury are mediated by the proteoglycan component chondroitin sulfate (CS), which is synthesized by

Topics: Animals; Carbohydrate Sulfotransferases; Cell Line, Tumor; Chemical and Drug Induced Liver Injury; Chondroitin Sulfates; Cytokines; Disease Models, Animal; Endothelin Receptor Antagonists; Fibrosis; Glycosaminoglycans; Humans; Liver; Mice; Receptor, Endothelin A; Receptor, Endothelin B; Sulfotransferases

Mucopolysaccharidoses are a class of lysosomal storage diseases, characterized by enzymatic deficiency in the degradation of specific glycosaminoglycans (GAG). Pathological accumulation of excess GAG leads to multiple clinical symptoms with systemic character, most severely affecting bones, muscles and connective tissues. Current therapies include periodic intravenous infusion of supplementary recombinant enzyme (Enzyme Replacement Therapy-ERT) or bone marrow transplantation. However, ERT has limited efficacy due to poor penetration in some organs and tissues. Here, we investigated the potential of the β-D-xyloside derivative odiparcil as an oral GAG clearance therapy for Maroteaux-Lamy syndrome (Mucopolysaccharidosis type VI, MPS VI). In vitro, in bovine aortic endothelial cells, odiparcil stimulated the secretion of sulphated GAG into culture media, mainly of chondroitin sulphate (CS) /dermatan sulphate (DS) type. Efficacy of odiparcil in reducing intracellular GAG content was investigated in skin fibroblasts from MPS VI patients where odiparcil was shown to reduce efficiently the accumulation of intracellular CS with an EC50 in the range of 1 μM. In vivo, in wild type rats, after oral administrations, odiparcil was well distributed, achieving μM concentrations in MPS VI disease-relevant tissues and organs (bone, cartilage, heart and cornea). In MPS VI Arylsulphatase B deficient mice (Arsb-), after chronic oral administration, odiparcil consistently stimulated the urinary excretion of sulphated GAG throughout the treatment period and significantly reduced tissue GAG accumulation in liver and kidney. Furthermore, odiparcil diminished the pathological cartilage thickening observed in trachea and femoral growth plates of MPS VI mice. The therapeutic efficacy of odiparcil was similar in models of early (treatment starting in juvenile, 4 weeks old mice) or established disease (treatment starting in adult, 3 months old mice). Our data demonstrate that odiparcil effectively diverts the synthesis of cellular glycosaminoglycans into secreted soluble species and this effect can be used for reducing cellular and tissue GAG accumulation in MPS VI models. Therefore, our data reveal the potential of odiparcil as an oral GAG clearance therapy for MPS VI patients.

Topics: Administration, Oral; Animals; Cattle; Cells, Cultured; Chondroitin Sulfates; Dermatan Sulfate; Disease Models, Animal; Endothelial Cells; Female; Glycosaminoglycans; Glycosides; Humans; In Vitro Techniques; Male; Mice; Mice, Inbred C57BL; Mice, Mutant Strains; Mucopolysaccharidosis VI; Rats; Rats, Sprague-Dawley

Sea cucumber body wall contains several naturally occurring bioactive components that possess health-promoting properties.

Topics: Animals; Anti-Inflammatory Agents; Chondroitin Sulfates; Colitis; Dextran Sulfate; Disease Models, Animal; Glycosaminoglycans; HaCaT Cells; Humans; In Vitro Techniques; Mexico; Mice; Otitis; Sea Cucumbers; Tetradecanoylphorbol Acetate; Tissue Extracts

Oligo-peptides, including monomeric amino acids, have received much attention as bioactive molecules and drugs. One of the biggest problems of these compounds, however, is their very short bioavailability due to instant metabolism and rapid excretion. To solve this problem, we newly designed a poly(ethylene glycol) (PEG)-block-polypeptide self-assembling based drug for the treatment of acute liver injury. Here, PEG-block-poly(L-Ornithine) (PEG-b-POrn) was synthesized via a ring opening polymerization, and a nano-sized polyion self-assembling complex (Nano

Topics: Acetaminophen; Ammonia; Animals; Cattle; Cell Survival; Chemical and Drug Induced Liver Injury; Chondroitin Sulfates; Disease Models, Animal; Drug Delivery Systems; Endothelial Cells; Hyperammonemia; Macrophages; Male; Mice; Mice, Inbred C57BL; Mice, Inbred ICR; Nanoparticles; Ornithine; RAW 264.7 Cells

Blood cytokines affect the development of inflammatory processes in both normal and pathological states. We have studied changes in the concentration of interleukins (ILs) - 1β, IL-4, IL-10, IL-12B p40, transforming growth factor β (TGF β), tumor necrosis factor (TNF-α) in acute carrageenan-induced inflammation and degenerative-dystrophic changes of knee joint caused by monoiodoacetate-induced Osteoarthritis (OA) in experimental models on rats. We also investigated the change in the cytokine profile during prophylactic and therapeutic administration of chondroitin sulfate to animals under experimental conditions.. The concentration of the cytokines was measured in blood serum by enzyme-linked immunosorbent assay.. The manifestation of articular lesions was characterized by a disturbance in the balance between proinflammatory (IL-1β, IL-12B p40, TNF-α) and anti-inflammatory (IL-4, IL-10, TGF -β) cytokines.. A reduction in the concentration of proinflammatory cytokines in blood serum after prophylactic and therapeutic administration of chondroitin sulfate to the rat with experimental models of acute inflammation of the hind limb and degenerative-dystrophic changes in the knee joint with OA is associated with anti-inflammatory and regenerative properties of the drug.

Topics: Animals; Arthritis, Experimental; Carrageenan; Chondroitin Sulfates; Cytokines; Disease Models, Animal; Edema; Iodoacetic Acid; Male; Osteoarthritis; Random Allocation; Rats; Rats, Wistar; Sensitivity and Specificity

The two related tumor necrosis factor members a proliferation-inducing ligand (APRIL) and B-cell activation factor (BAFF) are currently targeted in autoimmune diseases as B-cell regulators. In multiple sclerosis (MS), combined APRIL/BAFF blockade led to unexpected exacerbated inflammation in the central nervous system (CNS) of patients. Here, we investigate the role of the APRIL/BAFF axis in the CNS.. APRIL expression was analyzed in MS lesions by immunohistochemistry. The in vivo role of APRIL was assessed in the murine MS model, experimental autoimmune encephalitis (EAE). Functional in vitro studies were performed with human and mouse astrocytes.. APRIL was expressed in lesions from EAE. In its absence, the disease was worst. Lesions from MS patients also showed APRIL expression upon infiltration of macrophages. Notably, all the APRIL secreted by these macrophages specifically targeted astrocytes. The upregulation of chondroitin sulfate proteoglycan, sometimes bearing chondroitin sulfate of type E sugar moieties, binding APRIL, in reactive astrocytes explained the latter selectivity. Astrocytes responded to APRIL by producing a sufficient amount of IL-10 to dampen antigen-specific T-cell proliferation and pathogenic cytokine secretion. Finally, an intraspinal delivery of recombinant APRIL before disease onset, shortly reduced EAE symptoms. Repeated intravenous injections of recombinant APRIL before and even at disease onset also had an effect.. Our data show that APRIL mediates an anti-inflammatory response from astrocytes in MS lesions. This protective activity is not shared with BAFF. ANN NEUROL 2019;85:406-420.

Topics: Adult; Aged; Animals; Astrocytes; B-Cell Activating Factor; Cell Proliferation; Chondroitin Sulfate Proteoglycans; Chondroitin Sulfates; Cytokines; Disease Models, Animal; Encephalomyelitis, Autoimmune, Experimental; Female; Humans; Immunohistochemistry; Interleukin-10; Macrophages; Male; Mice; Mice, Knockout; Middle Aged; Multiple Sclerosis; Reverse Transcriptase Polymerase Chain Reaction; T-Lymphocytes; Tumor Necrosis Factor Ligand Superfamily Member 13

Familial members of urolithiasis have high risk for stone development. We observed the low sulfated glycosaminoglycan (GAG) excretion in urolithiasis patients and their descendants. In this study, we investigated urinary excretion of sulfated GAG, chondroitin sulfate (CS), heparan sulfate (HS) and hyaluronic acid (HA) in urolithiasis and their children, and explored the effect of CS and HA supplement in urolithic hyperoxaluric rats. The 24-hour urines were collected from urolithiasis patients (28) and their children (40), as well as healthy controls (45) and their children (33) to measure urinary sulfated GAG, CS, HS and HA excretion rate. Our result showed that urinary sulfated GAG and CS were diminished in both urolithiasis patients and their children, while decreased HS and increased HA were observed only in urolithiasis patients. Percentage of HS per sulfated GAG increased in both urolithiasis patients and their children. In hyperoxaluric rats induced by ethylene glycol and vitamin D, we found that CS supplement could prevent stone formation, while HA supplement had no effect on stone formation. Our study revealed that decreased urinary GAG and CS excretion are common in familial members of urolithiasis patients, and CS supplement might be beneficial in calcium oxalate urolithiasis prophylaxis for hyperoxaluric patients.

Topics: Adult; Animals; Child; Chondroitin Sulfates; Creatinine; Dietary Supplements; Disease Models, Animal; Female; Glycosaminoglycans; Heparitin Sulfate; Humans; Hyaluronic Acid; Kidney; Male; Middle Aged; Rats; Rats, Wistar; Urolithiasis

Topics: Animals; Apoptosis; Caspases; Cell Survival; Chondrocytes; Chondroitin Sulfates; Disease Models, Animal; DNA Fragmentation; Humans; Hydrogen Peroxide; Mitochondria; Mitochondrial Dynamics; Osteoarthritis; Rats

The aim of this study was to compare the effects of combined treatment with chondroitin sulfate and glucosamine sulfate (CS/Gl) and photobiomodulation (PBM) on the degenerative process related to osteoarthritis (OA) in the articular cartilage in rats. Forty male Wistar rats were randomly divided into four groups: OA control group (CG); OA animals submitted to PBM treatment (PBM); OA animals submitted to CS/Gl treatment (CS/Gl); OA submitted to CS/GS associated with PBM treatments (GS/Gl + PBM). The CS/Gl started 48 h after the surgery, and they were performed for 29 consecutive days. Moreover, PBM was performed after the CS/Gl administration on the left joint. Morphological characteristics and immunoexpression of interleukin 10 (IL-10) and 1 beta (IL-1β) and collagen type II (Col II) of the articular cartilage were evaluated. The results showed that all treated groups (CS/Gl and PBM) presented attenuation signs of degenerative process (measured by histopathological analysis) and lower density chondrocytes [PBM (p = 0.0017); CS/Gl (p = 0.0153) and CS/Gl + PBM (p = 0.002)]. Additionally, CS/Gl [associated (p = 0.0089) or not with PBM (p = 0.0059)] showed significative lower values for OARSI grade evaluation. Furthermore, CS/GS + PBM decreased IL-1β protein expression (p = 0.0359) and increased IL-10 (p = 0.028) and Col II imunoexpression (p = 0.0204) compared to CG. This study showed that CS/Gl associated with PBM was effective in modulating inflammatory process and preventing the articular tissue degradation in the knees OA rats.

Topics: Animals; Chondrocytes; Chondroitin Sulfates; Collagen Type II; Combined Modality Therapy; Disease Models, Animal; Glucosamine; Immunohistochemistry; Interleukin-10; Interleukin-1beta; Low-Level Light Therapy; Male; Osteoarthritis; Rats, Wistar

Extracellular histones induce lethal thrombosis by promoting platelet aggregation, neutrophil migration, and cell injuries. Heparin, which has negative charges, can bind to extracellular histones; however, heparin strongly inhibits the activation of coagulation. Since chondroitin sulfate (CS) shows less effect on the coagulation system than heparin does, CS has the potential to become an effective drug for lethal thrombosis with high risk of bleeding. To elucidate the therapeutic mechanisms of CS in lethal thrombosis, we investigated the interaction between CS and extracellular histones. Mouse vascular endothelial cells were incubated with histones in the presence of heparin or CS, and the expression of caspase-3/7 was measured. The interactions between histones and heparin or CS were measured by surface plasmon resonance analysis. Vascular permeability, platelet counts, liver and renal functions, and coagulation times were evaluated in an in vivo assay. The apoptosis induced by histones was inhibited by treatment with heparin or CS. Heparin and CS showed strong binding to histones and inhibited vascular hyperpermeability. The platelet counts as well as liver and renal functions were not decreased by the treatment with heparin or CS. Moreover, CS showed less effect on the coagulation system than heparin did. These results suggested that CS can be a novel agent for lethal thrombosis with the risk for hemorrhage. Since vascular endothelial cell injuries occur at an early stage of lethal thrombosis, administration of CS might be a useful approach.

Topics: Animals; Anticoagulants; Blood Coagulation; Capillary Permeability; Chondroitin Sulfates; Disease Models, Animal; Endothelial Cells; Hemorrhage; Heparin; Histones; Humans; Male; Mice; Mice, Inbred DBA; Platelet Aggregation; Platelet Function Tests; Surface Plasmon Resonance; Thrombosis; Treatment Outcome

Porcine glutaraldehyde-fixed pericardium is widely used to replace human heart valves. Despite the stabilizing effects of glutaraldehyde fixation, the lack of endothelialization and the occurrence of immune reactions contribute to calcification and structural valve deterioration, which is particularly significant in young patients, in whom valve longevity is crucial. This report shows an optimization system with which to enhance endothelialization of fixed pericardium to mimic the biological function of a native heart valve. The glutaraldehyde detoxification, together with the application of a biodegradable methacrylated chondroitin sulfate hydrogel, reduces aldehydes cytotoxicity, increases the migration and proliferation of endothelial cells and the recruitment of endothelial cell progenitors, and confers thromboresistance in fixed pericardium. The combination of glutaraldehyde detoxification and a coating with chondroitin sulfate hydrogel promotes in situ mechanisms of endothelialization in fixed pericardium. We offer a new solution for improving the long life of bioprosthetic valves and exploring the means of making valves suitable to endothelialization.

Topics: Animals; Cell Movement; Cell Proliferation; Chondroitin Sulfates; Clinical Deterioration; Disease Models, Animal; Endothelial Cells; Glutaral; Heart Valves; Humans; Hydrogel, Polyethylene Glycol Dimethacrylate; Pericardium; Swine

Chondroitin sulfate (CS) is an important component of the extracellular matrix of cartilage and has been widely used as one of the main drugs for the treatment of joint pain-related nutraceuticals and medicines. Sturgeon bone (SB) is the main waste during deep processing of sturgeons in fishery production. The present study was evaluated the therapeutic effect of CS from SB (CSSB) on monosodium iodoacetate (MIA)-induced osteoarthritis (OA) pain and further explored the potential medicinal value of CSSB. The OA pain model was induced by intra-articular injection of MIA and then treated with CSSB. The results showed that, on the organismic level, CSSB can significantly reduce the joint swelling, reduce the pathological injury of the joints, decrease the levels of IL-1, TNF-α and PGE

Topics: Analgesics; Animals; Arthralgia; Arthritis, Experimental; Biological Products; Biomarkers; Chondroitin Sulfates; Cytokines; Disease Models, Animal; Inflammation Mediators; Iodoacetates; Male; Osteoarthritis; Rats

Chondroitin sulphate (CS) has long been used to treat osteoarthritis. Some investigations have also shown that the treatment with CS could reduce coronary events in patients with heart disease but no studies have identified the mechanistic role of these therapeutic effects. We aimed to investigate how the treatment with CS can interfere with the progress of atherosclerosis. The aortic arch, thoracic aorta and serum were obtained from apolipoprotein E (ApoE) knockout mice fed for 10 weeks with high-fat diet and then treated with CS (300 mg/kg,

Topics: Animals; Anti-Inflammatory Agents; Aorta, Thoracic; Aortic Diseases; Atherosclerosis; Blood Glucose; C-Reactive Protein; Chondroitin Sulfates; Diet, High-Fat; Disease Models, Animal; Endothelial Cells; Foam Cells; Humans; Inflammation; Inflammation Mediators; Lipids; Lipoproteins, LDL; Male; Mice, Knockout, ApoE; Monocytes; Plaque, Atherosclerotic; THP-1 Cells

Fucosylated chondroitin sulfate (fCS) and its depolymerized derivative (DfCS), prepared from sea cucumbers, are well-known for their anticoagulant activity. However, their other functional activities are poorly understood. Recently, we obtained fCS oligosaccharides from Isostichopus Badionotus by a modified controllable Fenton-system, named as DfCS-Ib. The functional activities of these oligosaccharides are still unclear. The present study investigated anti-hyperlipidemic activity of DfCS-Ib using a high-fat diet (HFD)-fed mice model. The results indicated that DfCS-Ib reduced obesity, hyperlipidemia, and inflammation caused by HFD. Meanwhile, DfCS-Ib increased the mRNA expression of PPARγ and decreased the mRNA expression of leptin, aP2, and F4/80 in fat tissue. Transcriptome analysis indicated that DfCS-Ib normalized the expressions of genes regulating lipid metabolism. Our results suggested that DfCS-Ib can alleviated lipid disorder by reducing lipid synthesis and promoting lipid lipidolysis. DfCS-Ib can act as a functional agent to regulate lipid disorder.

Topics: Animals; Chondroitin Sulfates; Dietary Fats; Disease Models, Animal; Hyperlipidemias; Male; Mice; Obesity; Oligosaccharides; Sea Cucumbers

Fucosylated chondroitin sulfate

Topics: Animals; Anti-Inflammatory Agents; Cartilage; Chondroitin Sulfates; Circular Dichroism; Cucumaria; Disease Models, Animal; Female; Humans; Magnetic Resonance Spectroscopy; Molecular Structure; Peptones; Peritonitis; Rats; Rats, Wistar; Salmo salar; Structure-Activity Relationship; Treatment Outcome

The present study is aimed at testing the antidepressant--like effects and probable mechanisms of action of low molecular mass chondroitin sulfate (LMMCS) on depression induced by chronic unpredictable mild stress (CUMS) in mice. Four weeks of CUMS exposure resulted in depressive-like behavior, expressed by a significant decrease in the locomotor activity and sucrose consumption and increased immobility time in the forced swim test. Further, there was a significant reduction of 5-HT level in the hippocampus region of depressed mice. Treatment of mice for four weeks with LMMCS ameliorated significantly both the behavioral and biochemical changes induced by CUMS. These novel results suggest that LMMCS produces an antidepressant-like effect in mice subjected to CUMS, which might be related, at least in part, to the increase of 5-HT concentration in the hippocampus.

Topics: Animals; Antidepressive Agents; Behavior, Animal; Chondroitin Sulfates; Depression; Disease Models, Animal; Hippocampus; Locomotion; Male; Mice; Molecular Weight; Serotonin; Stress, Psychological; Sucrose; Swimming

To evaluate the crosslinking effect of functionalized chondroitin sulfate (CS) in an ex vivo rabbit cornea model.. Chondroitin sulfate molecules were chemically modified with the N-hydroxysuccinimide (NHS) group. Enucleated rabbit eyes were crosslinked with 2, 5, or 10 mg/mL CS-NHS solution for 30 or 60 minutes. The CS-NHS penetration, corneal swelling ratio, Young's modulus, and ultrastructure of the crosslinked corneas were characterized. In addition, rabbit corneas were further treated with a collagenase-chondroitinase solution to create an ex vivo keratoconus (KC)-like model. The KC model corneas were crosslinked with a standard riboflavin-ultraviolet (UV) method or alternatively with CS-NHS. Corneal mechanics, ultrastructure, and keratocyte gene expression were evaluated after UV and CS-NHS crosslinking.. CS-NHS effectively penetrated into the corneal stroma within 60 minutes of treatment initiation. CS-NHS crosslinking reduced the swelling ratio by 35%, increased Young's modulus by 20%, and increased collagen fibril diameter and density. CS-NHS crosslinking improved corneal mechanics of KC model corneas to levels comparable to those with UV crosslinking. Moreover, CS-NHS crosslinking demonstrated significant downregulation of proinflammatory gene expression of keratocytes, indicating a potential protective effect imparted by CS-NHS during crosslinking.. Our results demonstrated that CS-NHS can reinforce normal and KC model corneal mechanics, and restore collagen density and alignment in KC model corneas without causing extensive keratocyte apoptosis and proinflammatory gene upregulation. Therefore, CS-NHS crosslinking can potentially provide an effective, safe, and biocompatible means of corneal reinforcement.

Topics: Animals; Biomechanical Phenomena; Chondroitin Sulfates; Collagen; Cornea; Corneal Keratocytes; Cross-Linking Reagents; Disease Models, Animal; Elastic Modulus; Photosensitizing Agents; Rabbits; Ultraviolet Rays

Osteoarthritis is the most common chronic joint disorder especially during aging. Although with controversies, glucosamine, both in its forms of sulfate and hydrochloride, and chondroitin sulfate are commonly employed to treat osteoarthritis. Due to the modest improve in the symptoms observed in patients treated with these drugs alone, a formulation combining both agents has been considered. The discrepant results achieved for pain control or structural improvement in osteoarthritis patients has been attributed to the quality of chemical formulations or different bias in clinical studies. The current study has been designed to test the effects of two different combined formulations with adequate pharmaceutical grade of these drugs in osteoarthritic joints, and to explore the underlying mechanisms modulated by both formulations in different osteoarthritis target tissues. Knee osteoarthritis was surgically induced in experimental rabbits. Some animals received the combined therapy (CT)1, (chondroitin sulfate 1200mg/day + glucosamine sulfate 1500mg/day), or the CT2 ((chondroitin sulfate 1200mg/day + glucosamine hydrochloride 1500mg/day). Neither CT1 nor CT2 significantly modified the cartilage damage or the synovial inflammation observed in osteoarthritic animals. Treatments were also unable to modify the presence of pro-inflammatory mediators, and the synthesis of metalloproteinases in the cartilage or in the synovium of osteoarthritic animals. Combined therapies did not modify the decrease in the subchondral bone mineral density observed in osteoarthritic rabbits. Therapies of chondroitin sulfate plus glucosamine sulfate or chondroitin sulfate plus glucosamine hydrochloride failed to improve structural damage or to ameliorate the inflammatory profile of joint tissues during experimental osteoarthritis.

Topics: Animals; Bone Density; Cartilage, Articular; Chondroitin Sulfates; Cytokines; Disease Models, Animal; Drug Interactions; Glucosamine; Knee Joint; Male; Matrix Metalloproteinases; Osteoarthritis, Knee; Osteoprotegerin; Rabbits; RANK Ligand; Synovial Membrane

Fucosylated chondroitin sulfate (FucCS) is a potent anticoagulant polysaccharide extracted from sea cucumber. Its anticoagulant activity is attributed to the presence of unique branches of sulfated fucose. Although this glycosaminoglycan exerts an antithrombotic effect following oral administration, high doses are necessary to achieve the maximum effect. The diminished activity of FucCS following oral administration is likely due to its degradation in the gastrointestinal tract and its limited ability to cross the intestinal cell membranes. The latter aspect is particularly difficult to overcome. However, gastro-resistant tablet formulation may help limit the degradation of FucCS in the gastrointestinal tract. In the present work, we found that the oral administration of FucCS as gastro-resistant tablets produces a more potent and prolonged anticoagulant effect compared with its administration as an aqueous solution, with no significant changes in the bleeding tendency or arterial blood pressure. Experiments using animal models of arterial thrombosis initiated by endothelial injury demonstrated that FucCS delivered as gastro-protective tablets produced a potent antithrombotic effect, whereas its aqueous solution was ineffective. However, there was no significant difference between the effects of FucCS delivered as gastro-resistant tablets or as aqueous solution in a venous thrombosis model, likely due to the high dose of thromboplastin used. New oral anticoagulants tested in these experimental models for comparison showed significantly increased bleeding tendencies. Our study provides a framework for developing effective oral anticoagulants based on sulfated polysaccharides from marine organisms. The present results suggest that FucCS is a promising oral anticoagulant.

Topics: Administration, Oral; Animals; Anticoagulants; Blood Coagulation; Carotid Artery Diseases; Chondroitin Sulfates; Disease Models, Animal; Drug Compounding; Female; Hemorrhage; Male; Rats, Wistar; Tablets, Enteric-Coated; Thrombosis; Time Factors; Venous Thrombosis

Although tendon-exposed or bone-exposed wounds can be resurfaced with flaps, such surgeries may not be feasible in patients with poor general or local conditions. Biosynthetic artificial skin is an alternative for critical wound coverage. We designed a new artificial skin bilayer to close difficult wounds permanently.. This study compares incorporation and wound contraction between silicone acellular porcine dermis (SAPD) and the Integra graft (Integra Life Sciences Corp., Billerica, Mass) in a rat model.. The SAPD was manufactured according to our previously described standard procedures. Integra grafts were obtained commercially. We included 24 male adult Sprague-Dawley rats and divided them into 2 groups. After creating a 3 × 4-cm full-thickness wound on the back, we transplanted the same-sized SAPD and Integra grafts onto the rat wounds. Autologous full-thickness skin (FTS) was grafted onto the acellular porcine dermal matrix (APDM) of the SAPD and the Integra dermal matrix (IDM) 2 weeks later. We measured the wound size and contraction rate of recipient wounds, studied the incorporation of FTS on the dermal matrix, and did pathological examination. Generalized estimating equations were used to assess the data from repeated wound and scar contraction measurements using SAS v9.2.. The sizes of wounds of both groups decreased over time. No difference in wound contraction was observed between the SAPD and Integra groups at weeks 2, 4, or 6 after grafting. However, the contraction rates in both groups increased significantly. The pathological examination showed that the FTS was well incorporated in the APDM and IDM. The recipient wounds showed new vessels and cell infiltration in the new matrix, but no severe inflammation. Skin appendages were regenerating in the FTS. There was no rejection sign.. Both SAPD and Integra are double-layered artificial skin products. Our results demonstrate that APDM and IDM are good templates and show excellent incorporation with autologous FTS graft. The results also demonstrated gradual wound contraction over time, but the contraction rate was not different between SAPD and Integra 6 weeks after grafting in a rat model.

