heparitin-sulfate and Sepsis

Sepsis-associated encephalopathy (SAE) is characterized by a diffuse cerebral dysfunction that accompanies sepsis in the absence of direct central nervous system infection. The endothelial glycocalyx is a dynamic mesh containing heparan sulfate linked to proteoglycans and glycoproteins, including selectins and vascular/intercellular adhesion molecules (V/I-CAMs), which protects the endothelium while mediating mechano-signal transduction between the blood and vascular wall. During severe inflammatory states, components of the glycocalyx are shed into the circulation and can be detected in soluble forms. Currently, SAE remains a diagnosis of exclusion and limited information is available on the utility of glycocalyx-associated molecules as biomarkers for SAE. We set out to synthesize all available evidence on the association between circulating molecules released from the endothelial glycocalyx surface during sepsis and sepsis-associated encephalopathy.. MEDLINE (PubMed) and EMBASE were searched since inception until May 2, 2022 to identify eligible studies. Any comparative observational study: i) evaluating the association between sepsis and cognitive decline and ii) providing information on level of circulating glycocalyx-associated molecules was eligible for inclusion.. Four case-control studies with 160 patients met the inclusion criteria. Meta-analysis of biomarkers ICAM-1 (SMD 0.41; 95% CI 0.05-0.76; p = 0.03; I2 = 50%) and VCAM-1 (SMD 0.55; 95% CI 0.12-0.98; p = 0.01; I2 = 82%) revealed higher pooled mean concentration in patients with SAE compared to the patients with sepsis alone. Single studies reported elevated levels of P-selectin (MD 0.80; 95% CI -17.77-19.37), E-selectin (MD 96.40; 95% Cl 37.90-154.90), heparan sulfate NS2S (MD 19.41; 95% CI 13.37-25.46), and heparan sulfate NS+NS2S+NS6S (MD 67.00; 95% CI 31.00-103.00) in patients with SAE compared to the patients with sepsis alone.. Plasma glycocalyx-associated molecules are elevated in SAE and may be useful for early identification of cognitive decline in sepsis patients.

Topics: Biomarkers; Cell Adhesion Molecules; Glycocalyx; Heparitin Sulfate; Humans; Observational Studies as Topic; Sepsis; Sepsis-Associated Encephalopathy

The endothelial glycocalyx has a profound influence at the vascular wall on the transmission of shear stress, on the maintenance of a selective permeability barrier and a low hydraulic conductivity, and on attenuating firm adhesion of blood leukocytes and platelets. Major constituents of the glycocalyx, including syndecans, heparan sulphates and hyaluronan, are shed from the endothelial surface under various acute and chronic clinical conditions, the best characterized being ischaemia and hypoxia, sepsis and inflammation, atherosclerosis, diabetes, renal disease and haemorrhagic viral infections. Damage has also been detected by in vivo microscopic techniques. Matrix metalloproteases may shed syndecans and heparanase, released from activated mast cells, cleaves heparan sulphates from core proteins. According to new data, not only hyaluronidase but also the serine proteases thrombin, elastase, proteinase 3 and plasminogen, as well as cathepsin B lead to loss of hyaluronan from the endothelial surface layer, suggesting a wide array of potentially destructive conditions. Appropriately, pharmacological agents such as inhibitors of inflammation, antithrombin and inhibitors of metalloproteases display potential to attenuate shedding of the glycocalyx in various experimental models. Also, plasma components, especially albumin, stabilize the glycocalyx and contribute to the endothelial surface layer. Though symptoms of the above listed diseases and conditions correlate with sequelae expected from disturbance of the endothelial glycocalyx (oedema, inflammation, leukocyte and platelet adhesion, low reflow), therapeutic studies to prove a causal connection have yet to be designed. With respect to studies on humans, some clinical evidence exists for benefits from application of sulodexide, a preparation delivering precursors of the glycocalyx constituent heparan sulphate. At present, the simplest option for protecting the glycocalyx seems to be to ensure an adequate level of albumin. However, also in this case, definite proof of causality needs to be delivered.