Topics: Acellular Dermis; Animals; Chondroitin Sulfates; Cicatrix; Collagen; Contracture; Disease Models, Animal; Male; Rats; Rats, Sprague-Dawley; Skin Transplantation; Swine; Transplantation, Autologous; Wound Closure Techniques; Wound Healing

Chondroitin/dermatan sulfate proteoglycans (CSPGs/DSPGs) are major components of the extracellular matrix. Their expression is generally upregulated after injuries to the adult mammalian central nervous system, which is known for its low ability to restore function after injury. Several studies support the view that CSPGs inhibit regeneration after injury, whereas the functions of DSPGs in injury paradigms are less certain. To characterize the functions of DSPGs in the presence of CSPGs, we studied young adult dermatan-4O-sulfotransferase1-deficient (Chst14(-/-)) mice, which express chondroitin sulfates (CSs), but not dermatan sulfates (DSs), to characterize the functional outcome after severe compression injury of the spinal cord. In comparison to their wild-type (Chst14(+/+)) littermates, regeneration was reduced in Chst14(-/-) mice. No differences between genotypes were seen in the size of spinal cords, numbers of microglia and astrocytes neither in intact nor injured spinal cords after injury. Monoaminergic innervation and re-innervation of the spinal cord caudal to the lesion site as well as expression levels of glial fibrillary acidic protein (GFAP) and myelin basic protein (MBP) were similar in both genotypes, independent of whether they were injured and examined 6weeks after injury or not injured. These results suggest that, in contrast to CSPGs, DSPGs, being the products of Chst14 enzymatic activity, promote regeneration after injury of the adult mouse central nervous system.

Topics: Animals; Behavior, Animal; Chondroitin Sulfates; Dermatan Sulfate; Disease Models, Animal; Mice; Motor Activity; Nerve Regeneration; Recovery of Function; Spinal Cord Injuries; Sulfotransferases

Osteoarthritic patients treated with high doses of chondroitin sulfate (CS) have a lower incidence of coronary heart disease--but the mechanistic aspects of these beneficial effects of CS remain undefined. We examined how CS treatment affects the formation of atheroma via interaction with endothelial cells and monocytes.. We characterized arterial atheromatous plaques by multiphoton microscopy and serum pro-inflammatory cytokines by immunoenzymatic techniques in obese mice receiving CS (1 g/kg/day, i.p.) or vehicle for 6 days. Effects of CS on signaling pathways, cytokine secretion and macrophage migration were evaluated in cultures of human coronary endothelial cells and in a monocyte cell line stimulated with TNF-α by Western blot, immunoenzymatic techniques and transwell migration assays.. Treatment of obese mice with CS reduced the extension of foam cell coverage in atheromatous plaques of arterial bifurcations by 62.5%, the serum concentration of IL1β by 70%, TNF-α by 82% and selected chemokines by 25-35%. Cultures of coronary endothelial cells and monocytes stimulated with TNF-α secreted less pro-inflammatory cytokines in the presence of CS (P < 0.01). CS reduced the activation of the TNF-α signaling pathway in endothelial cells (pErk 36% of reduction, and NFκB 33% of reduction), and the migration of activated monocytes to inflamed endothelial cells in transwells (81 ± 6 vs. 13 ± 2, P < 0.001).. CS interferes with the pro-inflammatory activation of monocytes and endothelial cells driven by TNF-α thus reducing the propagation of inflammation and preventing the formation of atherosclerotic plaques.

Topics: Animals; Atherosclerosis; Cell Line; Chondroitin Sulfates; Disease Models, Animal; Endothelium, Vascular; Inflammation; Male; Mice; Mice, Obese; Obesity

To evaluate the potential of a bioactive coating based on chondroitin sulfate (CS) and tethered epidermal growth factor (EGF) for improvement of healing around stent grafts (SGs).. The impact of the bioactive coating on cell survival was tested in vitro on human vascular cells using polyethylene terephthalate films (PET) as a substrate. After being transferred onto a more "realistic" material (expanded polytetrafluoroethylene [ePTFE]), the durability and mechanical behavior of the coating and cell survival were studied. Preliminary in vivo testing was performed in a canine iliac aneurysm model reproducing type I endoleaks (three animals with one control and one bioactive SG for each).. CS and EGF coatings significantly increased survival of human smooth muscle cells and fibroblasts compared with bare PET or ePTFE (P < .05). The coating also displayed good durability over 30 days according to enzyme-linked immunosorbent assay and cell survival tests. The coating did not affect mechanical properties of ePTFE and was successfully transferred onto commercial SGs for in vivo testing. No difference was observed on computed tomography and macroscopic examinations in endoleak persistence at 3 months, but the bioactive coating deposited on the abluminal surface of the SG (exposed to the vessel wall) increased the percentage of healed tissue in the aneurysm. No adverse effect, such as neointima formation or thrombosis, was observed.. The bioactive coating promoted in vitro cell survival, displayed good durability, and was successfully transferred onto a commercial SG. Preliminary in vivo results suggest improved healing around bioactive SGs.

Topics: Animals; Blood Vessel Prosthesis; Blood Vessel Prosthesis Implantation; Cell Adhesion; Cell Survival; Cells, Cultured; Chondroitin Sulfates; Coated Materials, Biocompatible; Computed Tomography Angiography; Disease Models, Animal; Dogs; Endoleak; Epidermal Growth Factor; Fibroblasts; Humans; Iliac Aneurysm; Iliac Artery; Materials Testing; Microscopy, Confocal; Myocytes, Smooth Muscle; Pilot Projects; Polyethylene Terephthalates; Polytetrafluoroethylene; Prosthesis Design; Stents; Time Factors; Wound Healing

The efficacy of the combination chondroitin sulfate-glucosamine (CS-GlcN) in the treatment of knee osteoarthritis (OA) has been suggested in recent clinical studies. In vitro reports have also suggested anti-inflammatory and anti-resorptive effects of this combination.. The aim of this study was to characterize the effects of CS-GlcN on joint degradation in vivo including the assessment of inflammation and bone metabolism in a model of OA.. We have used the OA model induced by anterior cruciate ligament transection (ACLT) in ovariectomised rats. CS-GlcN was administered daily (oral gavage) from week 0 until week 12 after ovariectomy at the dose of 140 (CS)+175 (GlcN)(HCl) mg/kg. Histochemical analyses were performed, the levels of biomarkers and inflammatory mediators were measured by luminex or ELISA and bone microstructure was determined by μCT.. CS-GlcN protected against cartilage degradation and reduced the levels of inflammatory mediators such as interleukin-1β and tumor necrosis factor-α in the affected knee. In addition, serum biomarkers of inflammation and cartilage and bone degradation including matrix metalloproteinase-3, C-telopeptide of type II collagen and the ratio receptor activator of nuclear factor κB ligand/osteoprotegerin were significantly decreased by CS-GlcN. This treatment also tended to improve some bone microstructural parameters without reaching statistical significance.. These results demonstrate the chondroprotective effects of CS-GlcN in vivo, in the experimental model of ACLT in ovariectomised rats, and suggest that this combination may be useful to control the joint catabolic effects of inflammatory stress. These findings could have clinical relevance related to the prevention of joint degradation by CS-GlcN and support the potential development of OA treatments based on this combination.

Topics: Animals; Anterior Cruciate Ligament; Anterior Cruciate Ligament Injuries; Biomarkers; Bone and Bones; Cartilage, Articular; Chondroitin Sulfates; Disease Models, Animal; Drug Therapy, Combination; Female; Glucosamine; Inflammation Mediators; Joints; Osteoarthritis, Knee; Ovariectomy; Protective Agents; Rats, Wistar; X-Ray Microtomography

Plasmodium falciparum infection during pregnancy leads to abortions, stillbirth, low birth weight, and maternal mortality. Infected erythrocytes (IEs) accumulate in the placenta by adhering to chondroitin sulfate A (CSA) via var2CSA protein exposed on the P. falciparum IE membrane. Plasmodium berghei IE infection in pregnant BALB/c mice is a model for severe placental malaria (PM). Here, we describe a transgenic P. berghei parasite expressing the full-length var2CSA extracellular region (domains DBL1X to DBL6ε) fused to a P. berghei exported protein (EMAP1) and characterize a var2CSA-based mouse model of PM. BALB/c mice were infected at midgestation with different doses of P. berghei-var2CSA (P. berghei-VAR) or P. berghei wild-type IEs. Infection with 10(4) P. berghei-VAR IEs induced a higher incidence of stillbirth and lower fetal weight than P. berghei At doses of 10(5) and 10(6) IEs, P. berghei-VAR-infected mice showed increased maternal mortality during pregnancy and fetal loss, respectively. Parasite loads in infected placentas were similar between parasite lines despite differences in maternal outcomes. Fetal weight loss normalized for parasitemia was higher in P. berghei-VAR-infected mice than in P. berghei-infected mice. In vitro assays showed that higher numbers of P. berghei-VAR IEs than P. berghei IEs adhered to placental tissue. Immunization of mice with P. berghei-VAR elicited IgG antibodies reactive to DBL1-6 recombinant protein, indicating that the topology of immunogenic epitopes is maintained between DBL1-6-EMAP1 on P. berghei-VAR and recombinant DBL1-6 (recDBL1-6). Our data suggested that impairments in pregnancy caused by P. berghei-VAR infection were attributable to var2CSA expression. This model provides a tool for preclinical evaluation of protection against PM induced by approaches that target var2CSA.

Topics: Animals; Antibodies, Protozoan; Antigens, Protozoan; Chondroitin Sulfates; Disease Models, Animal; Epitopes; Erythrocytes; Female; Fetal Weight; Immunization; Immunoglobulin G; Malaria; Malaria, Falciparum; Mice; Mice, Inbred BALB C; Parasite Load; Parasitemia; Placenta; Plasmodium berghei; Plasmodium falciparum; Pregnancy; Pregnancy Complications, Parasitic; Protein Domains; Recombinant Fusion Proteins; Stillbirth

Artificial dermal templates play an important role in physiologic wound closure after injury. In addition to contributing to stable, durable and flexible wound closure, they provide a scaffold for tissue repair. Several dermal templates are commercially available, with animal-derived Integra(®) dermal regeneration template perhaps the most widely used. NovoSorb™ Biodegradable Temporising Matrix (BTM) is a fully synthetic alternative that eliminates any risk of cross-species residual antigenicity. In this study, we aimed to compare early response after application of NovoSorb™ BTM with Integra(®) in terms of temporary wound closure, host cell infiltration, neovascularisation and collagen deposition in a mouse model.. Twenty athymic nude mice received full-thickness skin excision followed by grafting of the dermal template (n=10 NovoSorb™ BTM, n=10 Integra(®)), with the grafts excised and assessed after two weeks.. All twenty mice achieved temporary wound closure with no evidence of wound contracture. Microscopically, all twenty grafts became infiltrated with host cells along the entire length of the template, with NovoSorb™ BTM demonstrating a particular abundance of host inflammatory cells. Evidence of new collagen deposition and neovascularisation was observed in both templates, with NovoSorb™ BTM demonstrating a more extensive vascular network at this time point. However, a greater inflammatory response was also observed in the NovoSorb™ BTM grafts at this time point.. In this study, NovoSorb™ BTM demonstrates favourable properties as a dermal template, but further investigation is required to assess the significance of the differing inflammatory and vascular response to its implantation compared with Integra(®).

Topics: Animals; Burns; Chondroitin Sulfates; Collagen; Disease Models, Animal; Male; Mice; Mice, Nude; Skin Transplantation; Skin, Artificial; Tissue Scaffolds; Wound Healing

Chondroitin sulfate (CS) plays important roles in the complement system. However, the CS structure is complicated due to different sources and the number and positions of sulfate groups. The objective of this study was to prepare different low molecular weight chondroitin sulfates (LMWCSs) and to investigate the biological activity in anti-complement capacity. A series of LMWCSs was prepared from different sources and characterized by ultraviolet-visible (UV) spectroscopy, high-performance liquid chromatography (HPLC), size exclusion chromatography-multiangle laser light scattering (SEC-MALLS) and nuclear magnetic resonance (NMR) spectroscopy. Hemolytic, anti-complement 3 deposition capacity and cell viability assays were carried out to investigate the biological activities in vitro. The results showed that LMWCS prepared from shark cartilage with the oxidative degradation method (LMWCS-S-O) had the best anti-complement capacity. LMWCS-S-O could inhibit the alternative pathway of the complement system and protect chondrocytes from cell death. The attenuating effect of LMWCS-S-O on Osteoarthritis (OA) was investigated by destabilization of the medial meniscus (DMM) model in vivo. Functional wind-up, histological and C5b-9 analyses were used to evaluate the treatment effect on the OA model. In vivo results showed that LMWCS-S-O could attenuate OA. LMWCS-S-O with a high content of ΔDi-2,6diS and ΔDi-6S could be used for attenuating OA through regulating the complement system.

Topics: Animals; Cell Survival; Chondrocytes; Chondroitin Sulfates; Disease Models, Animal; Male; Mice; Mice, Inbred C57BL; Molecular Weight; Osteoarthritis; Random Allocation; Treatment Outcome

Osteoarthritis (OA) is a degenerative joint disease, which has no complete treatment with medication yet. Intraarticular hyaluronan (HA) injection can decrease pain and modify the natural course of OA. This study was designed to provide long term delivery of an MMP (matrix-metalloproteinase) inhibitor agent-doxycycline, together with matrix regenerative agent-chondroitin sulfate for treating OA which progress with matrix degenerations. Doxycycline (D) and doxycycline-chondroitin sulfate (D-CS) loaded poly-ɛ-caprolactone (PCL) microspheres (MS) were prepared as intraarticular delivery systems. Bio-effectiveness of developed microspheres was first evaluated with three-dimensional in vitro model of OA where both MS showed significant reduction in MMP-13 levels compared to untreated OA-chondrocytes at 15 and 24 days. Significant decrease was observed in GAG release into the media for both D MS and D-CS MS treated groups at 15 and 24 days. Second, the microspheres were injected to rabbit knee in hyaluronan (HA) to evaluate the effectiveness of the treatment. Radiographic scores of D MS and D-CS MS groups improved after 8 weeks when compared to OA group. Mankin-Pitzker histological scores similarly showed improvement with D MS and D-CSMS groups when compared to OA group. Ex vivo hardness tests of cartilages demonstrated superior hardness values with both doses of D-CSMS compared to OA group. D MS showed promising improvement of OA in histology results. Although, both MS groups had similar effects on cells in the in vitro model, D-CSMS had a positive contribution on all in vivo treatment outcomes and showed potential as a new strategy for treatment when applied to OA knee joints.

Topics: Animals; Chondroitin Sulfates; Disease Models, Animal; Doxycycline; Drug Evaluation, Preclinical; Injections, Intra-Articular; Microspheres; Osteoarthritis; Rabbits

We analyzed the urothelium of cats diagnosed with feline interstitial cystitis to determine whether abnormalities in protein expression patterns could be detected and whether the expression pattern was similar to that in patients with human interstitial cystitis/bladder pain syndrome. The proteins analyzed are involved in cell adhesion and barrier function, comprise the glycosaminoglycan layer or are differentiation markers.. Formalin fixed biopsies from 8 cats with feline interstitial cystitis and from 7 healthy control cats were labeled by immunohistochemistry and scored with a modified version of a system previously used for human samples. Cluster analysis was performed to investigate relationships between markers and samples.. Of the feline interstitial cystitis bladders 89% showed abnormal protein expression and chondroitin sulfate patterns while only 27% of normal tissues showed slight abnormalities. Abnormalities were found in most feline interstitial cystitis samples, including biglycan in 87.5%, chondroitin sulfate, decorin, E-cadherin and keratin-20 in 100%, uroplakin in 50% and ZO-1 in 87.5%. In feline interstitial cystitis bladders about 75% of chondroitin sulfate, biglycan and decorin samples demonstrated absent luminal staining or no staining. Cluster analysis revealed that feline interstitial cystitis and normal samples could be clearly separated into 2 groups, showing that the urothelium of cats with feline interstitial cystitis is altered from normal urothelium.. Feline interstitial cystitis produces changes in luminal glycosaminoglycan and several proteins similar to that in patients, suggesting some commonality in mechanism. Results support the use of feline interstitial cystitis as a model of human interstitial cystitis.

Topics: Animals; Biomarkers; Cats; Cell Differentiation; Chondroitin Sulfates; Cystitis, Interstitial; Disease Models, Animal; Humans; Immunohistochemistry; Proteins; Urothelium

This study investigates the effect of a new combination of glucosamine hydrochloride, chondroitin sulfate, methylsulfonylmethane, Harpagophytum procumbens root extract (standardized to 3% harpagoside) and bromelain extract (GCMHB) on formalin-induced damage to cartilage tissue in the rat knee joint and evaluates this combination in comparison with another combination of glucosamine hydrochloride, chondroitin sulfate and methylsulfonylmethane (GKM).. Animals in the control group were injected with formalin into the knee joint (FCG). Animals in the GCMHB-500 group were given 500mg/kg GCMHB+formalin, and those in the GKM-500 group were given 500mg/kg GKM+formalin. Finally, a healthy group (HG) was also used. GCMHB and GKM were administered to rats orally once a day for 30days. At the end of this period, the rats were sacrificed and the levels of MDA, NO, 8-OH/Gua, and tGSH in the knee joint tissue were measured. Analysis of IL-1β and TNF-α gene expression was done and the tissue was evaluated histopathologically.. MDA, NO and 8-OH/Gua levels and IL-1β and TNF-α gene expression were significantly lower in the GCMHB-500 group compared to the FCG group, whereas tGSH was significantly higher in the GCMHB-500 group than in the FCG group. No significant difference was found for the IL-1β, TNF-α and oxidant/antioxidant parameters between the GKM and FCG groups. The histopathological analysis showed that GCMHB could prevent damage to the cartilage joint, whereas GKM could not.. GCMHB may be used clinically by comparing with GKM in the treatment of osteoarthritis.

Topics: Animals; Bromelains; Cartilage; Chondroitin Sulfates; Dimethyl Sulfoxide; Disease Models, Animal; Drug Combinations; Formaldehyde; Gene Expression Regulation; Glucosamine; Harpagophytum; Interleukin-1beta; Knee Joint; Male; Osteoarthritis; Plant Extracts; Rats; Rats, Wistar; Sulfones; Tumor Necrosis Factor-alpha

Osteoarthritis is thought to be the most prevalent chronic and disabling joint disease in animals and humans and its treatment is a major orthopaedic challenge because there is no ideal drug treatment to preserve joint structure and function, as well as to ameliorate the symptomatology of the disease. The aim of the present study was to assess, using histology, histomorphometry and micro-CT, the effects of the treatment with several drugs of the SYSADOA group and a bisphosphonate in a model of early osteoarthritis, comparing all the results obtained.. Osteoarthritis was surgically induced by anterior cruciate ligament transection and partial meniscectomy on one knee of 56 rabbits; treatment was started three weeks after surgery and lasted 8 weeks; at the end of this period, the animals were sacrificed. Animals were divided into seven groups (8 animals each), one for each regimen of treatment (glucosamine sulfate, chondroitin sulfate, hyaluronic acid, diacerein, risedronate and a combination of risedronate and glucosamine) and one for the control (placebo-treated) group. Following sacrifice, femoral osteochondral cylinders and synovial membrane samples were obtained for their evaluation by qualitative and quantitative histology and micro-CT.. The model induced osteoarthritic changes in the knee joints and none of the treatments showed a significantly better efficacy over the others. Regarding cartilage thickness and volume, all the treatments achieved scores halfway between control groups, without statistical differences. Regarding the synovial membrane, diacerein and risedronate showed the best anti-inflammatory profile, whereas glucosamine and chondroitin did not present any effect standing the hyaluronic acid results between the others. Regarding the subchondral bone, there were no differences in thickness or volume, but risedronate, diacerein and hyaluronic acid seemed to have considerably modified the orientation of the trabecular lattice.. Out of the different drugs evaluated in the study for OA treatment, diacerein and risedronate showed, in all the parameters measured, a better profile of effectiveness; hyaluronic acid ameliorated cartilage swelling and promoted bone formation, but with a poor synovial effect; and finally, chondroitin and glucosamine sulfate prevented cartilage swelling in a similar way to the others, but had no effect on cartilage surface, synovial membrane or subchondral bone.

Topics: Animals; Anthraquinones; Anti-Inflammatory Agents; Arthrography; Bone Density Conservation Agents; Cartilage, Articular; Chondroitin Sulfates; Disease Models, Animal; Drug Therapy, Combination; Female; Femur; Glucosamine; Hyaluronic Acid; Joints; Osteoarthritis; Rabbits; Risedronic Acid; Synovial Membrane; Tomography, X-Ray Computed

In the present study, we investigated the effects of low molecular weight chondroitin sulfate (LMWCS) on amyloid beta (Aβ)-induced neurotoxicity in vitro and in vivo. The in vitro results showed that LMWCS blocked Aβ25-35-induced cell viability loss and apoptosis, decreased intracellular calcium concentration, reactive oxygen species (ROS) levels, the mitochondrial membrane potential (MMP) depolarization, and the protein expression of Caspase-3. During in vivo experiments, LMWCS improved the cognitive impairment induced by Aβ1-40, increased the level of choline acetyltransferase (ChAT), superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px), and decreased the level of malondialdehyde (MDA) and acetylcholinesterase (AChE) in the mouse brain. Moreover, LMWCS decreased the density of pyramidal cells of CA1 regions, and suppressed the protein expression of Bax/Bcl-2 and Caspase-3, -9 in the hippocampus of mice. In conclusion, LMWCS possessed neuroprotective properties against toxic effects induced by Aβ peptides both in vitro and in vivo, which might be related to anti-apoptotic activity. LMWCS might be a useful preventive and therapeutic compound for Alzheimer's disease.

Topics: Acetylcholinesterase; Alzheimer Disease; Amyloid beta-Peptides; Animals; Brain Injuries; Calcium; Caspase 3; Cell Line; Choline O-Acetyltransferase; Chondroitin Sulfates; Disease Models, Animal; Exploratory Behavior; Male; Maze Learning; Membrane Potentials; Mice; Mice, Inbred BALB C; Neuroblastoma; Neuroprotective Agents; Peptide Fragments; Rats; Reactive Oxygen Species

Atherogenesis is associated with the early retention of low-density lipoproteins (LDL) in the arterial intima by interaction with glycosaminoglycan (GAG)-side chains of proteoglycans. Retained LDL undergo reactive oxygen species-mediated oxidation. Oxidized LDL trigger oxidative stress (OS) and inflammation, contributing to atherosclerosis development. Recently, we reported the preventive anti-atherogenic properties of the chimeric mouse/human monoclonal antibody (mAb) chP3R99-LALA, which were related to the induction of anti-chondroitin sulfate antibody response able to inhibit chondroitin sulfate dependent LDL-enhanced oxidation. In the present work, we aimed at further investigating the impact of chP3R99-LALA mAb vaccination on progressive atherosclerosis in apolipoprotein E-deficient mice (apoE(-/-)) fed with a high-fat high-cholesterol diet receiving 5 doses (50 µg) of the antibody subcutaneously, when ~5% of the aortic area was covered by lesions. Therapeutic immunization with chP3R99-LALA mAb halted atherosclerotic lesions progression. In addition, aortic OS was modulated, as shown by a significant (p<0.05) reduction of lipid and protein oxidation, preservation of antioxidant enzymes activity and reduced glutathione, together with a decrease of nitric oxide levels. chP3R99-LALA mAb immunization also regulated aortic NF-κB activation, diminishing the proinflammatory IL1-β and TNF-α gene expression as well as the infiltration of macrophages into the arterial wall. The therapeutic immunization of apoE(-/-) with progressive atheromas and persistent hypercholesterolemia using chP3R99-LALA mAb arrested further development of lesions, accompanied by a decrease of aortic OS and NF-κB-regulated pro-inflammatory cytokine gene expression. These results contribute to broaden the potential use of this anti-GAG antibody-based immunotherapy as a novel approach to target atherosclerosis at different phases of progression.

Topics: Animals; Antibodies, Monoclonal; Apolipoproteins E; Atherosclerosis; Chondroitin Sulfates; Disease Models, Animal; Enzyme-Linked Immunosorbent Assay; Glycosaminoglycans; Humans; Immunohistochemistry; Male; Mice; Mice, Knockout; Oxidation-Reduction; Polymerase Chain Reaction; Recombinant Fusion Proteins; Vaccination

Glycosaminoglycan (GAG) chains of proteoglycans (PGs) play important roles in fibrosis through cell-matrix interactions and growth factor binding in the extracellular matrix. We investigated the expression and regulation of PG core protein (versican) and key enzymes (xylosyltransferase [XT]-I, β1,3-glucuronosyltransferase [GlcAT]-I, chondroitin-4-sulfotransferase [C4ST]) implicated in synthesis and sulfation of GAGs in bleomycin (BLM) and adenovirus-transforming growth factor (TGF)-β1-induced lung fibrosis in rats. We also studied the role of GlcAT-I or TGF-β1 and the signaling pathways regulating PG-GAG production in primary lung fibroblasts isolated from saline- or BLM-instilled rats. The mRNA for XT-I, GlcAT-I, C4ST, and versican was increased in the lung 14 days after BLM injury. In vitro studies indicate that fibrotic lung fibroblasts (FLFs) expressed more XT-I, C4ST, and chondroitin sulfate (CS)-GAGs than did normal lung fibroblasts at baseline. TGF-β1 enhanced the expression of XT-I, C4ST-I, and versican in normal lung fibroblasts, whereas SB203580 or SB431542, by targeting p38 mitogen-activated protein kinase or TGF-β type-1 receptor/activin receptor-like kinase 5, respectively, attenuated the response to both TGF-β1 and FLFs on PG-GAG expression. Neutralizing anti-TGF-β1 antibody abrogated FLF-conditioned medium-stimulated expression of XT-I, GlcAT-I, versican, and CS-GAG. Forced expression of TGF-β1 in vivo enhanced versican, XT-I, GlcAT-I, and C4ST-I expression and PG-GAG deposition in rat lungs. Finally, induced expression of GlcAT-I gene in rat lung fibroblasts increased GAG synthesis by these cells. Together, our results provide new insights into the basis for increased PG-GAG deposition in lung fibrosis; inhibition of TGF-β1-mediated or fibrosis-induced PG-GAG production by activin receptor-like kinase 5/p38 inhibitors may contribute to antifibrotic activity.

Topics: Animals; Antibodies, Neutralizing; Bleomycin; Cells, Cultured; Chondroitin Sulfates; Disease Models, Animal; Fibroblasts; Gene Expression Regulation, Enzymologic; Glucuronosyltransferase; Glycosaminoglycans; Glycosyltransferases; Lung; Male; p38 Mitogen-Activated Protein Kinases; Pentosyltransferases; Protein Kinase Inhibitors; Protein Serine-Threonine Kinases; Pulmonary Fibrosis; Rats; Rats, Sprague-Dawley; Receptor, Transforming Growth Factor-beta Type I; Receptors, Transforming Growth Factor beta; RNA, Messenger; Signal Transduction; Sulfotransferases; Time Factors; Transfection; Transforming Growth Factor beta1; UDP Xylose-Protein Xylosyltransferase; Up-Regulation; Versicans

To investigate beneficial effect of the readily available colchicine through its intravesical application on protamine/lipopolysaccharide induced interstitial cystitis model in rat and to compare its efficacy to the chondroitin sulphate available for clinical use.. Twenty-four Wistar female rats were assigned to control (C), interstitial cystitis (IC), chondroitin sulphate (CS) and colchicine (Col) groups. IC, CS and Col groups received protamine sulphate and lipopolysaccharide (PS/LPS) instillation. Testing agents CS and Col were administered a day after PS/LPS inoculation into the bladders. Rats in Group C received saline solution. CS and Col groups received 1 mL CS (0.2%) and 1 mL Col (0.05 mg/mL). The treatment agents were left in bladders for one hour's duration. Animals were sacrificed 5 days after the inoculation and the bladder tissues were examined histologically to evaluate the amount of extravasated leucocytes, mast cell concentration (by counting total number of cells per 10 high power field (hpf; 1 hpf = ×400 magnification) as well as interstitial tissue edema for each bladder.. Intravesical application of CS reduced significantly the leucocyte and mast cell infiltration as well as interstitial edema compared to group C . The level of reduction in leucocyte and mast cell infiltration in Col group was comparable to that of CS, although the interstitial edema was not resolved.. Intravesical administration of Col decreased leucocyte and mast cell infiltration to the same extent of CS in PS/LPS induced bladder inflammation in rat. Col may be an alternative to other treatment modalities for painful bladder conditions such as IC.

Topics: Administration, Intravesical; Animals; Chondroitin Sulfates; Colchicine; Cystitis; Cystitis, Interstitial; Disease Models, Animal; Female; Lipopolysaccharides; Protamines; Rats; Rats, Wistar

Glial and fibrotic scars inhibit neural regeneration after spinal cord injury (SCI). N-[3,4-dimethoxycinnamoyl]-anthranilic acid (tranilast) inhibits transforming growth factor β, alleviates allergic reactions, and decreases hypertrophic skin scars. We evaluated its ability to improve motor function and inhibit the spread of tissue damage in rats with SCI.. Rats with SCI were divided into groups that received tranilast (30 mg/[kg · day]) by intravenous administration (group IV), tranilast (200mg/[kg · day]) by oral administration (group OR), and saline injections (control). Motor functions were assessed by determining Basso, Beattie, and Bresnahan (BBB) scores and %grip tests for 8 weeks after SCI. Histological evaluation of ionized calcium binding adaptor molecule 1 (Iba1) at 1 week after SCI and glial fibrillary acidic protein (GFAP), fibronectin, and chondroitin sulfate (CS) at week 8 was performed.. Motor function recovery, BBB score, and the %grip test were significantly higher in the tranilast-treated groups than in the control group. At week 1 after SCI, inflammatory-cell invasion was more severe and Iba1 expression was significantly higher in the control group. At week 8, although the number of GFAP-positive cells increased greatly from the impaction site to the proximal and distal sites in the control group, these cells were confined around a cavity in the tranilast-treated groups. GFAP distribution coincided with that of fibronectin. Anti-CS antibody level in the tranilast-treated groups was significantly lower than that in the control group.. Tranilast inhibits inflammation in the acute phase of SCI and reduces glial and fibrotic scars and could present a new method for treating SCI.

Topics: Animals; Calcium-Binding Proteins; Chondroitin Sulfates; Disease Models, Animal; Fibronectins; Glial Fibrillary Acidic Protein; Microfilament Proteins; Motor Activity; ortho-Aminobenzoates; Rats; Rats, Sprague-Dawley; Recovery of Function; Spinal Cord; Spinal Cord Injuries

We planned to report on the effect of two nonanimal chondroitin sulfates (CSs) with different molecular masses produced using an innovative biotechnological process in an adjuvant arthritis animal model.. The experiments included healthy animals, untreated arthritic animals and arthritic animals having been administered 900 mg/kg of either of the two CS samples daily. Arthritic score, γ-glutamyltransferase (GGT) activity in hind paw joint tissue homogenates, plasmatic C-reactive protein (CRP) and pro-inflammatory cytokines IL-1β and IL-6 were assayed.. Low-molecular-mass (LMM) CS significantly reduced the arthritic score by up to about 30% from 14 to 28 days. In contrast, no significant differences were observed for high-molecular-mass (HMM) CS, even if a trend in its capacity to decrease the arthritic score by up to about 11% was observed. Additionally, LMM CS was able to significantly decrease GGT activity by approximately 31% and plasmatic CRP levels by about 9%. Both nonanimal CS samples were effective in reducing plasmatic levels of proinflammatory cytokines. A greater efficacy was also observed for LMM CS compared with a pharmaceutical-grade CS of extractive origin, while the efficacy of the HMM CS sample was found to be rather similar. The greater effect of LMM CS in reducing arthritic parameters may be related to its lower molecular mass with respect to HMM CS and natural CS.

Topics: Animals; Anti-Inflammatory Agents; Arthritis; C-Reactive Protein; Chondroitin Sulfates; Disease Models, Animal; gamma-Glutamyltransferase; Interleukin-1beta; Interleukin-6; Male; Rats, Inbred Lew; Tarsal Joints

Dermatan sulfate (DS) is synthesized from chondroitin sulfate (CS) by epimerization of glucuronic acid of CS to yield iduronic acid. In the present study, the role of CS and DS was examined in mice that received transection of nigrostriatal dopaminergic pathway followed by injection of glycosaminoglycan degrading enzymes into the lesion site. Two weeks after injury, fibrotic and glial scars were formed around the lesion, and transected axons did not regenerate beyond the fibrotic scar. Injection of chondroitinase ABC (ChABC), which degrades both CS and DS, completely suppressed the fibrotic scar formation, reduced the glial scar, and promoted the regeneration of dopaminergic axons. Injection of the DS-degrading enzyme chondroitinase B (ChB) also yielded similar results. By contrast, injection of chondroitinase AC (ChAC), a CS-degrading enzyme, did not suppress the fibrotic and glial scar formation, but reduced CS immunoreactivity and promoted the axonal regeneration. Addition of transforming growth factor-β1 (TGF-β1) to a co-culture of meningeal fibroblasts and cerebral astrocytes induces a fibrotic scar-like cell cluster. The effect of TGF-β1 on cluster formation was suppressed by treatment with ChABC or ChB, but not by ChAC. TGF-β1-induced cell cluster repelled neurites of neonatal cerebellar neurons, but addition of ChABC or ChAC suppressed the inhibitory property of clusters on neurite outgrowth. The present study is the first to demonstrate that DS and CS play different functions after brain injury: DS is involved in the lesion scar formation, and CS inhibits axonal regeneration.

Topics: Animals; Astrocytes; Axons; Brain Injuries; Chondroitin Sulfates; Cicatrix; Coculture Techniques; Dermatan Sulfate; Disease Models, Animal; Fibroblasts; Fluorescent Antibody Technique; Immunohistochemistry; Male; Mice; Mice, Inbred ICR; Nerve Regeneration; Rats; Rats, Sprague-Dawley

Bacterial chondroitinase ABC (ChaseABC) has been used to remove the inhibitory chondroitin sulfate chains from chondroitin sulfate proteoglycans to improve regeneration after rodent spinal cord injury. We hypothesized that the mammalian enzyme arylsulfatase B (ARSB) would also enhance recovery after mouse spinal cord injury. Application of the mammalian enzyme would be an attractive alternative to ChaseABC because of its more robust chemical stability and reduced immunogenicity. A one-time injection of human ARSB into injured mouse spinal cord eliminated immunoreactivity for chondroitin sulfates within five days, and up to 9 weeks after injury. After a moderate spinal cord injury, we observed improvements of locomotor recovery assessed by the Basso Mouse Scale (BMS) in ARSB treated mice, compared to the buffer-treated control group, at 6 weeks after injection. After a severe spinal cord injury, mice injected with equivalent units of ARSB or ChaseABC improved similarly and both groups achieved significantly more locomotor recovery than the buffer-treated control mice. Serotonin and tyrosine hydroxylase immunoreactive axons were more extensively present in mouse spinal cords treated with ARSB and ChaseABC, and the immunoreactive axons penetrated further beyond the injury site in ARSB or ChaseABC treated mice than in control mice. These results indicate that mammalian ARSB improves functional recovery after CNS injury. The structural/molecular mechanisms underlying the observed functional improvement remain to be elucidated.

Topics: Animals; Bacterial Proteins; Chondroitin ABC Lyase; Chondroitin Sulfates; Disease Models, Animal; Female; Humans; Locomotion; Mice; N-Acetylgalactosamine-4-Sulfatase; Recombinant Proteins; Recovery of Function; Spinal Cord Injuries

Loxoscelism is the designation given to clinical symptoms evoked by Loxosceles spider's bites. Clinical manifestations include skin necrosis with gravitational spreading and systemic disturbs. The venom contains several enzymatic toxins. Herein, we describe the cloning, expression, refolding and biological evaluation of a novel brown spider protein characterized as a hyaluronidase. Employing a venom gland cDNA library, we cloned a hyaluronidase (1200 bp cDNA) that encodes for a signal peptide and a mature protein. Amino acid alignment revealed a structural relationship with members of hyaluronidase family, such as scorpion and snake species. Recombinant hyaluronidase was expressed as N-terminal His-tag fusion protein (∼45 kDa) in inclusion bodies and activity was achieved using refolding. Immunoblot analysis showed that antibodies that recognize the recombinant protein cross-reacted with hyaluronidase from whole venom as well as an anti-venom serum reacted with recombinant protein. Recombinant hyaluronidase was able to degrade purified hyaluronic acid (HA) and chondroitin sulfate (CS), while dermatan sulfate (DS) and heparan sulfate (HS) were not affected. Zymograph experiments resulted in ∼45 kDa lytic zones in hyaluronic acid (HA) and chondroitin sulfate (CS) substrates. Through in vivo experiments of dermonecrosis using rabbit skin, the recombinant hyaluronidase was shown to increase the dermonecrotic effect produced by recombinant dermonecrotic toxin from L. intermedia venom (LiRecDT1). These data support the hypothesis that hyaluronidase is a "spreading factor". Recombinant hyaluronidase provides a useful tool for biotechnological ends. We propose the name Dietrich's Hyaluronidase for this enzyme, in honor of Professor Carl Peter von Dietrich, who dedicated his life to studying proteoglycans and glycosaminoglycans.

Topics: Animals; Arachnida; Arthropod Proteins; Chondroitin Sulfates; Cloning, Molecular; Disease Models, Animal; Hyaluronic Acid; Hyaluronoglucosaminidase; Insect Bites and Stings; Molecular Sequence Data; Molecular Weight; Phylogeny; Rabbits; Recombinant Proteins; Sequence Alignment; Sequence Analysis, DNA; Sequence Homology, Amino Acid; Substrate Specificity; Venoms

Aggrecan, a major macromolecule in cartilage, protects the extracellular matrix (ECM) from degradation during the progression of osteoarthritis (OA). However, aggrecan itself is also susceptible to proteolytic cleavage. Here, the use of a biomimetic proteoglycan (mAGC) is presented, which functionally mimics aggrecan but lacks the known cleavage sites, protecting the molecule from proteolytic degradation. The objective of this study is to test the efficacy of this molecule in ex vivo (human OA synovial fluid) and in vivo (Sprague-Dawley rats) osteoarthritic models. These results indicate that mAGC's may protect articular cartilage against the loss of key ECM components, and lower catabolic protein and gene expression in both models. This suppression of matrix degradation has the potential to provide a healthy environment for tissue repair.

Topics: Aged; Aged, 80 and over; Aggrecans; Animals; Biomimetic Materials; Cartilage, Articular; Cattle; Chondrocytes; Chondroitin Sulfates; Cytokines; Disease Models, Animal; Extracellular Matrix; Gene Expression Regulation; Humans; Hyaluronic Acid; Inflammation Mediators; Matrix Metalloproteinase 13; Middle Aged; Osteoarthritis; Rats; Rats, Sprague-Dawley

To evaluate the effects of hyperprolactinemia on the sulfated glycosaminoglycans (GAGs) of the murine uterus.. Experimental research.. University biochemistry laboratory.. Eighty female mice were divided into two groups of 40 animals each and treated with 0.2 mL of saline solution (controls, Ctr) and 200 μg of metoclopramide (experimental, HPrl). Treatments lasted for 50 consecutive days. The animals were divided into four subgroups of 10 animals each per treatment (Ctr and HPrl) and sacrificed according to the phase of the estrous cycle. The uterine horns were removed for biochemical analyses, and blood samples were collected for hormone measurements.. Induced hyperprolactinemia.. To quantify the sulfated GAGs, and PRL and sex steroid levels.. The endometrium during the estrus phase was significantly thicker in the HPrl animals than in the Ctr mice. The levels of chondroitin and dermatan sulfate were significantly increased in the HPrl group than in the Ctr group during all phases except metestrus. The amounts of heparan sulfate were lower during estrus and diestrus and higher in the metestrus phase in HPrl than in Ctr animals. Serum PRL levels were increased whereas the levels of E2 and P were decreased in all phases in the HPrl group than in the Ctr group.. The hyperprolactinemia changed the amounts of uterine sulfated GAGs. Our data suggest that these changes may not be correlated with ovarian steroid levels.

Topics: Animals; Biomarkers; Chondroitin Sulfates; Dermatan Sulfate; Disease Models, Animal; Estradiol; Estrous Cycle; Female; Glycosaminoglycans; Heparitin Sulfate; Hyperprolactinemia; Metoclopramide; Mice; Progesterone; Time Factors; Uterus

The study aimed to investigate the effect of the oral administration for 15 days of either Echinacea (E) or genuphil (a composite of chondroitin sulphate, glucosamine and methyl sulfonyl methane [GCM]) nutraceutical supplements on female rat model of acute or chronic arthritis induced by bacterial outer membrane protein (OMP) from faecal flora of healthy and rheumatic humans. Anti-cyclic citrullinated peptide (anti-CCP2), C-reactive protein (CRP) and rheumatoid factor (RF) values increased (p < 0.05) in both arthritic groups as compared to normal values. The rheumatic markers anti-CCP2, CRP and RF values decreased significantly in E- and GCM-treated groups compared to arthritic none-treated acute or chronic groups. The results of RF values of GCM-treated groups in acute and chronic models decreased exhibiting no statistical difference compared with the normal value. Histological examinations of the hind paw sections revealed moderate inflammation, oedema and mild proliferation of synovial cells in acute arthritic rats and more damage to cartilage and bone with severe inflammation in chronic ones. Echinacea acute treated group showed edema with proliferated synovial membrane and partial damage in cartilage and bone. While in the E-chronic treated group, rough edge with destructed cartilage and bone existed. However, the acute GCM group revealed mild cartilage damage. But the chronic GCM group showed mild synovial cells proliferation and revealed no inflammation with mild cartilage damage edge. Results demonstrated the OMP arthropathic property and through promising light on arthritis treatment using E- or GCM, with the advantage of GMC results over that of E-. The composite GCM is needed for further studies over the dose and duration to assess its preventive effects against the bacterial OMP arthrogenicity.

Topics: Animals; Antirheumatic Agents; Arthritis, Experimental; Arthritis, Rheumatoid; Chondroitin Sulfates; Complex Mixtures; Dietary Supplements; Dimethyl Sulfoxide; Disease Models, Animal; Echinacea; Female; Glucosamine; Lysosomes; Plant Extracts; Rats; Stifle; Sulfones

The aim of this study was to investigate the correlations of severity of osteoarthritis (OA) and serum biomarkers including keratan sulfate (KS), hyaluronic acid (HA) and chondroitin sulfate (CS) 846 epitope. We also investigated the effect of glucosamine and fish collagen peptide (FCP) on OA. OA was induced in 12 rabbits (12 weeks of age) by anterior cruciate ligament transection (ACLT). After the surgery, the rabbits were orally administered FCP (F group), glucosamine (G group) or FCP and glucosamine (FG group) for 4 weeks. The control group was provided water ad libitum (C group). Blood was collected before surgery (pre-ACLT) and before euthanasia (post-ACLT) for serum marker measurement. Biomarker levels were measured by using commercial kits. We evaluated OA severity both macroscopically and histologically. Macroscopic evaluation showed mildly eroded condylar surfaces in the C group. Histological findings were significantly different from the FG and other groups. There were no significant differences between each group at post-ACLT in terms of serum KS, HA and CS 846. Histological assessment and serum biomarker measurements performed at post-ACLT showed a significant correlation between HA concentration and OA severity. Variations in the CS 846 concentration at pre-ACLT and post-ACLT were significantly correlated with OA severity. Administration of glucosamine and FCP had chondroprotective effects in the ACLT model. Serum biomarker concentrations were significantly correlated with cartilage injury. Serum biomarker measurement would be useful for monitoring articular cartilage damage in the clinical setting.

Topics: Animals; Anterior Cruciate Ligament; Biomarkers; Cartilage, Articular; Chondroitin Sulfates; Collagen; Disease Models, Animal; Glucosamine; Histocytochemistry; Hyaluronic Acid; Keratan Sulfate; Osteoarthritis; Rabbits; Statistics, Nonparametric

Dermal templates, such as Matriderm® and Integra®, are widely used in plastic and reconstructive surgery, often as two-step procedures. A recent development is the application of thin dermal templates covered with split thickness skin grafts in one-step procedures. In this experimental study, we compare the two thin matrices Matriderm® 1 mm and Integra® Single Layer in a one-step procedure with particular focus on neodermis formation.. Matriderm® 1 mm and Integra® Dermal Regeneration Template-Single Layer (1.3 mm) were compared in a rat model. In three groups of five animals each, a full thickness wound was covered with (a) Matriderm® 1 mm and neonatal rat epidermis, (b) Integra® Single Layer and neonatal rat epidermis, or, (c) neonatal rat epidermis only (control). Histological sections 2 weeks post transplantation were analyzed with regard to take of template and epidermis, neodermal thickness, collagen deposition, vascularization, and inflammatory response.. Take of both templates was complete in all animals. The Matriderm®-based neodermis was thinner but showed a higher cell density than the Integra®-based neodermis. The other parameters were similar in both matrices.. The two templates demonstrate a comparable biological behavior early after transplantation. The only difference was found regarding neodermal thickness, probably resulting from faster degradation of Matriderm®. These preliminary data suggest that both dermal templates appear similarly suitable for transplantation in a one-step procedure.

Topics: Animals; Animals, Newborn; Chondroitin Sulfates; Collagen; Dermatologic Surgical Procedures; Disease Models, Animal; Elastin; Epidermis; Female; Follow-Up Studies; Plastic Surgery Procedures; Rats; Rats, Nude; Skin; Skin Transplantation; Skin, Artificial; Wound Healing; Wounds and Injuries

Histological and molecular changes were examined to investigate the effects of long-term administration of glucosamine (GlcN) and chondroitin sulfate (CS) in a model of spontaneous osteoarthritis (OA) in Hartley guinea pigs. Three groups of female 3-week-old Hartley guinea pigs received GlcN, CS, and neither agent, respectively. Five animals in each group were sacrificed at 8, 12, and 18 months of age. At 8 months of age, Hartley guinea pigs had severe degeneration of knee joint cartilage, chondrocyte apoptosis, marked reduction of tissue total RNA, decreases of aggrecan and collagen type 2 mRNAs, and increases in MMP-3 and MMP-8 mRNAs. Long-term administration of GlcN and CS reduced cartilage degeneration at 8 months of age. The marked loss of total RNA and the increase in MMP-3 mRNA were also inhibited by GlcN and CS. Thus, long-term oral administration of GlcN or CS inhibits OA progression, maintains total RNA and down-regulates MMP-3 mRNA in a spontaneous OA model in Hartley guinea pigs.

Topics: Animals; Apoptosis; Cartilage, Articular; Chondrocytes; Chondroitin Sulfates; Disease Models, Animal; Drug Evaluation, Preclinical; Female; Glucosamine; Guinea Pigs; Matrix Metalloproteinase 3; Osteoarthritis; RNA, Messenger; Up-Regulation

A vaccine protecting women against placental malaria could be based on the sub-domains of the VAR2CSA antigen, since antibodies against the DBL4ɛ-ID4 subunit of the VAR2CSA protein can inhibit parasite binding to the placental ligand chondroitin sulphate A (CSA). Here we tested the ability of DBL4ɛ-ID4 to induce binding-inhibitory antibodies when formulated with adjuvants approved for human use. We have characterized the immune response of DBL4ɛ-ID4 in combination with Freund's complete and incomplete adjuvant and with three adjuvants currently being used in clinical trials: Montanide(®) ISA 720, Alhydrogel(®) and CAF01. Antibodies induced against DBL4ɛ-ID4 in combination with these adjuvants inhibited parasite binding to CSA from 82% to 99%. Although, different epitope recognition patterns were obtained for the different formulations, all adjuvant combinations induced strong Th1 and Th2 type responses, resulting in IgG with similar binding strength, with to the DBL4ɛ-ID4 antigen. These results demonstrate that the DBL4ɛ-ID4 antigen is highly immunogenic and that binding inhibitory antibodies are induced when formulated with any of the tested adjuvants.

Topics: Adjuvants, Immunologic; Animals; Antibodies, Protozoan; Antigens, Protozoan; Cell Adhesion; Chondroitin Sulfates; Disease Models, Animal; Female; Malaria Vaccines; Malaria, Falciparum; Placenta Diseases; Pregnancy; Rats; Rats, Wistar

To investigate whether a physiologic effect of "glycosaminoglycan (GAG) replenishment therapy" altered recruitment of inflammatory cells in an acute bladder damage model. Replacement of the GAG layer with intravesically administered GAGs is an effective therapy for interstitial cystitis in at least some patients. Intravesically administered chondroitin sulfate was previously shown to bind to and restore the impermeability of surface-damaged ("leaky") urothelium to small ions.. Rat bladders were damaged with 10 mM HCl. Negative control bladders were treated with phosphate-buffered saline. On the following day, the animal bladders were treated with 20 mg/mL chondroitin sulfate in phosphate-buffered saline, and the negative and positive controls were treated with phosphate-buffered saline alone. At 2 and 4 days after treatment with chondroitin sulfate, the rats were killed, and sections of their bladders were analyzed using toluidine blue staining for mast cell immunohistochemical labeling using antibodies against CD45 for lymphocytes and myeloperoxidase for neutrophils.. Chondroitin sulfate treatment reduced the recruitment, in a statistically significant manner, of inflammatory cells, including neutrophils and mast cells to the suburothelial space but did not alter recruitment of CD45-positive lymphocytes.. For the first time, we have demonstrated that intravesical GAG replenishment therapy also produces a physiologic effect of decreasing recruitment of inflammatory cells in an acute model of the damaged bladder. These findings support the use of intravesically administered GAG for bladder disorders that result from a loss of impermeability, including interstitial, radiation, and chemical cystitis, and possibly others as well.