Topics: Animals; Anti-Inflammatory Agents; Endothelium, Vascular; Enzyme Inhibitors; Glycocalyx; Glycosaminoglycans; Heparitin Sulfate; Humans; Hyaluronic Acid; Peptide Hydrolases; Renal Insufficiency; Reperfusion Injury; Sepsis; Serum Albumin; Syndecans

Pasteurella (P) multocida exists in a variety of animals and causes diverse infections in humans due to animal bites and scratches, usually by cats or dogs, and oral and respiratory infection. We report a case of P multocida sepsis due to a scratch from a pet cat, complicated with disseminated intravascular coagulation in a post-chemotherapy neutropenic patient with non-Hodgkin lymphoma. The patient was a febrile 79-year-old woman with disturbed consciousness and subcutaneous abscess in her right hand due to a scratch from a pet cat. She was successfully treated with empirical antibiotic therapy with cefepime and administrations of granulocyte colony-stimulating factor and danaparoid. The minimum inhibitory concentration of cefepime against the isolate from this case was <2mg/L. Although a few days are required before a diagnosis of P multocida infection can be made from a bacteriological study, the infection can be successfully treated against febrile neutropenia with empirical cefepime. In a literature review, 7 cases, including ours, with hematological malignancies complicated with P multocida infection were identified and we summarized the clinical characteristics of these cases. These cases demonstrate the importance of the prevention of close contact between pet animals and immunocompromised hosts such as post-chemotherapy neutropenic patients.

Topics: Aged; Animals; Animals, Domestic; Anti-Bacterial Agents; Anticoagulants; Bites and Stings; Cats; Cefepime; Cephalosporins; Chondroitin Sulfates; Dermatan Sulfate; Female; Granulocyte Colony-Stimulating Factor; Heparitin Sulfate; Humans; Lymphoma, Non-Hodgkin; Neutropenia; Pasteurella Infections; Pasteurella multocida; Sepsis

TLRs are usually thought to recognize substances produced by microorganisms and thus, to initiate host defenses. This concept, however, fails to explain some functions of this family of receptors. Recognition of endogenous substances may explain the broader functions of TLRs in physiology and disease. Activation of TLRs by endogenous substances necessitates vigorous control of the function of the receptors. This communication will summarize a line of research, which points to an endogenous agonist for TLR4 and a putative mechanism for controlling the function of that receptor.

Topics: Animals; Heparitin Sulfate; Humans; Sepsis; Systemic Inflammatory Response Syndrome; Toll-Like Receptors

Myocardial dysfunction in sepsis has been linked to inflammation caused by pathogen-associated molecular patterns (PAMPs) as well as by host danger-associated molecular patterns (DAMPs). These include soluble heparan sulfate (HS), which triggers the devastating consequences of the pro-inflammatory cascades in severe sepsis and septic shock. Thus, there is increasing interest in the development of anti-infective agents, with effectiveness against both PAMPs and DAMPs. We hypothesized that a synthetic antimicrobial peptide (peptide 19-2.5) inhibits inflammatory response in murine cardiomyocytes (HL-1 cells) stimulated with PAMPs, DAMPs or serum from patients with septic shock by reduction and/or neutralization of soluble HS. In the current study, our data indicate that the treatment with peptide 19-2.5 decreases the inflammatory response in HL-1 cells stimulated with either PAMPs or DAMPs. Furthermore, our work shows that soluble HS in serum from patients with Gram-negative or Gram-positive septic shock induces a strong pro-inflammatory response in HL-1 cells, which can be effectively blocked by peptide 19-2.5. Based on these findings, peptide 19-2.5 is a novel anti-inflammatory agent interacting with both PAMPs and DAMPs, suggesting peptide 19-2.5 may have the potential for further development as a broad-spectrum anti-inflammatory agent in sepsis-induced myocardial inflammation and dysfunction.

Topics: Aged; Animals; Antimicrobial Cationic Peptides; Cell Line; Female; Heparitin Sulfate; Humans; Inflammation; Male; Mice; Middle Aged; Myocytes, Cardiac; Sepsis; Serum

Sepsis is a lethal syndrome manifested by an unregulated, overwhelming inflammation from the host in response to infection. Here, we exploit the use of a synthetic heparan sulfate octadecasaccharide (18-mer) to protect against sepsis. The 18-mer not only inhibits the pro-inflammatory activity of extracellular histone H3 and high mobility group box 1 (HMGB1), but also elicits the anti-inflammatory effect from apolipoprotein A-I (ApoA-I). We demonstrate that the 18-mer protects against sepsis-related injury and improves survival in cecal ligation and puncture mice and reduces inflammation in an endotoxemia mouse model. The 18-mer neutralizes the cytotoxic histone-3 (H3) through direct interaction with the protein. Furthermore, the 18-mer enlists the actions of ApoA-I to dissociate the complex of HMGB1 and lipopolysaccharide, a toxic complex contributing to cell death and tissue damage in sepsis. Our study provides strong evidence that the 18-mer mitigates inflammatory damage in sepsis by targeting numerous mediators, setting it apart from other potential therapies with a single target.