Topics: Animals; Burns, Chemical; Chemotaxis, Leukocyte; Chondroitin Sulfates; Cystitis; Disease Models, Animal; Drug Evaluation, Preclinical; Edema; Hydrochloric Acid; Leukocyte Common Antigens; Lymphocytes; Mast Cells; Neutrophils; Permeability; Peroxidase; Rats; Rats, Sprague-Dawley

Subendothelial retention of proatherogenic lipoproteins by proteoglycans is critical in atherosclerosis. The aim of this study was to characterize the recognition and antiatherogenic properties of a chimeric monoclonal antibody (mAb) that reacts with sulfated molecules.. chP3R99 mAb recognized sulfated glycosaminoglycans, mainly chondroitin sulfate (CS), by ELISA. This mAb blocked ≈70% of low-density lipoprotein (LDL)-CS association and ≈80% of LDL oxidation in vitro, and when intravenously injected to Sprague-Dawley rats (n=6, 1 mg/animal), it inhibited LDL (4 mg/kg intraperitoneally, 1 hour later) retention and oxidation in the artery wall. Moreover, subcutaneous immunization of New Zealand White rabbits (n=19) with chP3R99 mAb (100 μg, 3 doses at weekly intervals) prevented Lipofundin-induced atherosclerosis (2 mL/kg, 8 days) with a 22-fold reduction in the intima-media ratio (P<0.01). Histopathologic and ultrastructural studies showed no intimal alterations or slight thickening, with preserved junctions between endothelial cells and scarce collagen fibers and glycosaminoglycans. In addition, immunization with chP3R99 mAb suppressed macrophage infiltration in aorta and preserved redox status. The atheroprotective effect was associated with the induction of anti-CS antibodies in chP3R99-immunized rabbits, capable of blocking CS-LDL binding and LDL oxidation.. These results support the use of anti-sulfated glycosaminoglycan antibody-based immunotherapy as a potential tool to prevent atherosclerosis.

Topics: Animals; Antibodies, Monoclonal; Antibody Specificity; Atherosclerosis; Biological Transport; Cell Line; Chondroitin Sulfates; Disease Models, Animal; Drug Combinations; Enzyme-Linked Immunosorbent Assay; Foam Cells; Glycosaminoglycans; Immunization; Lipoproteins, LDL; Mice; Oxidation-Reduction; Oxidative Stress; Phospholipids; Rabbits; Rats; Rats, Sprague-Dawley; Sorbitol

Skeletal bone losses are mainly filled with autologous graft or artificial materials. Osteoblasts are essential to maintain bone homeostasis and bone repair through a matrix synthesis. We have previously demonstrated that adherence and regenerative matrix composition are fundamental to bone healing, even in critical situations. In this work the critical size defect technique was used to evaluate the systemic activity on bone regeneration of a novel mixture of extracellular polysaccharides. A 5mm diameter hole was made in each parietal bone of male Wistar rats. The right parietal bone hole was filled with a mixture of hyaluronic acid, chondroitin 6 sulphate, and dermatan sulphate mixed with 2.5% NaCl solution, while the left hole was left free of material and untreated and considered as control. Twenty-one days after surgery, the holes and surrounding tissues were examined visually, using X-rays, and by histological staining. Using the matrix substitute, bone healing was almost complete after 21 days in the treated hole and always complete in the control side due to some systemic effect. Neovascularization was also observed along with organized trabecular bone on both sides. No abnormal bone growth or connective tissue abnormalities were noted. At the end of the experiment, 95.1% (± 3.2) bone healing (n=20) was observed on the treated side; conversely, healing bone and histological structure were better on the control side.

Topics: Animals; Biocompatible Materials; Bone Diseases; Bone Regeneration; Chondroitin Sulfates; Dermatan Sulfate; Disease Models, Animal; Hyaluronic Acid; Male; Neovascularization, Physiologic; Osteogenesis; Parietal Bone; Periosteum; Rats; Rats, Wistar; Time Factors; Wound Healing

Chondroitin sulfate proteoglycans (CSPGs) play a pivotal role in many neuronal growth mechanisms including axon guidance and the modulation of repair processes following injury to the spinal cord or brain. Many actions of CSPGs in the central nervous system (CNS) are governed by the specific sulfation pattern on the glycosaminoglycan (GAG) chains attached to CSPG core proteins. To elucidate the role of CSPGs and sulfated GAG chains following traumatic brain injury (TBI), controlled cortical impact injury of mild to moderate severity was performed over the left sensory motor cortex in mice. Using immunoblotting and immunostaining, we found that TBI resulted in an increase in the CSPGs neurocan and NG2 expression in a tight band surrounding the injury core, which overlapped with the presence of 4-sulfated CS GAGs but not with 6-sulfated GAGs. This increase was observed as early as 7 days post injury (dpi), and persisted for up to 28 dpi. Labeling with markers against microglia/macrophages, NG2+ cells, fibroblasts, and astrocytes showed that these cells were all localized in the area, suggesting multiple origins of chondroitin-4-sulfate increase. TBI also caused a decrease in the expression of aggrecan and phosphacan in the pericontusional cortex with a concomitant reduction in the number of perineuronal nets. In summary, we describe a dual response in CSPGs whereby they may be actively involved in complex repair processes following TBI.

Topics: Animals; Antigens; Brain Injuries; Cerebral Cortex; Chondroitin Sulfates; Disease Models, Animal; Male; Mice; Mice, Inbred C57BL; Nerve Regeneration; Nerve Tissue Proteins; Neurocan; Proteoglycans; Recovery of Function

We evaluated whether viscoelastics have protective effects on the corneal endothelial cell damage in a toxic anterior segment syndrome (TASS) animal model depending on the types of viscoelastics.. A TASS animal model was established with an injection of 0.1 mL o-phthaldehyde solution (0.14%) into the anterior chamber of New Zealand white rabbits. One of two different viscoelastics, 1% sodium hyaluronate (cohesive group) or a 1:3 mixture of 4% chondroitin sulfate and 3% sodium hyaluronate (dispersive group), was injected into the anterior chamber. After five minutes, it was removed using a manual I/A instrument, and then 0.1 mL of o-phthaldehyde solution (0.14%) was injected into the anterior chamber. Damage to corneal endothelial cells was compared between the two groups.. The corneal thickness increased quickly in both groups after the disinfectant injection. However, the dispersive group showed relatively mild corneal edema compared to the cohesive group. The mean corneal haze score in the dispersive group also was lower than that of the cohesive group. These partial protective effects of the dispersive viscoelastic were demonstrated by the different findings of a live/dead cell assay, TUNEL staining, and scanning electron microscopy between the two groups.. The TASS animal model seems to be a useful means to evaluate corneal endothelial cell damage caused by toxic substances to find ways to protect or reduce endothelial cell damage. Dispersive viscoelastics were shown to have partial protective effects against corneal endothelial cell damage caused by a toxic disinfectant.

Topics: Animals; Anterior Eye Segment; CD11b Antigen; Chondroitin Sulfates; Corneal Edema; Corneal Endothelial Cell Loss; Disease Models, Animal; Disinfectants; Endothelium, Corneal; Hyaluronic Acid; o-Phthalaldehyde; Ophthalmic Solutions; Rabbits; Viscoelastic Substances; Viscosupplements

Lymphatic dysfunction in lymphedema results in chronic accumulation of interstitial fluid and life-long tissue swelling. In the absence of restored lymphatic drainage via adequate lymphangiogenesis, the interstitial environment can remodel in ways that decrease the elevated interstitial stress. Presently, relatively little is known about the glycosaminoglycans (GAGs) that become upregulated in the interstitium during lymphedema. We employed a mouse tail model of acute lymphedema that reproduces important features of the chronic human condition to establish a relationship between hyaluronan (HA) and sulfated GAG concentration with tissue swelling. We found that HA was upregulated by tissue injury at day 5 and became upregulated again by skin swelling (HA content increasing by 27% relative to controls at days 15 and 20). Surprisingly, the second phase of HA expression was associated with the declining phase of the tail skin swelling (tail diameter significantly decreasing by 17% from day 10 peak to day 20), demonstrating that HA is upregulated by tissue swelling and may help to counteract the edema in the mouse tail. This finding was confirmed by intradermal injection of an HA degrading enzyme (hyaluronidase) to the swollen tail, which was found to worsen the tail swelling. Sulfated GAGs, including chondroitin sulfate (CS), were not regulated by tissue swelling. The results demonstrate that HA, but not sulfated GAGs, is upregulated in the interstitium by acute tissue swelling. We speculate that HA expression during lymphedema may be part of a natural adaptive mechanism of the interstitial environment to reduce capillary filtration and increase interstitial fluid outflow following lymphatic obstruction and fluid accumulation.

Topics: Acute Disease; Animals; Chondroitin Sulfates; Disease Models, Animal; Glycosaminoglycans; Humans; Hyaluronic Acid; Hyaluronoglucosaminidase; Lymphedema; Mice; Tail; Time Factors

Bioabsorbable unfilled synthetic nerve conduits have been used in the reconstruction of small segmental nerve defects with variable results, especially in motor nerves. We hypothesized that providing a synthetic mimic of the Schwann cell basal lamina in the form of a collagen-glycosaminoglycan (GAG) matrix would improve the bridging of the nerve gap and functional motor recovery.. A unilateral 10-mm sciatic nerve defect was created in eighty-eight male Lewis rats. Animals were randomly divided into four experimental groups: repair with reversed autograft, reconstruction with collagen nerve conduit (1.5-mm NeuraGen, Integra LifeSciences), reconstruction with collagen nerve conduit filled with collagen matrix, and reconstruction with collagen nerve conduit filled with collagen-GAG (chondroitin-6-sulfate) matrix. Nerve regeneration was evaluated at twelve weeks on the basis of the compound muscle action potential, maximum isometric tetanic force, and wet muscle weight of the tibialis anterior muscle, the ankle contracture angle, and nerve histomorphometry.. The use of autograft resulted in significantly better motor recovery compared with the other experimental methods. Conduit filled with collagen-GAG matrix demonstrated superior results compared with empty conduit or conduit filled with collagen matrix with respect to all experimental parameters. Axon counts in the conduit filled with collagen-GAG matrix were not significantly different from those in the reversed autograft at twelve weeks after repair.. The addition of the synthetic collagen basal-lamina matrix with chondroitin-6-sulfate into the lumen of an entubulation repair significantly improved bridging of the nerve gap and functional motor recovery in a rat model.. Use of a nerve conduit filled with collagen-GAG matrix to bridge a motor or mixed nerve defect may result in superior functional motor recovery compared with commercially available empty collagen conduit. However, nerve autograft remains the gold standard for reconstruction of a segmental motor nerve defect.

Topics: Absorbable Implants; Animals; Chondroitin Sulfates; Collagen; Disease Models, Animal; Electromyography; Glycosaminoglycans; Guided Tissue Regeneration; Male; Motor Skills; Nerve Regeneration; Peripheral Nerves; Peripheral Nervous System Diseases; Random Allocation; Rats; Rats, Inbred Lew; Recovery of Function; Reference Values; Sciatic Nerve; Transplantation, Autologous; Treatment Outcome

Heat stroke is a condition characterized by high body temperature that can lead to hemorrhage and necrosis in multiple organs. Anticoagulants, such as danaparoid sodium (DA), inhibit various types of inflammation; however, the anti-inflammatory mechanism of action is not well understood. Given that heat stroke is a severe inflammatory response disease, we hypothesized that DA could inhibit inflammation from heat stress and prevent acute heat stroke.. Male Wistar rats were given a bolus injection of saline or DA (50 U/kg body weight) into the tail vein just prior to heat stress (42 °C for 30 min). Markers of inflammation were then determined in serum and tissue samples.. In rats pretreated with DA, induction of cytokines (interleukin [IL]-1β, IL-6, and tumor necrosis factor [TNF]-α), nitric oxide (NO), and high mobility group box 1 (HMGB1) protein were reduced compared with saline-treated rats. Histologic changes observed in lung, liver, and small intestine tissue samples of saline-treated rats were attenuated in DA-treated rats. Moreover, DA pretreatment improved survival in our rat model of heat stress-induced acute inflammation.. Collectively, our findings demonstrate that DA pretreatment may have value as a new therapeutic tool for heat stroke.

Topics: Acute Disease; Animals; Anticoagulants; Antithrombin III; Chondroitin Sulfates; Cytokines; Dermatan Sulfate; Disease Models, Animal; Fibrin Fibrinogen Degradation Products; Heat Exhaustion; Heat Stroke; Heparitin Sulfate; HMGB1 Protein; Inflammation; Intestine, Small; Liver; Lung; Male; Nitrates; Nitric Oxide; Nitrites; Peptide Hydrolases; Rats; Rats, Wistar; Survival Rate

Pulmonary fibrosis (PF) is characterized by increased deposition of proteoglycans (PGs), in particular core proteins. Glycosaminoglycans (GAGs) are key players in tissue repair and fibrosis, and we investigated whether PF is associated with changes in the expression and structure of GAGs as well as in the expression of β1,3-glucuronosyltransferase I (GlcAT-I), a rate-limiting enzyme in GAG synthesis. Lung biopsies from idiopathic pulmonary fibrosis (IPF) patients and lung tissue from a rat model of bleomycin (BLM)-induced PF were immunostained for chondroitin sulfated-GAGs and GlcAT-I expression. Alterations in disaccharide composition and sulfation of chondroitin/dermatan sulfate (CS/DS) were evaluated by fluorophore-assisted carbohydrate electrophoresis (FACE) in BLM rats. Lung fibroblasts isolated from control (saline-instilled) or BLM rat lungs were assessed for GAG structure and GlcAT-I expression. Disaccharide analysis showed that 4- and 6-sulfated disaccharides were increased in the lungs and lung fibroblasts obtained from fibrotic rats compared with controls. Fibrotic lung fibroblasts and transforming growth factor-β(1) (TGF-β(1))-treated normal lung fibroblasts expressed increased amounts of hyaluronan and 4- and 6-sulfated chondroitin, and neutralizing anti-TGF-β(1) antibody diminished the same. TGF-β(1) upregulated GlcAT-I and versican expression in lung fibroblasts, and signaling through TGF-β type I receptor/p38 MAPK was required for TGF-β(1)-mediated GlcAT-I and CS-GAG expression in fibroblasts. Our data show for the first time increased expression of CS-GAGs and GlcAT-I in IPF, fibrotic rat lungs, and fibrotic lung fibroblasts. These data suggest that alterations of sulfation isomers of CS/DS and upregulation of GlcAT-I contribute to the pathological PG-GAG accumulation in PF.

Topics: Animals; Bleomycin; Cells, Cultured; Chondroitin Sulfates; Dermatan Sulfate; Disease Models, Animal; Fibroblasts; Glucuronosyltransferase; Humans; Hyaluronic Acid; Idiopathic Pulmonary Fibrosis; Lung; Male; Pulmonary Fibrosis; Rats; Rats, Sprague-Dawley; RNA, Messenger; Tissue Distribution; Transforming Growth Factor beta1; Up-Regulation

To construct a suitable ex vivo model for the research of molecular mechanisms and the pharmacological screening of fungal adherence on the corneal surface.. Mouse eyes were divided into three groups as follows: a control group with normal corneal epithelium, a group with corneal epithelium that was needle-scarified, and a group with corneal epithelium that was completely debrided. All 96 corneas were placed in organ culture and inoculated with 5 μl spore suspensions of Candida albicans at 10⁹, 10⁸, or 10⁷ colony-forming units (CFU)/ml and incubated for 0, 30, 60, or 120 min. The corneas were homogenated and diluted for quantification by counting the CFU. The effects of amphotericin B or chondroitin sulfate on the adherence of the fungal spores were evaluated with the ex vivo organ culture model and were also compared with the human corneal epithelium monolayer model in vitro.. Compared with the normal corneas with intact epithelium, the corneas with scarified and debrided epithelium adhered more spores for above two and four folds. The spore adhesion on the corneal surface was in an inoculation concentration- and incubation time-dependent manner. Moreover, both amphotericin B and chondroitin sulfate inhibited the adhesion of C. albicans spores on the corneal surface, but the inhibitory rates were different between the ex vivo corneal organ culture model and the in vitro corneal epithelium monolayer model.. The corneal organ culture was a suitable ex vivo model for the research of fungal adhesion mechanisms and drug screening.

Topics: Amphotericin B; Animals; Bacterial Adhesion; Candida albicans; Candidiasis; Chondroitin Sulfates; Colony Count, Microbial; Cornea; Corneal Injuries; Corneal Ulcer; Disease Models, Animal; Eye Infections, Fungal; Mice; Mice, Inbred BALB C; Organ Culture Techniques; Time Factors

Advancement in tissue engineering provides a promising approach to recover the functionality of the degenerated intervertebral disc. In our study, a nucleus pulposus (NP) cell-seeded collagen II/hyaluronan/chondroitin-6-sulfate (CII/HyA/CS) tri-copolymer construct was implanted into the disc space directly after nucleotomy in a rabbit model.. The aim of this study was to investigate whether the NP cell-seeded CII/HyA/CS tri-copolymer constructs could regenerate the degenerated disc in vivo after implantation into the rabbit nucleotomy model.. Nucleotomy is one of the most prevalent surgical modalities to treat degenerative disc disease, which could achieve good short-term effects of pain relieve, whereas removal of the entire or partial NP changes the biomechanical characteristics of the remaining disc and the adjacent vertebral segments and a series of long-term complications such as accelerated annulus and the facet joints degeneration may ensue. Therefore, it is necessary to think about possible procedures immediately after the primary nucleotomy surgery to avoid these complications.. NP cells isolated from thoracic and lumbar spines of New Zealand White rabbits of approximately 3 weeks of age and 1 kg in weight were labeled with a 5- (and-6) -carboxyflurescein diacetate succinimidyl ester (CFDA-SE) fluorescent dye and seeded within the CII/HyA/CS scaffold by a centrifugation method. After in vitro culture for 1 week, NP cell-seeded CII/HyA/CS tri-copolymer constructs were allografted into the disc defects of recipient rabbit immediately after nucleotomy of the lumbar spine. The Bradner Disc Index and the T2-weighted signal intensity index were determined using lateral plane radiographs and magnetic resonance imaging at 4, 12, and 24 weeks after the operation. Finally, the operated discs were explanted for gross morphological observation, histological evaluation, and cell viability assessment. Animals with only nucleotomy and cell-free CII/HyA/CS scaffold implantation served as controls.. In our study, we could demonstrate that the T2-weighted signal intensity index of the operated discs decreased in all three groups 1 month after surgery and the index of the cell-containing scaffold insertion group was significantly higher than that of the other two groups. After 24 weeks, the index of the cell-containing scaffold insertion group increased significantly. However, further decline was observed in both the noninsertion group and the scaffold insertion group. In radiographic analysis, the narrowing of the intervertebral disc space was significantly retarded by the cell-scaffold hybrids implantation up to 24 postoperative weeks. Furthermore, the gross morphology and histological evaluation indicated that the allografted NP cells were viable and showed extracellular matrix production.. In our study, we had constructed rabbit NP cell-seeded CII/HyA/CS tri-copolymer implants in vitro. Immediately after nucleotomy of the recipient rabbit, we allografted the precultured cell-scaffold hybrids into the lacuna of the disc. Results documented survival of the allografted NP cells and extracellular matrix deposition, which finally resulted in maintenance of disc height and restoration of T2-weighted signal intensity on magnetic resonance imaging.

Topics: Animals; Cell Culture Techniques; Cell Survival; Cell Transplantation; Cells, Cultured; Chondroitin Sulfates; Collagen Type II; Disease Models, Animal; Female; Graft Survival; Hyaluronic Acid; Intervertebral Disc; Intervertebral Disc Degeneration; Lumbar Vertebrae; Magnetic Resonance Imaging; Male; Polymers; Rabbits; Radiography; Regeneration; Tissue Engineering; Tissue Scaffolds; Transplantation, Homologous; Treatment Outcome

During 2007-2008, serious adverse events were reported following iv administration of certain batches of commercially available heparin in humans. Anaphylactoid reactions with acute hypotension were the hallmark of these cases. Subsequently, it was shown that a contaminant, oversulfated chondroitin sulfate (OSCS), was responsible for these adverse events. The present study was undertaken to further elucidate the risks related to OSCS-contaminated heparin preparations. Using an anesthetized rat hemodynamic model, marked diastolic blood pressure drops were induced with a single iv injection of a contaminated heparin (1000 IU/kg; 34% wt/wt OSCS). OSCS alone (0.8 and 20 mg/kg) or in combination (0.8-1.7 mg/kg) with uncontaminated heparin produced a similar hypotensive effect, whereas heparin spiked with 0.2 or 0.4 mg/kg OSCS produced no hemodynamic changes. In conscious rats, acute hypotensive effects were seen following single iv administration of OSCS-spiked heparin (1.7 or 3.0 mg/kg). Conversely, no hemodynamic effects were observed with same doses when administered sc. Pretreatment with a bradykinin-2 receptor antagonist (HOE140) fully abolished the hypotensive response after iv OSCS (1.7 mg/kg) administration, whereas pretreatment with the histamine (H1) receptor antagonist cetirizine did not. In vitro, OSCS (25 and 250 μg/ml) induced a robust, dose-related increase in kallikrein activity in rat and human plasma with a lower amplitude of response in dog and pig. The data suggest that the adverse events associated with OSCS-contaminated heparin are dependent upon the concentration of contaminant and its route of administration. Furthermore, the kallikrein-kinin system plays a pivotal role in the initiation of OSCS-related vascular effects.

Topics: Anaphylaxis; Animals; Bradykinin; Chondroitin Sulfates; Disease Models, Animal; Dogs; Dose-Response Relationship, Drug; Drug Contamination; Female; Heparin; Humans; Hypotension; Kallikrein-Kinin System; Male; Rats; Rats, Sprague-Dawley; Swine

Cisplatin (CDDP) is an anticancer drug. The clinical limitations associated with CDDP have stimulated the development of macromolecular drug-carrier systems, in attempts to decrease its toxicity. A complex (CDDP-CSA-23) between CDDP and chondroitin sulfate (CSA), a natural polysaccharide with a mean molecular weight of 23 kDa, proved to have the same anticancer activity as CDDP. A toxicodynamic study was performed on perfused kidneys to determine the effect of CDDP-CSA-23 on renal functions and the extent of platinum accumulation. The results showed that CDDP-CSA-23 attenuates the reduction in urine flow and creatinine clearance induced by CDDP. Moreover, significantly lower amounts of platinum were excreted into the urine in the case of CDDP-CSA-23, compared with CDDP alone. Meanwhile, CDDP-CSA-23 effectively retarded the rapid perfusion of platinum into kidney tissues, as occurs when CDDP is being perfused alone. The cytoprotective effects of CDDP-CSA on human proximal tubular (HK-2) cells were examined by measuring the growth of HK-2 cells in the presence of CDDP or CDDP-CSA-23. Interestingly, CDDP-CSA-23 was found to have a significantly reduced cytotoxicity, compared to CDDP. These results suggest that CDDP-CSA-23 greatly decreased the negative effects of CDDP on glomerular filtration and tubular transport in kidneys at early stages of its administration.

Topics: Animals; Antineoplastic Agents; Cells, Cultured; Chondroitin Sulfates; Cisplatin; Creatinine; Disease Models, Animal; Disease Progression; Drug Combinations; Glomerular Filtration Rate; Humans; Kidney Diseases; Kidney Tubules, Proximal; Male; Neoplasms, Experimental; Perfusion; Rats, Sprague-Dawley

Rodent models of osteoarthritis and rheumatoid arthritis are useful tools to study these disease processes. Adjuvant arthritis (AAR) is a model of polyarthritis widely used for preclinical testing of antiarthritis substances. We report the effect of two different doses of highly purified chondroitin sulfate (CS) pharmaceutical grade in the AAR animal model after oral administration.. AAR was induced by a single intradermal injection of heat-inactivated Mycobacterium butyricum in incomplete Freund's adjuvant. The experiments included healthy animals, untreated arthritic animals, arthritic animals having been administered 300 or 900 mg/kg of CS daily, 14 days before AAR induction until the end of the experiment (day 28), arthritic animals having been administered 300 or 900 mg/kg of CS daily, from day 1 until the end of the experiment.. CS was capable of significantly reducing the severity of arthritis along with oxidative stress, a consequence of chronic inflammatory processes occurring in AAR. The CS pre-treatment regimen was effective throughout the whole subacute phase, while treatment from day 1 proved effective only in the chronic period. The effects were confirmed by improved total antioxidant status and γ-glutamyltransferase activity. CS administered under a pre-treatment regimen was also able to reduce the production of pro-inflammatory cytokines, C-reactive protein in plasma, phagocytic activity and the intracellular oxidative burst of neutrophils.. CS proved to be effective in slowing down AAR development and in reducing disease markers, thus supporting its beneficial activity as a drug in humans.

Topics: Animals; Antioxidants; Arthritis, Experimental; C-Reactive Protein; Chondroitin Sulfates; Cytokines; Disease Models, Animal; gamma-Glutamyltransferase; Hindlimb; Male; Neutrophils; Oxidative Stress; Phagocytes; Rats; Rats, Inbred Lew

To evaluate the effect of topically administered pigment epithelium-derived factor (PEDF) on experimentally induced corneal neovascularization (NV) in a rat model.. Corneal chemical cauterization was induced in the left eye by using silver nitrate sticks in 160 anesthetized male Sprague-Dawley rats. The rats were randomly assigned to 4 groups with 40 rats each for topical administration of recombinant PEDF, chloramphenicol, chondroitin sulfate, and normal saline (as control). At different intervals (3, 10, 15, and 30 days) of the treatment, rats were euthanized and the corneas removed for immunohistochemistry and Western blot analyses to measure expression levels of PEDF, vascular endothelial growth factor (VEGF), and CD34, an endothelial maker. The right eyes were used as normal control.. There were high levels of PEDF expression and low levels of VEGF and CD34 in the normal cornea. VEGF and CD34 levels were significantly induced by chemical cauterization in the groups treated with chloramphenicol, chondroitin sulfate, and normal saline, demonstrating corneal NV. The VEGF and CD34 levels reached a plateau in the cornea on the 10th day after cauterization and remained at high levels thereafter. In contrast, the PEDF treatment prevented the overexpression of VEGF and CD34 induced by the cauterization.. PEDF downregulates VEGF expression and inhibits corneal NV induced by chemical cauterization. The results suggested that PEDF has therapeutic potential for corneal neovascular diseases.

Topics: Administration, Topical; Animals; Antigens, CD34; Blotting, Western; Cautery; Chloramphenicol; Chondroitin Sulfates; Corneal Neovascularization; Disease Models, Animal; Eye Proteins; Immunohistochemistry; Male; Nerve Growth Factors; Rats; Rats, Sprague-Dawley; Recombinant Proteins; Serpins; Silver Nitrate; Vascular Endothelial Growth Factor A

Familial Amyloidotic Polyneuropathy (FAP) is a disorder characterized by the extracellular deposition of fibrillar Transthyretin (TTR) amyloid, with a special involvement of the peripheral nerve. We had previously shown that doxycycline administered for 3 months at 40 mg/Kg/ml in the drinking water, was capable of removing TTR amyloid deposits present in stomachs of old TTR-V30M transgenic mice; the removal was accompanied by a decrease in extracellular matrix remodeling proteins that accompany fibrillar deposition, but not of non-fibrillar TTR deposition and/or markers associated with pre-fibrillar deposits. On the other hand, Tauroursodeoxycholic acid (TUDCA), a biliary acid, administrated to the same mouse model was shown to be effective at lowering deposited non-fibrillar TTR, as well as the levels of markers associated with pre-fibrillar TTR, but only at young ages. In the present work we evaluated different doxycycline administration schemes, including different periods of treatment, different dosages and different FAP TTR V30M animal models. Evaluation included CR staining, immunohistochemistry for TTR, metalloproteinase 9 (MMP-9) and serum amyloid P component (SAP). We determined that a minimum period of 15 days of treatment with a 8 mg/Kg/day dosage resulted in fibril removal. The possibility of intermittent treatments was also assessed and a maximum period of 15 days of suspension was determined to maintain tissues amyloid-free. Combined cycled doxycycline and TUDCA administration to mice with amyloid deposition, using two different concentrations of both drugs, was more effective than either individual doxycycline or TUDCA, in significantly lowering TTR deposition and associated tissue markers. The observed synergistic effect of doxycycline/TUDCA in the range of human tolerable quantities, in the transgenic TTR mice models prompts their application in FAP, particularly in the early stages of disease.

Topics: Amyloid Neuropathies, Familial; Animals; Biglycan; Biomarkers; Blotting, Western; Chondroitin Sulfates; Disease Models, Animal; Dose-Response Relationship, Drug; Doxycycline; Drug Administration Schedule; Drug Synergism; Endoplasmic Reticulum Chaperone BiP; Extracellular Matrix Proteins; Heat-Shock Proteins; Immunohistochemistry; Matrix Metalloproteinase 9; Mice; Prealbumin; Proteoglycans; Stomach; Taurochenodeoxycholic Acid

Pregnancy-associated malaria (PAM) is a serious consequence of the adhesion to the placental receptor chondroitin sulfate A (CSA) of Plasmodium falciparum-infected erythrocytes (PE) expressing the large cysteine-rich multi-domain protein var2CSA. Women become resistant to PAM, and develop strain-transcending immunity against CSA-binding parasites. The identification of var2CSA regions that could elicit broadly neutralizing and adhesion-blocking antibodies is a key step for the design of prophylactic vaccine strategies.. Escherichia coli expressed var2CSA DBL domains were refolded and purified prior to immunization of mice and a goat. Protein-G-purified antibodies were tested for their ability to block FCR3(CSA)-infected erythrocytes binding to placental (BeWo) and monkey brain endothelial (ScC2) cell lines using a flow cytoadhesion inhibition assay mimicking closely the physiological conditions present in the placenta at shear stress of 0.05 Pa. DBL5-ε, DBL6-ε and DBL5-6-ε induced cross-reactive antibodies using Alum and Freund as adjuvants, which blocked cytoadhesion at values ranging between 40 to 96% at 0.5 mg IgG per ml. Importantly, antibodies raised against recombinant DBL5-ε from 3 distinct parasites genotypes (HB3, Dd2 and 7G8) showed strain-transcending inhibition ranging from 38 to 64% for the heterologuous FCR3(CSA).. Using single and double DBL domains from var2CSA and Alum as adjuvant, we identified recombinant subunits inducing an immune response in experimental animals which is able to block efficiently parasite adhesion in a flow cytoadhesion assay that mimics closely the erythrocyte flow in the placenta. These subunits show promising features for inclusion into a vaccine aiming to protect against PAM.

Topics: Animals; Antibodies, Protozoan; Antigens, Protozoan; Cell Adhesion; Cell Line; Chondroitin Sulfates; Disease Models, Animal; Female; Flow Cytometry; Humans; Malaria; Mice; Mice, Inbred BALB C; Placenta; Plasmodium falciparum; Pregnancy; Pregnancy Complications, Parasitic; Protein Structure, Tertiary; Recombinant Proteins; Saimiri; Species Specificity

Evidence that combined glucosamine sulfate and chondroitin sulfate (Gluchon) or isolated glucosamine (Glu) modifies joint damage in osteoarthritis (OA) is still lacking. We studied joint pain and cartilage damage using the anterior cruciate ligament transection (ACLT) model. Wistar rats were subjected to ACLT of the right knee (OA) or sham operation. Groups received either Glu (500 mg/kg), Gluchon (500 mg/kg glucosamine +400 mg/kg chondroitin) or vehicle (non-treated--NT) per os starting 7 days prior to ACLT until sacrifice at 70 days. Joint pain was evaluated daily using the rat-knee joint articular incapacitation test. Structural joint damage was assessed using histology and biochemistry as the chondroitin sulfate (CS) content of cartilage by densitometry (microgram per milligram dried cartilage), comparing to standard CS. The molar weight (Mw) of the CS samples, used as a qualitative biochemical parameter, was obtained by comparing their relative mobility on a polyacrylamide gel electrophoresis to standard CS. Gluchon, but not Glu, significantly reduced joint pain (P < 0.05) compared to NT. There was an increase in CS content in the OA group (77.7 +/- 8.3 microg/mg) compared to sham (53.5 +/- 11.2 microg/mg) (P < 0.05). The CS from OA samples had higher Mw (4:62 +/- 10(4) g/mol) compared to sham (4:18 +/- 0.19 x 10(4) g/mol) (P < 0.05). Gluchon administration significantly reversed both the increases in CS content (54.4 +/- 12.1 microg/mg) and Mw (4:18 +/- 0.2 x 10(4) g/mol) as compared to NT. Isolated Glu decreased CS content though not reaching statistical significance. Cartilage histology alterations were also significantly prevented by Gluchon administration. Gluchon provides clinical (analgesia) and structural benefits in the ACLT model. This is the first demonstration that biochemical alterations occurring in parallel to histological damage in OA are prevented by Gluchon administration.

Topics: Animals; Anterior Cruciate Ligament; Arthralgia; Cartilage, Articular; Chondroitin Sulfates; Disease Models, Animal; Drug Therapy, Combination; Glucosamine; Male; Osteoarthritis, Knee; Rats; Rats, Wistar

To determine the short- and long-term changes in the biomechanical properties and metabolic activity of articular cartilage following the remote application of bipolar radiofrequency (bRF) and monopolar radiofrequency (mRF) energy within the rabbit stifle joint.. The rabbits were randomly assigned to either Group-1 (normal rabbit food), or they were assigned to Group-2 (2% Cosequin in the diet). Each rabbit underwent bilateral stifle arthroscopy with either bRF or mRF applied to the infrapatellar fat pad for 45 seconds. Cartilage samples were collected at zero, four, and 14 weeks after surgery. Data were analyzed with a mixed model analysis of variance (ANOVA) for chondrocyte death, amount of GAG synthesis, and the equilibrium compressive modulus.. A significant increase in histological damage was noted at weeks four and 14 compared to week zero. Most of the chondrocyte death noted with confocal laser microscopy (49 of 56 samples) was noted in the superficial region (outer 25%) of the articular cartilage. GAG synthesis was not significantly different between groups or devices at any time point. A significant difference was not noted in equilibrium compressive modulus throughout the study.. Remote application of bRF and mRF energy lead to immediate chondrocyte death. Most of the damage was superficial hence the metabolic activity and biomechanical properties of the extracellular matrix were maintained throughout this study. Treatment with Cosequin did not prevent superficial chondrocyte death caused by the application of radiofrequency (RF) energy with in the joint.

Topics: Analysis of Variance; Animal Feed; Animals; Arthroscopy; Cartilage, Articular; Catheter Ablation; Cell Survival; Chondrocytes; Chondroitin Sulfates; Disease Models, Animal; Glycosaminoglycans; Rabbits; Radiation Injuries, Experimental; Random Allocation; Stifle

The use of engineered tissue for the treatment of a variety of acute to chronic wounds has become a clinical standard, and a better understanding of the cellular mechanisms of re-vascularization and barrier integrity could enhance clinical outcomes. Here, we focus on the characterization of the re-vascularization of acellular grafts such as Integra in an animal model to better understand the physiological properties of blood vessels growing in the collagen-glycosaminoglycan matrix vs. wound margins. While Integra has been extensively studied in pre-clinical models, the re-modeling mechanisms of the capillary bed under these matrices are not well understood. Therefore, our first objective was to quantify the kinetics of re-vascularization. The second objective was to assess changes in vascular permeability (VP) of the wound bed compared to normal adjacent skin. The third objective was to establish a non-invasive and quantitative assay for the measurement of VP to facilitate the rapid and reproducible characterization of vascular integrity. Using an excisional wound model in mice, we characterize the appearance, growth, and maturation of blood vessels in an Integra graft over 28 days after surgery. Initial appearance of blood vessels in the graft was observed at 7 days, with angiogenesis peaking between 7 and 14 days. The onset of VP coincided with the increase in re-vascularization of the wound bed and there was a sustained elevation of VP that declined to baseline by 28 days. We propose a non-invasive strategy to assess VP of the wound capillary bed will facilitate a better understanding of the cell and molecular basis of angiogenesis in wound healing.

Topics: Animals; Capillary Permeability; Chondroitin Sulfates; Collagen; Disease Models, Animal; Graft Survival; Granulation Tissue; Male; Mice; Mice, Inbred C57BL; Neovascularization, Physiologic; Postoperative Period; Skin; Skin Transplantation; Skin, Artificial; Wound Healing

The degeneration of intervertebral disc (IVD) is a major cause of low back pain. However, there is no satisfactory preventive treatment for degenerative disc disease (DDD). In this study, we examined the effects of a novel cross-linked hyaluronate hydrogel and cross-linked chondroitin sulfate (CS) hydrogel on a rabbit model of IVD injury. We injected 300 microl of phosphate buffer saline, 1% sodium hyaluronate, cross-linked hyaluronate hydrogel, or cross-linked CS hydrogel into the injured IVDs. One, three or six months after treatment, the whole spinal columns were dissected and magnetic resonance (MR) images of the IVDs were examined. It was noted that the IVD, which was injected with cross-linked hyaluronate hydrogel or cross-linked CS hydrogel mostly retained the normal signal intensity of the MR images. These IVDs exhibited a higher degree of staining with safranin-O than the control discs or 1% sodium hyaluronate-injected discs, suggesting that the intradiscal application of cross-linked hyaluronate hydrogel or cross-linked CS hydrogel probably inhibits the degenerative cascade of the DDD. The intradiscal administration of these drugs is safe, easy and costs less. In the near future, these intradiscal injections may become the standard therapy for the treatment of DDD instead of the spine surgeries.

Topics: Alginates; Animals; Chondroitin Sulfates; Cross-Linking Reagents; Disease Models, Animal; Humans; Hyaluronic Acid; Intervertebral Disc; Intervertebral Disc Displacement; Rabbits; Treatment Outcome

Among the agents employed to manage osteoarthritis, chondroitin sulphate (CS) is a natural glycosaminoglycan with an anti-inflammatory effect on joint cells. CS might also influence the inflammatory component of atherosclerosis. Our aim was to examine the effect of CS administration on vascular injury and on markers of systemic inflammation in a rabbit model of atherosclerosis aggravated by systemic inflammation provoked by chronic antigen-induced arthritis.. Atherosclerosis was induced in rabbits by maintaining them on a hyperlipidaemic diet after producing an endothelial lesion in the femoral arteries. Simultaneously, chronic arthritis was induced in these animals by repeated intraarticular injections of ovalbumin in previously immunized rabbits. A group of these rabbits were treated prophylactically with CS (100 mg kg(-1)day(-1)) and when the animals were killed, serum and peripheral blood mononuclear cells (PBMC) were isolated. Furthermore, femoral arteries and thoracic aorta were used for gene expression studies and histological examination.. CS administration reduced the concentration of the proinflammatory molecules C-reactive protein and IL-6 in serum. Likewise, CS inhibited the expression of CCL2/monocyte chemoattractant protein (MCP)-1 and cyclooxygenase (COX)-2 in PBMC, and reduced the nuclear translocation of nuclear factor-kappaB. In the femoral lesion, CS also diminished the expression of CCL2 and COX-2, as well as the ratio of the intima/media thickness. Moreover, CS decreased the percentage of rabbits with atherosclerosis and chronic arthritis that developed vascular lesions in the aorta.. These findings suggest that CS treatment may to some extent impede the progression of atherosclerosis.

Topics: Animals; Anti-Inflammatory Agents; Aorta; Arthritis, Experimental; Atherosclerosis; C-Reactive Protein; Chemokine CCL2; Chondroitin Sulfates; Cyclooxygenase 2; Disease Models, Animal; Disease Progression; Gene Expression Regulation; Inflammation; Interleukin-6; Male; NF-kappa B; Rabbits

Whereas the alterations of diverse tissues in cellular and molecular levels have been investigated during leg lengthening via microscopy and biochemical studies, little is known about the response of deep fascia. This study aims to investigate the changes of the extracellular matrix in deep fascia in response to leg lengthening.. Animal model of leg lengthening was established in New Zealand white rabbits. Distraction was initiated at a rate of 1 mm/day and 2 mm/day in two steps, and preceded until increases of 10% and 20% in the initial length of tibia had been achieved. Alcian blue stain and picrosirius-polarization method were used for the study of the extracellular matrix of deep fascia samples. Leica DM LA image analysis system was used to investigate the quantitative changes of collagen type I and III.. Alcian blue stain showed that glycosaminoglycans of fascia of each group were composed of chondroitin sulphate and heparin sulphate, but not of keratan sulphate. Under the polarization microscopy, the fascia consisted mainly of collagen type I. After leg lengthening, the percentage of collagen type III increased. The most similar collagen composition of the fascia to that of the normal fascia was detected at a 20% increase in tibia length achieved via a distraction rate of 1 mm/d.. The changes in collagen distribution and composition occur in deep fascia during leg lengthening. Although different lengthening schemes resulted in varied matrix changes, the most comparable collagen composition to be demonstrated under the scheme of a distraction rate of 1 mm/day and 20% increase in tibia length. Efficient fascia regeneration is initiated only in certain combinations of the leg load parameters including appropriate intensity and duration time, e.g., either low density distraction that persist a relatively short time or high distraction rates.

Topics: Alcian Blue; Animals; Biomarkers; Bone Lengthening; Chondroitin Sulfates; Collagen Type I; Collagen Type III; Coloring Agents; Disease Models, Animal; Extracellular Matrix; Fascia; Fasciotomy; Heparin; Hindlimb; Image Processing, Computer-Assisted; Microscopy, Polarization; Proteoglycans; Rabbits; Stress, Mechanical

To determine whether chondroitin sulphate (CS) impedes the migration of primary articular chondrocytes.. Articular chondrocytes were isolated from young and skeletally mature bovine animals. Boyden chambers were used to quantify chondrocyte migration on aggrecan in the presence and absence of CS chains. A novel in vitro model of cell migration into articular cartilage explants was designed to visualise and quantify the migration of labelled chondrocytes into cartilage matrix which had been treated with chondroitinase ABC to remove CS chains present.. A consistent trend of increased migration with both age groups of a sub-population of chondrocytes was demonstrated on aggrecan in the absence of CS. These data were supported by results from the in vitro model of chondrocyte migration which demonstrated increasing numbers of a chondrocyte sub-population from both age groups of cartilage migrating into the chondroitinase ABC digested cartilage explants with time in culture. Minimal migration of these chondrocytes was demonstrated into phosphate buffered saline (PBS) treated control explants.. We confirm that a sub-population of chondrocytes isolated from both young and skeletally mature articular cartilages have the ability to migrate. We also demonstrate that CS chains inhibit the migration of these articular chondrocytes and that their removal by chondroitinase ABC digestion enhances the migration of these chondrocytes. Such findings may provide a clinical application for improving cell-based cartilage repair strategies by enhancing integration between endogenous and repair tissue.

Topics: Animals; Cartilage, Articular; Cattle; Cell Movement; Chondrocytes; Chondroitin Sulfates; Disease Models, Animal; Microscopy, Fluorescence; Wound Healing

Activation of nuclear factor kappaB (NF-kappaB) and caspases may greatly amplify inflammation and cell damage in addition to that directly exerted by free radicals. Since reactive oxygen species (ROS) are involved in acute pancreatitis, we studied whether the administration of chondroitin-4-sulphate (C4S), in addition to its antioxidant activity, was able to modulate NF-kappaB and caspase activation in an experimental model of caerulein-induced acute pancreatitis in mice. Hyperstimulating doses of caerulein (50 microg/ kg), five injections per mouse given at hourly intervals produced the following: high serum lipase and amylase activity; lipid peroxidation, evaluated by 8-isoprostane concentrations; loss of antioxidant defenses such as glutathione reductase (GR) activity; NF-kappaB activation and loss of cytoplasmic IkappaBalpha protein; increases in tumor necrosis factor-alpha (TNF-alpha), interleukin-6 (IL-6), caspase-3, and caspase-7 gene expression and their related protein; accumulation and activation of neutrophils in the damaged tissue, evaluated by elastase (ELA) determination; and pancreatic injury, evaluated by histologic analysis. Pretreatment of mice with different doses of C4S, given 1 hr before caerulein injections and 1 and 2 hrs after the last caerulein injection, reduced lipid peroxidation, inhibited NF-kappaB translocation and cytoplasmic IkappaBalpha protein loss, decreased TNF-alpha, IL-6, and caspase gene expression and their related protein levels, limited endogenous antioxidant depletion, and reduced tissue neutrophils accumulation and tissue damage. Since molecules with antioxidant activity can block NF-kappaB and apoptosis activation, we suggest that C4S administration is able to block NF-kappaB and caspase activation by reducing the oxidative burst.

Topics: Animals; Apoptosis; Ceruletide; Chondroitin Sulfates; Disease Models, Animal; Gene Expression Regulation, Enzymologic; Glutathione Reductase; Interleukin-6; Lipase; Male; Mice; NF-kappa B; Oxidative Stress; Pancreatitis; Tumor Necrosis Factor-alpha

Heparin is an excellent inhibitor of P- and L-selectin binding to the carbohydrate determinant, sialyl Lewis(x). As a consequence of its anti-selectin activity, heparin attenuates metastasis and inflammation. Here we show that fucosylated chondroitin sulfate (FucCS), a polysaccharide isolated from sea cucumber composed of a chondroitin sulfate backbone substituted at the 3-position of the beta-D-glucuronic acid residues with 2,4-disulfated alpha-L-fucopyranosyl branches, is a potent inhibitor of P- and L-selectin binding to immobilized sialyl Lewis(x) and LS180 carcinoma cell attachment to immobilized P- and L-selectins. Inhibition occurs in a concentration-dependent manner. Furthermore, FucCS was 4-8-fold more potent than heparin in the inhibition of the P- and L-selectin-sialyl Lewis(x) interactions. No inhibition of E-selectin was observed. FucCS also inhibited lung colonization by adenocarcinoma MC-38 cells in an experimental metastasis model in mice, as well as neutrophil recruitment in two models of inflammation (thioglycollate-induced peritonitis and lipopolysaccharide-induced lung inflammation). Inhibition occurred at a dose that produces no significant change in plasma activated partial thromboplastin time. Removal of the sulfated fucose branches on the FucCS abolished the inhibitory effect in vitro and in vivo. Overall, the results suggest that invertebrate FucCS may be a potential alternative to heparin for blocking metastasis and inflammatory reactions without the undesirable side effects of anticoagulant heparin.

Topics: Adenocarcinoma; Animals; Anticoagulants; Carbohydrate Conformation; Cell Adhesion; Chondroitin Sulfates; Disease Models, Animal; Dose-Response Relationship, Drug; Heparin; L-Selectin; Lipopolysaccharides; Lung Neoplasms; Mice; Neoplasm Metastasis; Neoplasms, Experimental; Neutrophil Infiltration; P-Selectin; Partial Thromboplastin Time; Peritonitis; Pneumonia; Sea Cucumbers; Thioglycolates

In juvenile tree shrews, positioning a negative-power lens in front of an eye produces a hyperopic shift in refractive state and causes a compensatory increase in axial length over several days so that the eye is myopic when the lens is removed. During negative lens compensation, the scleral extracellular matrix is remodeled. A biomechanical property of the sclera, creep rate, increases; during recovery from induced myopia, the creep rate decreases below normal levels. Changes in glycosaminoglycan (GAG) levels, including those of hyaluronan, may participate in these changes in creep rate and, in turn, participate in controlling the axial length and refractive state. This study investigated the unsulfated and sulfated GAG composition of the sclera during compensation for a -5 diopter (D) lens and during recovery.. Capillary electrophoresis was used to assess the relative levels (ng/mg dry scleral weight) of unsulfated GAGs (hyaluronan [HA] and chondroitin [C0S]), sulfated GAGs (chondroitin-4-sulfate [C4S], chondroitin-6-sulfate [C6S], and dermatan sulfate [DS]) in the sclera of groups of tree shrews (n = 5 per group) that wore a monocular -5 D lens for 1, 2, 4, or 11 days or had 11 days of -5 D lens wear followed by 1, 2, or 4 days of recovery from lens wear. The fellow eye served as an untreated control. Groups of normal and plano lens-treated animals provided age-matched values.. Expressed as a fraction of dry weight, levels of HA were lower after 1, 4, and 11 days of -5 D lens wear. Levels of C0S, C6S, and C4S were significantly lower after 4 and 11 days of lens wear. After 1 and 2 days of recovery, GAG levels in the treated eyes were not significantly different from those in control eyes. After 4 recovery days, HA levels were lower, but the levels of all other GAGs were not different in the recovering and control eyes. Some binocular changes also occurred.. The rapid differential decrease in HA levels during negative lens compensation and the absence of any difference after just 1 day of recovery suggest that HA levels may play a previously unrecognized early role in regulating the biomechanical property (creep rate) of the sclera. The reduced levels of the other GAGs, which occur when creep rate is at its peak elevation, and their rapid return to normal after 1 day of recovery suggest that they may also participate in regulating this biomechanical property of the sclera.

Topics: Animals; Chondroitin; Chondroitin Sulfates; Dermatan Sulfate; Disease Models, Animal; Electrophoresis, Capillary; Female; Hyaluronic Acid; Male; Myopia; Organ Size; Sclera; Sensory Deprivation; Tupaiidae

In experimental acute pancreatitis, a large amount of reactive oxygen species are produced, and in turn cytoskeletal changes may be induced in pancreatic tissue. These changes contribute to an imbalance of digestive enzyme segregation, transport, exocytosis and activation, resulting in cell injury. In this study, we assessed the effects of chondroitin sulfate (CS) on attenuation of oxidative damage and protection of F-actin in rats with acute necrotizing pancreatitis (ANP).. Ninety male Wistar rats were divided randomly into three groups. Group A was infused with 5% sodium taurocholate; group B was treated with CS; and group C served as control. Rats from the three groups were killed at 1, 3 or 8 hours. The levels were measured of malonyl dialdehyde (MDA), total superoxide dismutase (SOD), glutathione synthetase (GSH), serum amylase (SAM) and adenosine triphosphate (ATP). F-actin immunostained with rhodamine-phalloidin was analyzed using a confocal laser scanning system and the content of F-actin protein was determined.. The levels of SAM increased in groups A and B, whereas the levels of GSH, SOD and ATP in group A decreased markedly during pancreatitis, and MDA increased significantly. The levels of GSH, SOD and ATP in group B were higher than those in group A, but the level of MDA was lower than in group A. At the same time, ANP resulted in early disruption of the cytoskeleton with dramatic changes and a loss of F-actin. Administration of CS moderated the damage to the actin cytoskeleton.. Retrograde infusion of sodium taurocholate via the pancreatic duct may produce pancreatic necrosis and a marked increase in serum amylase activity, induce a severe depletion of ATP level, prime lipid peroxidation, and damage F-actin. Treatment with CS can ameliorate pancreatic cell conditions, limit cell membrane peroxidation, protect F-actin, and attenuate pancreatitis.

Topics: Actins; Amylases; Animals; Antioxidants; Chondroitin Sulfates; Colorimetry; Cytoskeleton; Disease Models, Animal; Lipid Peroxidation; Male; Malondialdehyde; Microscopy, Confocal; Pancreatitis, Acute Necrotizing; Rats; Rats, Wistar; Treatment Outcome

Intraperitoneally administered chondroitin sulfate E inhibited the development of antibody-induced arthritis, a model of rheumatoid arthritis, while chondroitin 4-sulfate showed no effects. Chondroitin sulfate E inhibited in vitro differentiation of osteoclasts, which play key roles in the etiology of rheumatoid arthritis. One of the targets of chondroitin sulfate E is midkine, a heparin-binding growth factor or cytokine. Indeed, a chimeric-type siRNA for midkine inhibited the development of antibody-induced arthritis and adhesion of the omentum to the injured abdominal wall. These results indicate the significance of midkine as a molecular target to treat or prevent rheumatoid arthritis and adhesion after surgery, and the utility of chondroitin sulfate E to inhibit midkine in vivo.

Topics: Animals; Antibodies; Arthritis, Rheumatoid; Cell Differentiation; Chondroitin Sulfates; Cytokines; Disease Models, Animal; Female; Injections, Intraperitoneal; Mice; Mice, Inbred BALB C; Midkine; Osteoblasts; RNA, Small Interfering

Mutations in the SLC26A2 cause a family of recessive chondrodysplasias that includes in order of decreasing severity achondrogenesis 1B, atelosteogenesis 2, diastrophic dysplasia and recessive multiple epiphyseal dysplasia. The gene encodes for a widely distributed sulfate/chloride antiporter of the cell membrane whose function is crucial for the uptake of inorganic sulfate that is needed for proteoglycan sulfation. To investigate the mechanisms leading to skeletal dysplasia, we generated a transgenic mouse with a mutation in Slc26a2 causing a partial loss of function of the sulfate transporter. Homozygous mutant mice were characterized by skeletal dysplasia with chondrocytes of irregular size, delay in the formation of the secondary ossification centre and osteoporosis of long bones. Impaired sulfate uptake was demonstrated in chondrocytes, osteoblasts and fibroblasts, but proteoglycan undersulfation was detected only in cartilage. The similarity with human diastrophic dysplasia makes this mouse a model to explore pathogenetic and therapeutic aspects of SLC26A2-related disorders.

Topics: Animals; Anion Transport Proteins; Chondrocytes; Chondroitin Sulfates; Disease Models, Animal; Epiphyses; Health; Mice; Mice, Transgenic; Sulfate Transporters; Sulfates

Fucosylated chondroitin sulfate is a potent anticoagulant polysaccharide extracted from sea cucumber. Its anticoagulant activity is attributed to the presence of sulfated fucose branches. We have shown that intravascular injection of fucosylated chondroitin sulfate inhibits thrombus formation in a venous and an arterial shunt model in rats. Since this compound resists digestion by enzymes that cleave mammalian glycosaminoglycans, we investigated the possibility that fucosylated chondroitin sulfate might be absorbed after oral administration. In fact, after oral administration of fucosylated chondroitin sulfate to rats, we observed a dose-dependent increase in the plasma anticoagulant activity, as assessed by assays for activated partial thromboplastin time (aPTT) and thrombin time (TT) (about 3- and 5-fold, respectively) and by anti-IIa activity. Furthermore, animals receiving daily oral doses of this glycosaminoglycan showed a decrease in thrombus weight on experimental models of venous and arterial shunt thrombosis. This antithrombotic action clearly has a strong relationship with anticoagulant activity. Similar doses of heparin administered orally had no effect on the plasma anticoagulant activity or on the thrombus weight. Finally, we observed that fucosylated chondroitin sulfate given orally to rats did not modify the bleeding time. Overall, our results indicate that fucosylated chondroitin sulfate is absorbed after oral administration and could become a promising oral anticoagulant.

Topics: Administration, Oral; Animals; Anticoagulants; Bleeding Time; Blood Coagulation; Chondroitin Sulfates; Disease Models, Animal; Dose-Response Relationship, Drug; Fibrinolytic Agents; Heparin; Partial Thromboplastin Time; Prothrombin; Rats; Rats, Wistar; Sea Cucumbers; Thrombin Time; Thrombosis; Time Factors

To provide a less expensive and more convenient protocol for the treatment of tympanic membrane perforations (TMPs).. Several materials were prepared and compared for TMP repair including Carbylan-SX, Gelatin-DTPH-PEGDA (GX), Carbylan-S/Gelatin-DTPH (Carbylan-GSX) (injectable and sponge), Gelfoam, Epifilm, and crosslinked thiolated chondroitin sulfate (CS-DTPH-PEGDA [CS-SX]). Hartley pigmented guinea pigs (Elm Hill) underwent bilateral myringotomy with 1 ear left as a control and the other treated with one of the previously mentioned materials.. Carbylan-GSX (injectable and sponge), Gelfoam with saline, and CS-SX had the shortest time for TMP closure. Epifilm, Carbylan, and gelatin preparations resulted in closure rates similar to controls. CS-SX showed a marked inflammatory reaction compared with controls and other materials based on neutrophil, lymphocyte, epitheloid counts, and degree of fibrosis.. This study shows the validity of Carbylan-GSX compared with Gelfoam as a material to promote TMP closure in an acute TMP guinea pig model.

Topics: Animals; Chondroitin Sulfates; Disease Models, Animal; Extracellular Matrix; Gelatin; Gelatin Sponge, Absorbable; Guinea Pigs; Humans; Hyaluronic Acid; Hydrogels; Polyethylene Glycols; Tympanic Membrane Perforation

The aim of this study was to evaluate histologically the action of chondroitin sulphate in osteoarthritis experimentally induced by continuous immobilization. Fourteen young female Norfolk rabbits aged 2.5-3 months at the beginning of the experiment were divided into two equitable groups submitted to immobilization of the right knee for a period of 12 weeks. The treated group received 1.0 ml/animal/s.c. of 12% chondroitin sulphate, once a week for 12 weeks, and the untreated group did not receive any treatment. Two additional animals were not submitted to knee immobilization (sham group). Microscopical examination of knee preparations stained with haematoxylin-eosin and Masson trichrome showed lesions of both joints in treated and untreated groups, with no significant difference between the scores obtained for the right and left knees. Examination of preparations stained with picrosirius red showed collagen fibre alignment and misalignment in the right and left knees of the animals of all groups, but statistic analysis could not be performed. It was not possible to differentiate the proteoglycan concentration between limbs or groups (treated and untreated) by safranin O or toluidine blue staining. It was possible to conclude that the chondroitin sulphate was not able to reduce the histological changes induced by this osteoarthritis experimental model.

Topics: Animals; Chondroitin Sulfates; Disease Models, Animal; Female; Immobilization; Knee Joint; Osteoarthritis; Rabbits; Treatment Outcome

Stroke induces axonal sprouting in peri-infarct cortex. A set of growth-associated genes important in axonal sprouting in peripheral nervous system regeneration and cortical development has recently been defined. The expression profiles of these growth-associated genes were defined during the post-stroke axonal sprouting response using a model of stroke in barrel field cortex. Stroke induces sequential waves of neuronal growth-promoting genes during the sprouting response: an early expression peak (SPRR1), a mid expression peak (p21, Ta1 tubulin, L1, MARCKS), a late peak (SCG10, SCLIP), and an early/sustained pattern (GAP43, CAP23, c-jun). These expression peaks correspond to specific time points in the sprouting response. The expression of the growth-inhibiting chondroitin sulfate proteoglycans aggrecan, brevican, versican, and phosphacan are induced late in the sprouting process; except neurocan, which is increased during the peak of the growth-promoting gene expression. The developmentally associated growth inhibitors ephrin-A5, ephB1, semaphorin IIIa, and neuropilin 1 are also induced in the early phases of the sprouting response. At the cellular level, chondroitin sulfate proteoglycans, in the form of peri-neuronal nets, are reduced in the region of axonal sprouting, during the peak of growth-promoting gene expression. These results identify a unique profile of growth-promoting gene expression in adult cortex after stroke, the inhibitory molecules that are present during the sprouting response, and a region in which growth-promoting genes are increased, growth-inhibitory proteins are diminished and axonal sprouting occurs. This region may be a growth-promoting zone after stroke.

Topics: Aggrecans; Animals; Brain Infarction; Cerebral Cortex; Chondroitin Sulfate Proteoglycans; Chondroitin Sulfates; Disease Models, Animal; Extracellular Matrix Proteins; Gene Expression; Gene Expression Regulation; Immunohistochemistry; In Situ Hybridization; Lectins, C-Type; Male; Nerve Growth Factors; Nerve Tissue Proteins; Proteoglycans; Rats; Rats, Inbred F344; Receptor-Like Protein Tyrosine Phosphatases, Class 5; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Stroke; Time Factors; Versicans

The therapeutic effect of glucosamine hydrochloride (GH) and chondroitin sulfate (CS) in combination with fursultiamine, a vitamin B1 derivative, on the development of cartilage lesions was investigated in an animal model of osteoarthritis (OA).. The OA model was created by partial medial meniscectomy of the right knee joint (day 0). The rabbits were placed into three experimental groups: operated (OA) rabbits that received placebo treatment, OA rabbits that received GH (1000 mg/kg) + CS (800 mg/kg), and OA rabbits that received GH + CS + fursultiamine (100 mg/kg). Each treatment was initiated on day 3 and continued for 8 weeks. Macroscopic and histologic analyses were performed on the cartilage. The level of MMP-1 in OA cartilage chondrocytes was evaluated by immunohistochemistry.. Only the group receiving combined treatment with GH + CS + fursultiamine showed a significant reduction in the severity of macroscopic and histologic lesions on tibial plateau, which is the weight bearing cartilage surface of the tibia, compared with placebo-treated OA rabbits. This treatment group also revealed a small, but significant, decrease in the body weight gain of the rabbits. In cartilage from placebo-treated OA rabbits, a significantly higher percentage of chondrocytes in superficial layer stained positive for MMP-1 compared with unoperated control. Rabbits treated with the GH + CS + fursultiamine revealed a significant reduction in the level of MMP-1.. These results suggest that the chondroprotective effect of GH + CS is enhanced by the addition of fursultiamine in experimental OA. This effect was associated with a reduction in the level of MMP-1, which are known to play an important role in the pathophysiology of OA lesions.

Topics: Animals; Body Weight; Cartilage; Chondroitin Sulfates; Disease Models, Animal; Disease Progression; Fursultiamin; Glucosamine; Immunohistochemistry; Male; Matrix Metalloproteinase 1; Osteoarthritis; Protective Agents; Rabbits; Tibia

To determine whether a novel platelet-activating factor (PAF) antagonist prevents experimentally induced diffuse lamellar keratitis (DLK) after laser in situ keratomileusis (LASIK).. Department of Ophthalmology and Neuroscience Center of Excellence, Louisiana State University Health Science Center, New Orleans, Louisiana, USA.. Twenty eyes of 10 New Zealand albino rabbits were used. The left eyes were treated with a peribulbar injection of 0.5 mL of PAF receptor antagonist LAU 0901 (2,4,6-trimethyl-1,4-dihydropyridine-3,5-dicarboxylic acid ester) dissolved in 20 hydroxypropyl B cyclodextrin (30 microg /mL). Two rabbits were treated with a peribulbar injection of 0.5 mL of vehicle (cyclodextrin) alone and served as controls. A corneal flap was cut in all eyes, and the interface was exposed to Pseudomonas aeruginosa endotoxin. The left eyes were additionally treated with 1 drop of LAU 0901 4 times a day. Rabbits were killed on postoperative days 1, 2, 3, 5, and 8. The eyes were enucleated and processed for histopathology and immunohistochemical examination.. Corneas not treated with LAU 0901 and controls showed a severe inflammatory response in the flap margin and stromal interface, characterized by loss of keratocytes, activation of adjacent keratocytes and transformation to myofibroblasts, infiltration of polymorphonuclear leukocytes and monocytes, and presence of epithelial cells with necrosis and melting of adjacent stroma. Corneas of rabbits treated with LAU 0901 showed minimal loss of keratocytes and myofibroblast transformation, minimal inflammatory cell infiltration, and minimal presence of epithelial cells in the interface.. Induction of DLK was blocked by a PAF receptor antagonist in rabbit eyes. The histopathological evaluation and immunohistochemical studies showed that treatment with LAU 0901 blocked keratocyte apoptosis, transformation of fibroblasts to myofibroblasts and migration to the wound site, and chemotaxis of inflammatory cells, inhibiting the inflammatory response and promoting adequate healing of the flap interface and adjacent stroma.

Topics: Actins; Animals; Apoptosis; Cell Differentiation; Cell Movement; Cells, Cultured; Chondroitin Sulfates; Cornea; Dihydropyridines; Disease Models, Animal; Endotoxins; Fibroblasts; Fluorescent Antibody Technique, Indirect; Immunoenzyme Techniques; In Situ Nick-End Labeling; Keratitis; Ophthalmic Solutions; Platelet Membrane Glycoproteins; Pseudomonas; Rabbits; Receptors, G-Protein-Coupled

The purposes of this study were to establish a quantitative method for evaluating rabbit tear film status and investigate the efficacy of artificial tear preparations through ocular surface bathing or eye drop application.. The rabbit tear film was evaluated using a noninvasive specular reflection video recording system. The appearance of a tear break area (TBA) on the tear film images (7.4 mm2/mm) after 30 seconds of eye opening was quantified by image analysis. To induce disruption of the rabbit tear film, the ocular surface was challenged for 60 minutes with 1 ppm hypochloric acid (HOCl). Immediately after irrigation, artificial tear preparations composed of viscosity agents sodium hyaluronate (SH), hydroxypropylmethycellulose (HPMC), hydroxyethylcellulose (HEC), or chondroitin sulfate (CS) were applied to the rabbit eye through ocular surface bathing or eye drop application, and the recovery of the disrupted tear film was compared for each preparation.. A dramatic increase in TBA was observed immediately after the ocular surface was challenged with HOCl, and it returned to the initial level after 6 hours. Immediately after ocular surface bathing and eye drop application, a dramatic recovery of TBA was observed in all the test solution-treated eyes. One hour after treatments, prolonged amelioration of the tear film instability was observed after ocular surface bathing, but not by eye drop application, with the artificial tear preparations composed of HPMC or SH.. Ocular surface bathing with artificial tear preparations composed of a suitable viscosity agents could be useful in managing tear film instability.

Topics: Animals; Cellulose; Chondroitin Sulfates; Disease Models, Animal; Dry Eye Syndromes; Hyaluronic Acid; Hypochlorous Acid; Hypromellose Derivatives; Methylcellulose; Ophthalmic Solutions; Rabbits; Tears; Viscosity

Fucosylated chondroitin sulfate is a glycosaminoglycan from sea cucumber, made up of alternating beta-D-glucuronic acid and N-acetyl-beta-D-galactosamine units, like mammalian chondroitin sulfate. But the beta-D-glucuronic acid residues have branches of sulfated fucose, which confer high anticoagulant and antithrombotic properties on this compound. We have now compared the anticoagulant, bleeding and antithrombotic effects of this fucosylated chondroitin sulfate and its carboxyl-reduced derivative. Both compounds have similar anticoagulant action, mostly due to acceleration of thrombin inhibition in the presence of heparin cofactor II. The native glycosaminoglycan shows a correlation among anticoagulant, bleeding and antithrombotic effects. Inhibition of thrombosis in an arterio-venous shunt occurs at low doses, which are almost ineffective in modifying the anticoagulant activity of plasma. In a venous experimental model, on the contrary, antithrombotic activity requires high doses and occurs concomitantly with an increase in the anticoagulant activity of plasma. The action of fucosylated chondroitin sulfate on thrombosis is apparently unrelated to its effect on platelet aggregation. The carboxyl-reduced derivative of fucosylated chondroitin sulfate prevented thrombosis in the arterio-venous shunt, but not in the venous experimental model. This derivative did not increase bleeding, in spite of its potent anticoagulant activity. Therefore, our results reveal a dissociation of the anticoagulant, bleeding and antithrombotic effects of the glycosaminoglycan. Furthermore, we demonstrate that a polysaccharide may be a potent inhibitor of one type of thrombotic episode, but inactive on another. We propose that the different effects of fucosylated chondroitin sulfate and its carboxyl-reduced derivative on venous thrombosis may be related to adherence of the glycosaminoglycan to the vessel wall.

Topics: Acetylgalactosamine; Animals; Anticoagulants; Blood Coagulation; Carotid Arteries; Chondroitin Sulfates; Disease Models, Animal; Dose-Response Relationship, Drug; Female; Fucose; Glucuronic Acid; Hemorrhage; Heparin; Heparin Cofactor II; Male; Rats; Rats, Wistar; Sea Cucumbers; Venous Thrombosis

Hepatic fibrosis involves the interplay of many factors including reactive oxygen species. Recent reports described antioxidant properties of glycosaminoglycans (GAGs). Since several findings have shown that hyaluronic acid (HYA) and chondroitin-4-sulphate (C4S) may act as antioxidant molecules, the aim of this research was to evaluate the antioxidant effects of HYA and C4S treatment in a rat model of liver fibrosis. The effect on tissue inhibitors of metalloproteinases (TIMPs) was also studied. Liver fibrosis was induced in rats by eight intraperitoneal injections of CCl4, twice a week for 6 weeks. HYA or C4S alone (25 mg/kg) or HYA and C4S in combination (12.5 + 12.5 mg/kg) were administered daily by the same route during the 6 weeks. At the end of the 6-week treatment period (24 h after the last dose of GAGs), the following parameters were evaluated: (1) serum alanine aminotransferase (ALT) and aspartate aminotransferase (AST) activities, as index of hepatic cell disruption; (2) hepatic conjugated dienes (CD), as index of lipid peroxidation; (3) hepatic TIMPs activity and expression; (4) hepatic superoxide dismutase (SOD) and glutathione peroxidase (GPx) activity, as index of endogenous defences; (5) hepatic hydroxyproline, as index of collagen deposition. CCl4-induced liver fibrosis enhanced lipid peroxidation and TIMPs activation, increased ALT and AST, depleted antioxidants SOD and GPx, and caused collagen deposition in liver tissue. Treatment with GAGs, especially when in combination, successfully reduced ALT and AST rise, lipid peroxidation by evaluating conjugated dienes, TIMPs activation and mRNA expression, partially restored SOD and GPx activities, and limited collagen deposition in the hepatic tissue. The data obtained showed that these molecules were able to limit hepatic injury induced by chronic CCl4 intoxication and especially limited liver fibrosis. They also confirm that HYA and C4S may exert antioxidant mechanism, while reduction of TIMPs expression suggests that GAGs may influence MMPs and TIMPs imbalance in liver fibrosis.

Topics: Alanine Transaminase; Animals; Antioxidants; Aspartate Aminotransferases; Carbon Tetrachloride; Carbon Tetrachloride Poisoning; Chondroitin Sulfates; Collagen; Disease Models, Animal; Glycosaminoglycans; Hyaluronic Acid; Injections, Intraperitoneal; Lipid Peroxidation; Liver; Liver Cirrhosis; Male; Rats; Rats, Sprague-Dawley

Vascular wall protection can be achieved by preventive attachment to the vascular wall of antioxidants and elimination/neutralization of toxic products after their disproportioning. For this purpose we have prepared covalent conjugates between the vascular wall glycosaminglycan chondroitin sulfate (CHS) and the antioxidant enzymes superoxide dismutase (SOD) and catalase (CAT). The following conjugates were obtained: SOD-CHS, CAT-CHS and SOD-CHS-CAT. Their anti-thrombotic activity was compared in a rat model of arterial thrombosis by measuring the time of occlusion emergence and thrombus mass. It is noteworthy that the effectiveness of single bolus injections of SOD-CHS/CAT-CHS mixture was much lower than that of the bienzymic SOD-CHS-CAT conjugate. The conjugate SOD-CHS-CAT proved to be anti-thrombotically effective in doses two orders of magnitude lower than the native biocatalysts and an order of magnitude lower than SOD-CHS and CAT-CHS derivatives. For effective anti-thrombotic protection in oxidative conditions it is important to maintain the stable connection of SOD and CAT activity on the vascular wall and the large size of these conjugates. Covalent conjugate SOD-CHS-CAT is the best prospect for pharmaceutical development.

Topics: Animals; Antioxidants; Catalase; Chlorides; Chondroitin Sulfates; Disease Models, Animal; Ferric Compounds; Injections, Intravenous; Male; Rats; Structure-Activity Relationship; Superoxide Dismutase; Thrombosis

To test whether there is undersulfation of chondroitin sulfate in osteoarthritic bovine articular cartilage to support the hypothesis that sulfate deficiency is involved with the development of osteoarthritis.. Cartilage samples from bovine patellae (n = 32) were divided into 3 groups based on their osteoarthritic progression, as assessed by modified Mankin score. Uronic acid contents of the samples were determined. Fragmentation of the proteoglycans due to proteolytic processing was estimated with agarose gel electrophoresis. The molar ratios of chondroitin sulfate isoforms in the extracted proteoglycans were determined with fluorophore-assisted carbohydrate electrophoresis.. Loss of proteoglycans and accumulation of tissue water was evident in groups II and III, and progressive OA increased heterogeneity of aggrecan population in groups II and III. Importantly, the molar ratio of nonsulfated disaccharide was decreased in the osteoarthritic articular cartilage.. The structure of chondroitin sulfate in degenerated bovine cartilage did not support the hypothesis that sulfate depletion is present in osteoarthritic joint.

Topics: Animals; Biomarkers; Cartilage, Articular; Cattle; Chondroitin Sulfates; Disease Models, Animal; Disease Progression; Electrophoresis, Agar Gel; Glycosaminoglycans; Patella; Reference Values; Sensitivity and Specificity; Tissue Culture Techniques

As progress is made in the development of a tissue-engineered cardiac valve, the need for a reliable cell source is particularly important. A technique has been developed for the reliable biopsy of tricuspid valve leaflets. Expanding the harvested cells in culture is feasible and provides a source of leaflet cells that are structurally and functionally similar to the pulmonary and aortic valve leaflet cells that they may replace.. Thirteen sheep underwent tricuspid valve biopsy. Transthoracic echocardiography (TTE) was performed to evaluate function and guide the subsequent biopsy. Myofibroblasts were isolated from the biopsy samples, expanded in culture through 10 passages, and evaluated with immunocytochemistry for valve cell markers. Two animals were sacrificed acutely, two animals died during the immediate postoperative period, and nine animals survived for four weeks or more.. All preoperative and pre-explantation echocardiograms were normal. Both animals sacrificed acutely showed that the tricuspid valve leaflet was indeed biopsied with this technique. Two perioperative deaths occurred; one animal died secondary to injury of the chorda tendinea with subsequent destruction of the posterior leaflet; another died from disruption of the superior vena cava that led to irreversible cardiac tamponade. At sacrifice (2 to 17 weeks), all other animals showed intact tricuspid valves with normal leaflet anatomy. All cultured biopsies generated myofibroblasts that were immunocytochemically positive for alpha smooth muscle actin, chondroitin sulfate, vimentin and fibronectin.. Biopsy of the tricuspid valve to obtain recipient cardiac valve leaflet cells is possible, and the technique is simple and reliable. Biopsy of the leaflet does not compromise function. Interstitial cells can be harvested and expanded in culture. Cellular structure and function is preserved and is similar to that of other cardiac leaflet cells. Tricuspid valve leaflet biopsies are a potential source for harvesting cells to be used in the development of a tissue-engineered cardiac valve.

Topics: Actins; Animals; Biopsy; Cells, Cultured; Chondroitin Sulfates; Disease Models, Animal; Echocardiography, Transesophageal; Eosine Yellowish-(YS); Extracellular Matrix Proteins; Female; Fibroblasts; Fluorescent Dyes; Hematoxylin; Immunohistochemistry; Male; Models, Cardiovascular; Myocytes, Cardiac; Sheep; Staining and Labeling; Tissue Engineering; Tricuspid Valve; United States; Vimentin

This study was conducted to develop a new biomaterial to be used for an antioxidative drug. In this study, the hepatoprotective effect of chondroitin sulfate (CS) (100 mg/kg and 200 mg/kg body weight) was investigated at the antioxidative enzyme levels of liver total homogenate and mitochondria fraction. And the carbone tetrachloride (CCl(4))-induced rats were used as hepatotoxic models. The CCl(4) induced rat has been widely used as a hepatotoxic model due to its practicality, convenience and cost effectiveness since the generation of free oxygen radicals by CCl(4) injection was proposed as an important causative agent of hepatotoxicity. Malondialdehyde (MDA) levels were determined as well as the activities of superoxide dismutase (SOD), catalase (CAT), reduced-glutathione (GSH), oxidized-glutathione (GSSG) and glutathione peroxidase (GPx) in the liver. In addition, histopathology of liver tissue was investigated. Liver antioxidative enzyme activity was elevated while MDA concentration was decreased in all CS treated animals. The results demonstrated that CS protected oxidative stress in a dose dependent manner. Moreover, inflammation and cirrhosis in liver tissue of CS treated group were significantly decreased. It gave us an impression that CS might be a radical scavenger.

Topics: Animals; Antioxidants; Carbon Tetrachloride Poisoning; Catalase; Chemical and Drug Induced Liver Injury; Chondroitin Sulfates; Disease Models, Animal; Female; Glutathione; Glutathione Disulfide; Glutathione Peroxidase; Lipid Peroxidation; Malondialdehyde; Mitochondria, Liver; Rats; Rats, Sprague-Dawley; Superoxide Dismutase

We have previously demonstrated that enzymatic digestion of chondroitin sulfate proteoglycan (CSPG) at the scar promotes the axonal regrowth of Clarke's nucleus (CN) neurons into an implanted peripheral nerve graft after hemisection of the spinal cord. The present study examined whether degradation of CSPG using chondroitinase ABC promoted the regeneration of CN neurons through the scar into the rostral spinal cord in neonatal and adult rats. Following hemisection of the spinal cord at T11, either vehicle or chondroitinase ABC was applied onto the lesion site. The postoperative survival periods were 2 and 4 weeks. The regenerated CN neurons were retrogradely labeled by Fluoro-Gold injected at spinal cord level C7. In the sham group, there was no regeneration of injured CN neurons in both neonatal and adult rats. Treatment with 2.5 unit/ml chondroitinase ABC in neonates resulted in 11.8 and 8.3% of the injured CN neurons regenerated into the rostral spinal cord at 2 and 4 weeks, respectively. In adults, 9.4 and 12.3%, at 2 and 4 weeks, respectively, of the injured CN neurons regenerated their axons to the rostral spinal cord. The immunoreactivity for the carbohydrate epitope of CSPG was dramatically decreased around the lesion site after treatment with chondroitinase ABC compared to sham control in both neonatal and adult animals. Our results show that axonal regeneration in the spinal cord can be promoted by degradation of CSPG with chondroitinase ABC. This result further suggests that CSPG is inhibitory to the regeneration of neurons in the spinal cord after traumatic injury.

Topics: Age Factors; Animals; Animals, Newborn; Axons; Chondroitin ABC Lyase; Chondroitin Sulfate Proteoglycans; Chondroitin Sulfates; Cicatrix; Disease Models, Animal; Female; Immunohistochemistry; Nerve Regeneration; Neurons; Rats; Rats, Sprague-Dawley; Spinal Cord Injuries

Rheumatoid arthritis (RA) is a chronic, systemic, and inflammatory disease of connective tissue with unknown etiology. We investigated whether aberrant immune responses to glycosaminoglycans (GAGs), a major component of joint cartilage, joint fluid, and other soft connective tissue, causes this disease. Here we show that injection of GAGs such as hyaluronic acid, heparin, and chondroitin sulfates A, B, and C induce arthritis, tendosynovitis, dermatitis, and other pathological conditions in mice. We developed a technique by staining tissue specimens with fluorochrome- or biotin-labeled GAGs to visualize the direct binding between cells and GAGs. We discovered that inflammatory infiltrates from the affected tissue are dominated by a distinct phenotype of GAG-binding cells, a significant portion of which are CD4(+) T cells. GAG-binding cells seem to be expanded in bone marrow of GAG-immunized mice. Furthermore, we identified GAG-binding cells in inflamed synovial tissue of human patients with RA. Our findings suggest that carbohydrate self-antigenic GAGs provoke autoimmune dysfunctions that involve the expansion of GAG-binding cells which migrate to anatomical sites rich in GAGs. These GAG-binding cells might, in turn, promote the inflammation and pathology seen both in our murine model and in human RA.

Topics: Animals; Arthritis, Rheumatoid; Autoantibodies; Bone Marrow Cells; CD4-Positive T-Lymphocytes; Chondroitin Sulfates; Dermatan Sulfate; Disease Models, Animal; Female; Glycosaminoglycans; Heparin; Humans; Hyaluronic Acid; Immunophenotyping; Mice; Mice, Inbred C57BL

Current use of Integra, the collagen-based dermal analogue, requires a two-step grafting procedure to achieve wound closure with an "ultrathin" autograft.. A one-step operative procedure of meshed composite skin graft (MCSG) using Integra as a dermal template for a meshed split thickness autograft was developed in rats. The silicon layer of Integra was removed, the resulting dermal analogue was meshed (1:1.5), expanded, and placed on excised full thickness wound and covered with a meshed (1:1.5 or 1:6) split thickness autograft. Grafted wounds were dressed with BioBrane, Vaseline gauze, silver-impregnated nylon, or silver-nylon and direct current (SNDC). At scheduled intervals up to 3 months postgrafting, wounds were examined for epithelialization, collagen deposition and fibrosis, hair growth, and contraction. The results of wound closure and healing following the one-step procedure were compared with the outcome of the two-step grafting procedure where application of meshed Integra (step one) was followed in 14 days by removal of the silicon layer and application of the meshed autograft (step two).. The one-step procedure applied to meshed autograft/Integra (1:1.5/1:1.5) composite graft accelerated wound closure by 6-19 days when compared with the two-step procedure. At 3 months postgrafting, the contraction of the healed wound dressed with SNDC, BioBrane, or Vaseline gauze was reduced by 13-16% following the one-step procedure compared with the two-step procedure (p < 0.05). The one-step procedure allowed the expansion of the autograft layer to 1:6 while achieving wound healing results similar to grafting with 1:1.5 meshed autograft layer using the two-step grafting procedure.. Single-step application of meshed, thin, split thickness autograft over meshed Integra-derived dermal substitute allows more rapid wound closure with less contraction and more efficient use of graft donor skin than can be obtained with the commonly used two-step grafting procedure.

Topics: Analysis of Variance; Animals; Biocompatible Materials; Chondroitin Sulfates; Collagen; Dermis; Disease Models, Animal; Female; Occlusive Dressings; Outcome and Process Assessment, Health Care; Rats; Rats, Inbred Lew; Skin Transplantation; Skin, Artificial; Surgical Mesh; Time Factors; Transplantation, Autologous; Wound Healing; Wounds, Penetrating

Polyhydroxyethyl methacrylate (pHEMA) membranes coated on one side with chondroitin sulfate (CS) were used to block adhesion physically and to reduce friction between healing flexor tendons and the surrounding tissue in rabbit forepaws after surgical repair. Digits with pHEMA-only, standard tendon sheath repair, and with no sheath repair were the controls. Over 12 weeks the CS-coated membranes were evaluated for joint flexion, adhesion limitation, and tendon healing progress. The membranes initially allowed for better flexion (ie, for 6 weeks), but their relative superior effectiveness faded afterward. Histology showed that adhesions were less severe and healing was better in the CS-pHEMA membranes at 3 and 6 weeks. If further studies determine precise amounts or thicknesses of CS coats that will maximize its healing properties, CS-pHEMA should prove useful in clinical settings in which restoration of tendon sheath integrity with a minimum of adhesions is not possible.

Topics: Animals; Biocompatible Materials; Chondroitin Sulfates; Disease Models, Animal; Drug Combinations; Polyhydroxyethyl Methacrylate; Rabbits; Range of Motion, Articular; Tendon Injuries; Time Factors; Tissue Adhesions; Wound Healing

Colonic mucus is decreased in a rat model of spastic constipation, and some types of water-insoluble dietary fiber increase colonic mucus when consumed by rats for several weeks. However, little is known about the effect of water-soluble dietary fiber on the colonic mucus. The aim of the present study was to investigate the effect of various types of water-soluble dietary fiber on colonic mucus in a rat model of spastic constipation. Oral administration of 1.5 mg/day of carrageenan and chondroitin sulfate increased the fecal excretion, epithelial mucin production, thickness of the mucous layer, and amount of luminal mucus in loperamide-administered rats. Sodium alginate, 5 mg/day, thickened the mucus layer at the fecal surface. Cellulose, 5 mg/day, increased the fecal excretion but not the colonic mucus. Carrageenan, chondroitin sulfate, and sodium alginate, but not cellulose, increased colonic mucus in the rat model of spastic constipation.

Topics: Alginates; Animals; Carrageenan; Cellulose; Chondroitin Sulfates; Colon; Constipation; Dietary Fiber; Disease Models, Animal; Glucuronic Acid; Hexuronic Acids; Loperamide; Male; Mucus; Polysaccharides; Rats; Rats, Sprague-Dawley

Danaparoid and heparin, on the basis of anti-activated factor X (anti-FXa) activity, were equipotent in accelerating the rate of interaction of FXa and antithrombin III. In rat tissue factor-induced disseminated intravascular coagulation (DIC) models, an intravenous administration of danaparoid inhibited the decrease in plasma fibrinogen and platelet counts and the increase in serum fibrinogen degradation products. Expressed on the basis of anti-FXa activity, these effects were comparable with those of dalteparin and heparin. In rat mesenteric small artery and vein, less bleeding was observed after intravenous administration of danaparoid than after dalteparin or heparin. Danaparoid did not affect adenosine diphosphate- or collagen-induced platelet aggregation, and showed weaker inhibitory effects on aggregation induced by thrombin, or collagen + thrombin, than did dalteparin or heparin. These findings suggest that danaparoid may be useful for the prevention of DIC and has less tendency to cause bleeding than dalteparin or heparin, probably as a result of its weaker ability to inhibit platelet aggregation.

Topics: Animals; Anticoagulants; Antithrombin III; Bleeding Time; Chondroitin Sulfates; Dalteparin; Dermatan Sulfate; Disease Models, Animal; Disseminated Intravascular Coagulation; Drug Combinations; Drug Evaluation, Preclinical; Enzyme Inhibitors; Factor Xa; Factor Xa Inhibitors; Heparin; Heparitin Sulfate; Kinetics; Male; Platelet Aggregation; Rats; Rats, Wistar; Risk Assessment; Thromboplastin

Although none of these markers currently have any clinical applications, researchers have made strong advances in understanding molecular markers of OA and remain optimistic that the goal of identifying clinically useful markers of OA is attainable.(3-6,8,9) Much of this positive sentiment arises from the large array of molecules that have been identified.(8) Molecular markers of the greatest potential clinical use will be those that allow early detection of OA, permit disease progression to be monitored, or allow efficacy of various treatment regimens to be assessed. Earlier and more sensitive detection of OA changes may increase effective opportunities for treatment intervention during reversible phases of the OA disease process and allow more objective assessment of treatment efficacy and prognosis.(6)

Topics: Animals; Biomarkers; Cartilage, Articular; Chondroitin Sulfates; Disease Models, Animal; Dog Diseases; Dogs; Femur; Keratan Sulfate; Matrix Metalloproteinase 3; Osteoarthritis; Synovial Fluid

The susceptibility of 129/SvJ mice to infection with Strongyloides venezuelensis was compared with that of C57BL/6 mice. After a primary infection, daily egg output in faeces (EPG) from 129/SvJ mice was lower and terminated earlier than that from C57BL/6 mice. Adult worm recovery from the small intestine of 129/SvJ mice on day 7 was also lower than that of C57BL/6 mice. When the numbers of larvae recovered from the lungs were examined on days 2, 3 and 4 after a primary infection, they were comparable between the two strains. On the other hand, when an equal number of larvae recovered from the lungs of each strain on day 3 were implanted orally into homologous strain mice, the magnitude of EPG and the number of adult worms in the small intestine on day 5 after implantation were significantly lower in 129/SvJ than in C57BL/6 mice. The number of mucosal mast cells in the jejunum was not significantly different between 129/SvJ and C57BL/6 naive mice. Total chondroitin sulphate concentration in the gut washings obtained from naive mice was significantly higher in 129/SvJ (11.34 +/- 9.48) than in C57BL/6 mice (1.09 +/- 0.77, P < 0.05). These results indicate that the natural resistance of 129SvJ mice to S. venezuelensis infection is expressed at the intestine, probably due to higher concentration of chondroitin sulphate, which prevents establishment of S. venezuelensis.

Topics: Animals; Chondroitin Sulfates; Disease Models, Animal; Immunity, Innate; Intestine, Small; Male; Mast Cells; Mice; Mice, Inbred C57BL; Parasite Egg Count; Rats; Rats, Wistar; Strongyloides; Strongyloidiasis

Damage to the meniscus can lead to posttraumatic osteoarthritis. Early markers of joint injury and tissue disease may be useful in developing and administering clinical treatment. We investigated the effects of total medial meniscectomy on biomarkers measured serially in synovial lavage fluid each month for 3 months. Following meniscectomy in dogs, four biomarkers were evaluated: cartilage oligomeric matrix protein, keratan sulfate epitope (5D4), the 3B3(-) neoepitope of chondroitin-6-sulfate, and the 3B3(+) chondroitinase-generated epitope of chondroitin-6-sulfate. Meniscectomy led to statistically significant elevations of all four biomarkers, with levels peaking at 4 weeks. By 12 weeks, the level of the 5D4 epitope returned to the preoperative baseline level whereas that of cartilage oligomeric matrix protein, 3B3(-), and 3B3(+) remained above the baseline. Concentrations of these biomarkers in the knees not operated on did not change significantly from the baseline. The levels of cartilage oligomeric matrix protein and 3B3(-) relative to 3B3(+) remained constant in all knees. In contrast, the level of 5D4 relative to 3B3(+) declined over time in the knee operated on but remained constant in the knee not operated on. These results demonstrate a quantitative change in the molecular components of synovial fluid after meniscectomy, as well as a qualitative change evinced by an alteration in the relative proportions of these epitopes. Extensive analyses showed a strong correlation between serum levels of 3B3(-) from the femoral and cephalic veins; however, serum 3B3(-) was not correlated with synovial fluid 3B3(-). These findings support the hypothesis that the concentrations of select cartilage biomarkers in synovial fluid are altered following meniscectomy and are promising tools for objectively monitoring the induction of osteoarthritis in this model system.

Topics: Animals; Bacterial Proteins; Biomarkers; Cartilage, Articular; Chondroitin Sulfates; Disease Models, Animal; Dogs; Epitopes; Keratan Sulfate; Male; Membrane Proteins; Menisci, Tibial; Osteoarthritis; Synovial Fluid; Transferases

The antithrombotic activity of a fucosylated chondroitin sulfate extracted from the body wall of a sea cucumber was assessed using a stasis thrombosis model in rats. Intravenous administration of the polysaccharide reduced thrombosis in a dose-dependent manner. We also compared the antithrombotic action of the sea cucumber chondroitin sulfate with that of standard mammalian glycosaminoglycans, mainly heparin and dermatan sulfate. Intravascular injection of fucosylated chondroitin sulfate at the dose totally preventing thrombus formation produced a much more intense modification of the plasma anticoagulant activity than antithrombotic doses of unfractionated heparin, low-molecular-weight heparin and mammalian dermatan sulfate. Thus, it is possible that the mechanism of antithrombotic action of these polysaccharides are different. For fucosylated chondroitin sulfate, it depends mostly on modifications of the plasma anticoagulant activity, but it may involve additional effects in the case of mammalian glycosaminoglycans, perhaps modifications induced in the cells of the vessel wall. The anticoagulant and possibly the antithrombotic actions of fucosylated chondroitin sulfate are mostly dependent on heparin cofactor II activity, and both are markedly reduced with the decrease of the chain size of the polymer. Overall, the sulfated polysaccharide from the invertebrate revealed an unequivocal effect in preventing experimental venous thrombosis, is a useful tool to investigate the antithrombotic action in mammals and may offer an alternative for future development of a new therapeutic agent.

Topics: Animals; Anticoagulants; Carbohydrate Sequence; Chondroitin Sulfates; Dermatan Sulfate; Disease Models, Animal; Dose-Response Relationship, Drug; Echinodermata; Female; Fibrinolytic Agents; Fucose; Glycosaminoglycans; Heparin; Male; Molecular Sequence Data; Molecular Weight; Partial Thromboplastin Time; Rats; Sulfur Radioisotopes; Thrombosis

Supplements of glucosamine hydrochloride, low molecular weight chondroitin sulfate, and manganese ascorbate were tested separately and in combination for their ability to retard progression of cartilage degeneration in a rabbit instability model of osteoarthrosis. Computerized quantitative histologic evaluation of safranin O stained sections of the medial femoral condyles measured the grade and extent of tissue involvement of lesions. Severe lesions (Mankin grade greater than 7) were absent in all animals supplemented with a dietary mixture of glucosamine, chondroitin sulfate, and manganese ascorbate. Total linear involvement (mm of lesioned surface) and total grade (mean grade x number of lesions per animal) were reduced significantly in animals given the combination compared with controls (59% and 74% respectively). Animals supplemented with glucosamine, chondroitin sulfate, or manganese ascorbate alone had less moderate and severe tissue involvement than controls but not to the extent of the combined group. In vitro, a combination of glucosamine hydrochloride and chondroitin sulfate acted synergistically in stimulating glycosaminoglycan synthesis (96.6%). Chondroitin sulfate and manganese ascorbate but not glucosamine were effective in inhibiting degradative enzyme activity. These data suggest that the disease modifying effect (the ability to retard progression of cartilage degeneration) of a mixture of glucosamine, chondroitin sulfate, and manganese ascorbate is more efficacious than either agent alone.

Topics: Animals; Cartilage, Articular; Chondroitin Sulfates; Dietary Supplements; Disease Models, Animal; Drug Synergism; Drug Therapy, Combination; Glucosamine; Manganese; Osteoarthritis; Rabbits

Matrix degradation and apoptosis are two crucial features of osteoarthrosis (OA), but the relationship between them needs to be clarified. We developed a novel OA model in partial meniscectomisized rats for the assessment in situ of these processes. The surgical procedure used permitted us to identify and remove 30-50% of the meniscus. Furthermore, the inclusion of high impact exercise enhanced the cartilage damage in a short period of time. OA cartilage displayed a rough and eroded surface, fibrosis and even complete loss; we also found hypertrophy, chondrocyte clusters and a disrupted extracellular matrix. As evidence of matrix degradation we found a diminution of the extracellular components such as chondroitin sulfate-4, 6 (CS-4, 6) and proteoglycans (PGs), and an overall increased activity of the PGs-degrading enzyme, stromelysin-1 (SLN-1). Moreover, the frequency of apoptotic chondrocytes increased according to the severity of the matrix degradation, as detected by TUNEL technique and by morphology. All these changes were enhanced in OA rats with high-impact exercise. Trained rats showed a statistically highly significant difference in histological changes compared to untrained animals. We demonstrate that our model can be evaluated according to Mankin's histologically graded parameters. Also, it is an effective method to assess, in situ, apoptosis in an experimentally induced OA. Our results suggest that apoptosis could be involved in matrix degradation and that TUNEL might be a good method for spotting early apoptotic changes which, combined with careful morphological examinations, provide substantial help in improving apoptosis detection.

Topics: Animals; Apoptosis; Cell Count; Chondrocytes; Chondroitin Sulfates; Disease Models, Animal; Extracellular Matrix; Fluorescent Antibody Technique, Indirect; In Situ Nick-End Labeling; Male; Matrix Metalloproteinase 3; Menisci, Tibial; Microscopy, Confocal; Osteoarthritis; Physical Conditioning, Animal; Proteoglycans; Rats; Rats, Wistar

Individual antithrombotic activities of superoxide dismutase (SOD) and sodium chondroitin sulfate (CHS) as well as the activities of covalent and noncovalent complexes of SOD with CHS were compared in a rat model of arterial thrombosis induced by ferrous chloride. Covalent conjugate of SOD with CHS exerted the most potent antithrombotic effect, which was associated with adsorption of the conjugate on the glycocalyx of the vascular wall cells and stability of the covalent bond between CHS and SOD subunits. Theoretical and practical directions in the investigation of SOD and CHS preparations are outlined.

Topics: Animals; Carotid Arteries; Chondroitin Sulfates; Disease Models, Animal; Dose-Response Relationship, Drug; Drug Combinations; Drug Stability; Ferrous Compounds; Fibrinolytic Agents; Glycocalyx; In Vitro Techniques; Male; Rats; Superoxide Dismutase; Thrombosis

Coronal stepoffs of 0.5 mm (equal to the cartilage height) were created on the medial femoral condyles of adult, skeletally mature rabbits as a model for articular surface incongruity. After 3, 6, 12, and 24 weeks, tissue was analyzed histologically using hematoxylin and eosin and Safranin O staining, autoradiographs were made of the femoral condyles, and immunohistologic analysis was done for 3-B-3(-) and 7-D-4 chondroitin sulfate epitopes. An overlapping flap from the high toward the low side and an increase of the cartilage height on the low side of the defect were observed as permanent features of adaptation throughout the entire followup. Significant degeneration was not seen around the lesion or in the tibial cartilage opposing a stepoff defect. Autoradiography showed a three-phase response to the lesion: an early increase in radiolabeled sulfate (35SO4) uptake, a sharp decline of 35SO4 uptake, and finally a late recovery of the autoradiographic signal indicating partial recovery of proteoglycan synthetic activity. After an early increase, immunohistologic analysis for 3-B-3(-) showed a subsiding tendency by 24 weeks, and the staining with 7-D-4 remained elevated uniformly in the vicinity of the lesion. A rabbit femoral stepoff defect with an offset of 0.5 mm may remodel and not lead to degeneration within the first 6 months after injury in a stable joint.

Topics: Adaptation, Physiological; Animals; Autoradiography; Cartilage Diseases; Cartilage, Articular; Chondroitin Sulfates; Coloring Agents; Disease Models, Animal; Eosine Yellowish-(YS); Epitopes; Female; Femur Head; Fluorescent Dyes; Follow-Up Studies; Hematoxylin; Hindlimb; Immunohistochemistry; Phenazines; Proteoglycans; Rabbits; Radiopharmaceuticals; Range of Motion, Articular; Sulfates; Sulfur Radioisotopes

A decrease in anionic change and the loss of heparan sulfate proteoglycan have previously been observed in the glomerular basement membrane (GBM) during diabetic glomerulosclerosis. We studied the chronological changes in the anionic character and the glycosaminoglycan content in the GBM of WBN/ Kob rats with spontaneous diabetes. Two types of cationic probes were used: polyethyleneimine (PEI) and cationic colloidal gold (CCG). Immunogold labeling was performed with anti-monoclonal-heparan-sulfate-glycosaminoglycan (HS-GAG) and anti-chondroitin-sulfate-glycosaminoglycan (CS-GAG) antibodies. The GBM width, the anionic sites and the GAG sites were investigated in diabetic WBN/Kob rats at 2, 10 and 19 months, compared with control rats. Diabetes was confirmed in WBN/Kob rats after 8 months in this study. The GBM width gradually thickened with age. The PEI anionic sites significantly decreased in the lamina rara externa (LRE) at 19 months (vs. 2 and 10 months). The HS-GAG sites also significantly decreased in the LRE at 10 and 19 months (vs. 2 months). However, the CCG anionic sites and the CS-GAG sites significantly increased in the LRE and the lamina densa at 10 months (vs. 2 months) and, after 19 months, returned to the level seen at 2 months. Results indicate that there is an early transient increase in CS-GAG in the GBM while HS-GAG decreases. We noticed a transient increase in the CCG anionic sites at this early stage of diabetic glomerulosclerosis as well. The increase in CS-GAG may provide a marker for early diabetic changes in the GBM.

Topics: Animals; Anions; Antibodies, Monoclonal; Basement Membrane; Chondroitin Sulfates; Diabetes Mellitus, Type 2; Diabetic Nephropathies; Disease Models, Animal; Heparitin Sulfate; Immunohistochemistry; Kidney Glomerulus; Male; Rats; Rats, Wistar

To study how the concentrations of proteoglycans (PGs) and collagen change in various parts of tibial articular cartilage during aging, and to evaluate the development of spontaneous osteoarthrosis (OA) in guinea pigs.. PGs were extracted from guinea pig cartilage samples using 4M guanidine hydrochloride, and the amount of hydroxyproline was determined in the extraction remainder. The molecular size and aggregation of PGs were analyzed by electrophoresis, and the glycosaminoglycan composition was assessed by high-performance liquid chromatography.. The PG concentration was proportional to the load distribution. However, when OA became histologically manifest, the PG concentration decreased by 50% (from a mean of 44 microg to 22 microg per mg fresh tissue) and the collagen level decreased by 40% (from a mean of 17 microg to 10 microg per mg fresh tissue), while the proportion of water increased by 13% (from a mean of 710 mg to 800 mg per mg fresh tissue).. Unmineralized cartilage can, within physiologic load limits, respond to increased mechanical demands by increasing the PG and collagen concentrations. Beyond a certain limit, however, the cartilage can no longer compensate for further increases in stress, which results in cartilage degeneration and losses of matrix constituents. These losses seemed to appear earlier in the disease process than has been described in previous animal models of secondary OA.

Topics: Aging; Animals; Body Weight; Cartilage, Articular; Chondroitin Sulfates; Chromatography, High Pressure Liquid; Disease Models, Animal; Electrophoresis; Glycosaminoglycans; Guinea Pigs; Hyaluronic Acid; Male; Osteoarthritis; Proteoglycans

To investigate the effect of diacetyl rhein (DAR) on the synthesis, turnover and composition of cartilage in an experimental model of osteoarthritis in beagle bitches.. Osteoarthritis was induced in mature beagle bitches by the transection of the cranial cruciate ligament. Six animals received DAR 20 mg/kg daily for 11 weeks. A matched group received empty capsules daily for the same period. At 11 weeks, articular cartilage was examined for the ratio of the 6:4-sulfated disaccharides of chondroitin and the tissue concentration of hydroxyproline and glycosaminoglycan. In addition, labeling studies were performed to estimate the effect of DAR on proteoglycan synthesis and turnover.. DAR had no effect on body weight or food consumption but induced a mild diarrhea and slightly increased the incidence of vomiting. DAR tended to reduce proteoglycan synthesis, however, DAR did reduce proteoglycan turnover in the femoral cartilage. DAR produced changes in the composition of the osteoarthritic cartilage that could only partly be accounted for by changes in hydration and/or swelling. In addition, it was noted that induction of osteoarthritis increased the ratio of chondroitin 6-sulfated to chondroitin 4-sulfated disaccharides; DAR reduced the ratio in tibial plateau cartilage from osteoarthritic joints compared with untreated tissue from osteoarthritic joints. DAR showed moderate reduction on the biosynthesis of proteoglycans. DAR also produced a reduction in proteoglycan turnover from all anatomical areas compared with non-treated controls in both the lateral and medial femoral condyles.. DAR was well tolerated by the experimental animals, but did not produce significant changes in the synthesis or turnover of proteoglycans. The slight reduction in proteoglycan synthesis may prove to be biologically significant after chronic dosing. DAR's effects on the hydroxyproline and glycosaminoglycan content suggest, however, that it must influence the swelling of cartilage and loss of glycosaminoglycan. This indicates that small changes can translate, to significant differences in cartilage composition over an 11-week time period.

Topics: Animals; Anthraquinones; Anti-Inflammatory Agents, Non-Steroidal; Biomarkers; Cartilage, Articular; Cells, Cultured; Chondroitin Sulfates; Disease Models, Animal; Dogs; Epitopes; Glycosaminoglycans; Hydroxyproline; Osteoarthritis; Proteoglycans

To define the architecture and extracellular matrix composition of the lamina cribrosa in rodents, normal, adult pigmented rat and guinea pig eyes were frozen and sectioned for light microscopic immunohistochemistry. Antibodies specific for collagens I, III, IV and VI, laminin, elastin, and chondroitin and dermatan sulfate proteoglycans were exposed to longitudinal and cross-sections of optic nerve heads and their binding distributions observed with the avidin-biotin-peroxidase complex technique. Cross-sections of the intraocular portion of the rat optic nerve head revealed a horizontally oval shape with distinct, vertically oriented, laminar beams. The guinea pig optic nervehead cross-section was circular, with randomly oriented beams. In both animals, collagens I, III and VI were found throughout the laminar beams, along with elastin fibrils. Collagen IV and laminin antibodies deposited along laminar beam margins and within the beams, representing astrocytic and vascular endothelial cell basement membranes. Both animals showed evidence for dermatan and chondroitin sulfate-containing proteoglycans in all connective tissue structures of the nerve head. In the rat, chondroitin-4 sulfate proteoglycans appeared localized to the sclera and laminar beams. The rat and the guinea pig optic nerve head possess an identifiable lamina cribrosa with structural proteins nearly identical to that of the primate. Both animals may provide affordable alternative animal models for in vivo studies on the role of the lamina cribrosa in glaucomatous optic nerve damage.

Topics: Animals; Chondroitin Sulfates; Collagen; Dermatan Sulfate; Disease Models, Animal; Glaucoma; Guinea Pigs; Immunoenzyme Techniques; Optic Disk; Rats; Rats, Inbred BN

The dry eye syndrome is a chronic disease which can become a serious threat to useful vision. However, there is only a limited understanding regarding effective treatment or prevention of this disease. Establishing an effective mode of treatment requires the use of a satisfactory animal dry eye model. Ideally, such a model should rapidly determine the effectiveness of agents that inhibit the damaging effects of this syndrome. This paper presents a short-term dry eye model using rabbits, which combines mechanical prevention of blinking and methylene blue staining. This model is not intended to be a precise representation of the dry eye syndrome, since this disorder has recently become recognized to involve a primary pathological process of the corneal and conjunctival epithelium. However, by using this model, clinical signs of dry eye can be observed after a few hours in the form of acute desiccation. Corneal damage can easily be evaluated both qualitatively by methylene blue staining scores, and quantitatively by chronic assay. Visually observed corneal epithelial thinning was confirmed by scanning electron microscopy (SEM) to be due to loss of epithelial integrity. Using a 3% chondroitin sulfate solution, an already proven effective agent for dry eye, this model effectively demonstrated an 80% inhibition in the development of methylene blue positive lesion after a period of only 2 hours. This short term dry eye model is valuable in primarily screening the efficacy of potential therapeutic agents in the prevention and treatment of dry eye.

Topics: Animals; Chondroitin Sulfates; Coloring Agents; Cornea; Desiccation; Disease Models, Animal; Dry Eye Syndromes; Epithelium; Male; Methylene Blue; Microscopy, Electron, Scanning; Ophthalmic Solutions; Rabbits

Alterations in basement membrane components, notably proteoglycans, in a rat model of polycystic kidney disease have been investigated. Rats were fed phenol II (2-amino-4-hydroxyphenyl-5-phenyl thiazole) for 4 days and then changed to normal diet for a 7-day recovery period. Marked dilation of distal tubules and collecting ducts was observed by 4 days with phenol II treatment, but the morphology returned to normal after 7 days of subsequent normal diet. Staining of tissue sections with two mouse monoclonal antibodies to a recently described basement membrane chondroitin sulfate proteoglycan (BM-CSPG) core protein was markedly diminished in the basement membranes of dilated cystic tubules. Reduction in staining was evident as early as 2 days. During recovery, BM-CSPG increased in tubular basement membranes and returned to normal after 7 days. Staining with a polyclonal antibody to chondroitin sulfate chains confirmed these changes in cystic tubule basement membranes. During the recovery stage, interstitial chondroitin sulfate (representing a CSPG other than BM-CSPG) was greatly increased around these tubules, along with the glycoprotein fibronectin. Staining with antibody to a basement membrane heparan sulfate proteoglycan core protein related to perlecan did not diminish but rather stained affected tubules intensely, whereas laminin, on the other hand, was apparently diminished in the basement membranes of the cystic tubules. Type IV collagen staining did not change through disease onset or recovery. These results suggest that BM-CSPG, which was rapidly altered in distribution through the onset and recovery phases, may be a sensitive marker of the cystic state, and in addition, the expression of basement membrane proteoglycans may be specifically and separately regulated in this disease.

Topics: Aging; Animals; Basement Membrane; Chondroitin Sulfates; Collagen; Disease Models, Animal; Fibronectins; Fluorescent Antibody Technique; Kidney Tubules; Laminin; Male; Phenols; Polycystic Kidney Diseases; Rats; Rats, Sprague-Dawley; Staining and Labeling

Canine experimental models of osteoarthritis (OA) and disuse atrophy were used to study cartilage metabolism. The synovial fluids from the OA joints showed elevated levels of keratan sulfate (KS) epitope and link protein, indicating increased catabolism. Analysis of fluids from joints with disuse atrophy showed high levels of KS epitope, but no increase in link protein. Quantitation of a novel chondroitin sulfate (3B3) epitope showed it to be present only in the synovial fluids and articular cartilage of the OA joints. The results indicate that these may be important indicators, or markers, of degenerative joint disease.

Topics: Animals; Atrophy; Cartilage, Articular; Chondroitin Sulfates; Disease Models, Animal; Dogs; Keratan Sulfate; Osteoarthritis; Proteoglycans; Synovial Fluid

The effect of chondroitin sulfate upon the growth of calcium oxalate crystals was measured in vivo by using an experimental model in rats. Adult male Wistar rats were treated by chronic i.p. injections of chondroitin sulfate solutions (1, 5 or 10 mg in 0.3 ml of saline, every 2 days). This treatment led to a dose-dependent increase in the urinary chondroitin sulfate concentration. Urolithiasis was induced by the introduction of a calcium oxalate seed into the bladder of the animals. Urine samples were collected and the calculi formed were removed after 42 days. The chondroitin sulfate concentration have decreased in the lithiasic urines, as compared to controls and higher chondroitin sulfate doses correlated with larger calculi. The presence of chondroitin sulfate in the matrices of stones obtained from chondroitin sulfate-treated animals suggested that there was some adsorption of chondroitin sulfate on to the growth sites of the calcium oxalate crystals. In contrast to the chondroitin sulfate effect observed in vitro, which inhibits the growth of calcium oxalate crystals, our results suggest that in vivo chondroitin sulfate promotes the growth of stones in the urinary tract.

Topics: Adult; Animals; Calcium Oxalate; Child; Chondroitin Sulfates; Disease Models, Animal; Humans; Rats; Rats, Inbred SHR; Rats, Inbred Strains; Urinary Bladder Calculi

Out of 30 adult rabbits, 20 had one knee immobilized with a plaster of Paris cast for 6 or 12 weeks, and 10 rabbits were used as untreated controls. Prior to immobilization, 10 knees were injected with high-molecular weight hyaluronic acid. The articular cartilage of the femoral condyles was studied by light microscopy, whereas that of the patella and tibia was analyzed biochemically. Degenerative changes of the articular cartilage similar to those seen in arthrosis were observed after 6 weeks. The intraarticular injection of hyaluronic acid did not prevent these changes; instead, the reparative processes seemed inhibited.

Topics: Animals; Arthritis; Cartilage, Articular; Chondroitin Sulfates; Dermatan Sulfate; Disease Models, Animal; Evaluation Studies as Topic; Hyaluronic Acid; Immobilization; Injections, Intra-Articular; Keratan Sulfate; Knee Joint; Rabbits; Uronic Acids

Two experimental thrombosis models in rats have been compared with regard to the composition of the formed thrombi and the effects of various treatments on thrombus formation. In the first model thrombosis is induced in the vena cava by a combination of venous stasis and hypercoagulability; these thrombi consist merely of red cells and fibrin with only a few platelets. In the second model thrombosis is induced in an arterio-venous shunt in which the formed thrombi consist of red cells, fibrin and a large amount of platelet aggregates adhering to the foreign material. Antiplatelet serum and acetylsalicylic acid, which reduce blood platelet activity, inhibited thrombus formation only in the arteriovenous shunt model. Dicumoxane, an oral anticoagulant, was active in both models. The glycosaminoglycans heparin, Org 10172, Fragmin and the pentasaccharide, representing the AT-III binding sequence of heparin, were active in both models. However, there were qualitative and quantitative differences between the effects of the glycosaminoglycans suggesting differences in their modes of action.

Topics: Animals; Arteriovenous Shunt, Surgical; Aspirin; Blood Coagulation Factors; Blood Platelets; Chondroitin Sulfates; Coumarins; Dermatan Sulfate; Disease Models, Animal; Fibrin; Fibrinolytic Agents; Glycosaminoglycans; Heparin; Heparin, Low-Molecular-Weight; Heparitin Sulfate; Immune Sera; Ligation; Male; Oligosaccharides; Rats; Thrombosis; Venae Cavae; Vitamin K

The vascular proteoglycans probably have an important influence on the biomechanical properties of blood vessels and, therefore, may play a role in the development or maintenance of hypertension. In the aorta of the spontaneously hypertensive rat, the authors previously observed an increased content of chondroitin sulfate, an increased incorporation of [35S]sulfate into proteoglycans, and qualitative alterations in the [35S]polysaccharides compared to the normotensive Wistar Kyoto rat. To determine if these differences were related to hypertension or to strain variations, normotensive and hypertensive Dahl S rats were studied. There was a significant elevation (70%) in the aorta content of chondroitin sulfate, whereas the dermatan sulfate and hyaluronic acid contents were similar in the two groups. The in vitro incorporation of [35S]sulfate was increased 2.6-fold in the hypertensive animals. No differences between the two groups were observed with respect to the gel chromatographic profiles of the [35S]proteoglycans or the charge density of the [35S]glycosaminoglycans, as assessed by ion exchange chromatography. It was concluded that the increase in chondroitin sulfate and [35S]sulfate incorporation into proteoglycans occurred as a result of hypertension, regardless of genetic factors.

Topics: Animals; Aorta; Chondroitin Sulfates; Dermatan Sulfate; Disease Models, Animal; Hyaluronic Acid; Hypertension; Male; Proteoglycans; Rats; Sodium Chloride

Ocular surface drying was studied in rabbits using an experimental model of blinking interruption. This model produces a discontinuity in the pre-corneal tear film after a few minutes of exposure to air. Changes in superficial epithelial cell structure were studied using a scan microscope. A decrease of micro-folds and an increase in epithelial cell desquamation were observed. Prolonged exposure to air produced similar cellular lesions in deeper layers. The instillation of a single mucomimetic eye-drop containing chondroitin sulphate prevented total interruption of epithelial humidification, kept the epithelial surface intact and preserved normal cellular structure.

Topics: Animals; Chondroitin; Chondroitin Sulfates; Cornea; Disease Models, Animal; Epithelium; Microscopy, Electron, Scanning; Ophthalmic Solutions; Rabbits; Xerophthalmia

We studied changes in subchondral bone and articular cartilage in an animal model of osteoarthrosis. In this model we applied repetitive impulsive loads to rabbits' knees. Their legs were held in short leg splints so the rabbits were unable to dampen the peak applied load with ankle flexion. After sacrifice, at 1 day to 6 weeks, we studied proximal tibial load-bearing cartilage histologically, biochemically, and with radioactive sulfate uptake. We also studied the subchondral bone under that cartilage histologically, histomorphometrically, with bone scan (99mTc pyrophosphate), and by tetracycline labeling. An increase in 99mTc labeling of the subchondral bone was the first reliable change observed. This was followed by an increase in tetracycline labeling, bone formation, and a decrease in porosity, which has been associated with relative stiffening of bone. Horizontal splitting and deep fibrillation of the overlying articular cartilage followed the early bone changes. All of these changes preceded changes in content and characterization of cartilage proteoglycans or increased chondrocyte activity as manifested by incorporation of radioactive sulfate. In this model the early bone changes preceded changes in the articular cartilage. The deep splitting of articular cartilage occurred prior to metabolic alteration of that tissue.

Topics: Animals; Calcium Pyrophosphate; Cartilage, Articular; Chondroitin Sulfates; Disease Models, Animal; Female; Glycosaminoglycans; Keratan Sulfate; Knee Joint; Osteoarthritis; Proteoglycans; Rabbits; Synovial Membrane

Endotoxin-induced lung injury has previously been shown to produce lesions that resemble emphysema morphologically and biochemically as demonstrated by the reduction in the content of lung elastin. The purpose of this study was to define the changes in one other connective tissue component, glycosaminoglycans, during the acute phase of the lung injury. Intravenous administration of a single dose of endotoxin in rats resulted in an increase in the total synthesis of glycosaminoglycans by the pulmonary parenchyma. There was a significant increase in the proportion of dermatan sulfate synthesized during the first 48 hr and a concomitant decrease in heparin/heparan sulfate synthesis. At 48 hr the increased synthesis of dermatan sulfate had reached 7.3 times control values and began to decline, whereas the synthesis of chondroitin-4-sulfate rose from 4.1 to 10.7 times control values between 48 and 72 hr. Analysis of the rates of synthesis revealed that the total amount of heparin/heparan sulfate remained constant while the synthesis of chondroitin-6-sulfate increased proportionally to the overall synthesis of glycosaminoglycans. These findings indicate that dramatic changes in glycosaminoglycan synthesis are an integral part of endotoxin lung injury.

Topics: Animals; Chondroitin Sulfates; Dermatan Sulfate; Disease Models, Animal; Endotoxins; Female; Glycosaminoglycans; Heparin; Heparitin Sulfate; Lung; Pulmonary Emphysema; Rats; Rats, Inbred Strains; Time Factors

The pharmacological profile of Org 10172, a mixture of sulphated glycosaminoglycorunans derived from hog intestinal mucosa, has been assessed in experimental thrombosis and bleeding models in rats and compared with heparin USP. Org 10172 inhibited thrombus formation in arterio-venous shunts dose dependently, the dose required for 50% inhibition (ID50) of thrombus formation was 40 anti-Xa units/kg i.v. The ID50 for heparin USP was 70 anti-Xa units/kg i.v. Org 10172 hardly increased bleeding in doses upto 1600 anti-Xa units/kg i.v., whereas heparin USP dose dependently increased bleeding from 90 anti-Xa units/kg i.v. onwards. The benefit (anti-thrombotic)/risk (bleeding) ratio of Org 10172 was therefore considerably better than that of heparin USP. The improved profile of Org 10172 towards bleeding might be caused by differences in the interaction with blood platelets in comparison with heparin USP. Org 10172 had less effect on the platelet content in thrombi than heparin USP. Org 10172 did not inhibit collagen induced release of serotonin in contrast to heparin USP. Org 10172 inhibited factor Xa induced aggregation of rabbit platelets but only at anti-Xa levels which were fifteen times higher than for heparin USP. In contrast to heparin USP Org 10172 had only a very weak effect on the activated partial thromboplastin time (APTT) ex vivo.

Topics: Animals; Blood Coagulation; Blood Platelets; Chondroitin Sulfates; Dermatan Sulfate; Disease Models, Animal; Fibrinolytic Agents; Glycosaminoglycans; Heparin; Heparinoids; Heparitin Sulfate; Male; Platelet Aggregation; Rabbits; Rats; Rats, Inbred Strains; Thrombophlebitis

The experimental models used by investigators to study myocardial infarction have been considered as to their possible application for use in studies of the healing of myocardial infarction. The information concerning healing has also been surveyed. In general, the healing of the myocardial infarct in the dog and in the rat is by connective tissue replacement of the injured tissue resulting in a scar similar to skin scars. In the healing process, there is an early increase of glycoproteins, possibly from serum, and of hyaluronic acid in the injured tissue. Much of this is part of the general reaction to injury and may not be part of the healing process. Somewhat later (about 2-3 days in the dog) the chondroitin-4-sulfate fraction begins to rise. Collagen biosynthesis increases at the same time although this relationship is not well established. Much later (after 30 days in dog) the chondroitin-4-sulfate content of the injured tissue begins to decrease. At this time the scar is well formed. Much later (as late as 171 days) the scar in the myocardium still contains elevated amounts of chondroitin-4-sulfate. The dermatan sulfate is increased and the hyaluronic acid slightly decreased as compared to undamaged myocardium. These changes are typical in maturing scars of skin.

Topics: Animals; Chondroitin Sulfates; Coronary Vessels; Dermatan Sulfate; Disease Models, Animal; Dogs; Glycosaminoglycans; Heart; Humans; Hyaluronic Acid; Middle Aged; Myocardial Infarction; Rats; Wound Healing

The organic matrix of the long bones of the "normal" rat strain and of its mutant Op/Orl were investigated. This latter strain exhibits some anomalles similar to Albers-Schönberg disease as well as tooth retentions. Bones (tibia, femur) were incubated with 14C-glucose and also submitted to a fractional extraction procedure (EDTA, urea) to separate soluble and insoluble fractions. The proteins, hexoses, hexosamines, hydroxyproline content of these fractions was determined as well as the radioactivity of the soluble extracts Glycosaminoglycans were also studied by cellogel electrophoresis after pronase digestion of the EDTA-extracts. Male and female animals of each strain were studied separately. The relative amount of soluble proteins (EDTA + urea extracts) was the same in both strains, the mineral content of the final residue was however higher in the mutant Op/Orl strain. The hydroxyproline content of the mutant-extracts were lower than in the original strain suggesting a lower soluble collagen content. This may be due to a faster polymerisation, insolubillisation of freshly synthesised collagen. The hexosamine content of the mutant urea extracts and the final residue was higher than that found in the analogous extracts obtained from the normal "normal" strain, suggesting a higher proportion of structural glycoproteins in the mutant bone, uronic acid being low or absent. Some other parameters were identical between the mutant and original strains but differed between males and females. The radioactivity of the soluble extracts was higher in the males than in females. The glycosaminoglycans of the EDTA extracts are also different: male-extracts show two bands on cellogel electrophoresis, one corresponding to chondroitin-sulphate, the other to hyaluronate. Females showed only the chondroitin sulphate band. The aminoacid composition of the insoluble residue of males was higher in basic amino acids (lysine, histidine, arginine) than the female extracts. These results indicate the existence of discrete well defined differences between the organic matrix of the orginal and mutant strain on one side and between male and female bones on the other side.

Topics: Amino Acids; Animals; Bone Matrix; Chondroitin Sulfates; Disease Models, Animal; Edetic Acid; Femur; Glycosaminoglycans; Hexosamines; Hexoses; Hyaluronic Acid; Hydroxyproline; Osteopetrosis; Proteins; Rats; Rats, Inbred Strains; Sex Factors; Tibia; Urea

An experimental model of osteoarthritis resulting from laxity of the joint was induced in eighteen mature dogs (at least two years old) by sectioning the anterior cruciate ligament of the right knee (stifle) with a stab incision, the left knee providing a control. A sham operation was also performed in three other dogs, in which a stab incision was made but the ligament left intact. The dogs were killed at various intervals from one to forty-eight weeks later. Morphological changes in bone, cartilage, synovial membrane and joint capsule were examined in all the joints and biochemical changes in the cartilage of three dogs killed after two, eight, and sixteen weeks. All the changes resulting from the operation progressed with time and became indistinguishable from those found in three dogs with natural osteoarthritis of the knee. There were no changes in the joints which had sham operations. As the time of onset is known, this experimental model in a larger species enables a study to be made of the biochemical as well as the morphological changes in the early stages of osteoarthritis.

Topics: Animals; Bone Development; Cartilage, Articular; Chondroitin Sulfates; Disease Models, Animal; Dogs; Female; Femur; Galactosamine; Glucosamine; Keratan Sulfate; Knee Joint; Ligaments, Articular; Male; Menisci, Tibial; Osteoarthritis; Proteoglycans; Stress, Physiological; Synovial Membrane; Tibia; Uronic Acids

Extracts of human peripheral blood polymorphonuclear leukocyte granules, and two purified proteases derived from such extracts, an elastase and a chymotrypsin-like enzyme, degrade isolated bovine nasal cartilage proteoglycan at neutral pH. Viscosity studies indicate that the leukocyte granule extracts lack hyaluronidase activity and that their degradative effect on proteoglycan at physiological pH is due entirely to proteolytic action. Sepharose 4B gel chromatography and SDS-polyacrylamide gel electrophoresis of proteoglycan fractions treated with leukocyte granule enzymes at pH 7.0 indicate that they degrade one of the proteoglycan link proteins, release a fragment from the hyaluronic acid-binding portion of the proteoglycan subunit core protein, and break down the remainder of the proteoglycan subunit molecule into peptide fragments with varying numbers of chondroitin sulfate chains. Immunodiffusion studies indicate that the antigenic determinants of the proteoglycan subunit core protein and the link proteins survive treatment with granule proteases. Similar degradation of human articular cartilage proteoglycan by granule neutral proteases can be presumed to occur, in view of the similarity of structure of human articular and bovine nasal cartilage proteoglycans. The release of granule enzymes in the course of neutrophil-mediated inflammation can thus result in the degradation of cartilage matrix proteoglycan, leading to cartilage destruction and joint injury.

Topics: Animals; Arthritis; Cartilage, Articular; Cattle; Chemical Phenomena; Chemistry; Chondroitin Sulfates; Chromatography, Gel; Chymotrypsin; Cytoplasmic Granules; Disease Models, Animal; Electrophoresis, Polyacrylamide Gel; Glycosaminoglycans; Humans; Immunodiffusion; Joint Diseases; Leukocytes; Nasal Septum; Pancreatic Elastase; Proteoglycans; Sodium Dodecyl Sulfate; Viscosity