Topics: Animals; Apolipoprotein A-I; Disease Models, Animal; Endotoxemia; Heparitin Sulfate; HMGB1 Protein; Lipopolysaccharides; Mice; Sepsis

Heparan sulfate proteoglycans (HSPGs) are at the forefront of host-microbe interactions. Cell surface HSPGs are thought to promote infection as attachment and internalization receptors for many bacterial pathogens and as soluble inhibitors of host immunity when released from the cell surface by ectodomain shedding. However, the importance of HSPG-pathogen interactions in vivo has yet to be clearly established. Here we describe several representative methods to study the role of HSPGs in systemic bacterial infections, such as bacteremia and sepsis. The overall experimental strategy is to use mouse models to establish the physiological significance of HSPGs, to determine the identity of HSPGs that specifically promote infection, and to define key structural features of HSPGs that enhance bacterial virulence in systemic infections.

Topics: Animals; Bacterial Infections; Cell Membrane; Disease Models, Animal; Heparan Sulfate Proteoglycans; Heparitin Sulfate; Mice; Sepsis

Angiopoietin-2 (Ang-2) is a key mediator of vascular disease during sepsis, and elevated plasma levels of Ang-2 are associated with organ injury scores and poor clinical outcomes. We have previously observed that biomarkers of endothelial glycocalyx (EG) damage correlate with plasma Ang-2 levels, suggesting a potential mechanistic linkage between EG injury and Ang-2 expression during states of systemic inflammation. However, the cell signaling mechanisms regulating Ang-2 expression following EG damage are unknown. In the current study, we determined the temporal associations between plasma heparan sulfate (HS) levels as a marker of EG erosion and plasma Ang-2 levels in children with sepsis and in mouse models of sepsis. Second, we evaluated the role of shear stress-mediated 5'-adenosine monophosphate-activated protein kinase (AMPK) signaling in Ang-2 expression following enzymatic HS cleavage from the surface of human primary lung microvascular endothelial cells (HLMVECs). We found that plasma HS levels peaked before plasma Ang-2 levels in children and mice with sepsis. Further, we discovered that impaired AMPK signaling contributed to increased Ang-2 expression following HS cleavage from flow-conditioned HLMVECs, establishing a paradigm by which Ang-2 may be upregulated during sepsis.

Topics: AMP-Activated Protein Kinases; Angiopoietin-2; Animals; Biomarkers; Child; Endothelial Cells; Forkhead Box Protein O1; Glycocalyx; Heparitin Sulfate; Humans; Mice; Sepsis; Signal Transduction

Endotheliopathy is a key element in COVID-19 pathophysiology, contributing to both morbidity and mortality. Biomarkers distinguishing different COVID-19 phenotypes from sepsis syndrome remain poorly understood.. To characterize circulating biomarkers of endothelial damage in different COVID-19 clinical disease stages compared with sepsis syndrome and normal volunteers.. Patients with COVID-19 pneumonia (n = 49) were classified into moderate, severe, or critical (life-threatening) disease. Plasma samples were collected within 48 to 72 h of hospitalization to analyze endothelial activation markers, including soluble Vascular Cell Adhesion Molecule-1 (sVCAM-1), von Willebrand Factor (VWF), A disintegrin-like and metalloprotease with thrombospondin type 1 motif no. 13 (ADAMTS-13) activity, thrombomodulin (TM), and soluble TNF receptor I (sTNFRI); heparan sulfate (HS) for endothelial glycocalyx degradation; C5b9 deposits on endothelial cells in culture and soluble C5b9 for complement activation; circulating dsDNA for neutrophil extracellular traps (NETs) presence, and α2-antiplasmin and PAI-1 as parameters of fibrinolysis. We compared the level of each biomarker in all three COVID-19 groups and healthy donors as controls (n = 45). Results in critically ill COVID-19 patients were compared with other intensive care unit (ICU) patients with septic shock (SS, n = 14), sepsis (S, n = 7), and noninfectious systemic inflammatory response syndrome (NI-SIRS, n = 7).. All analyzed biomarkers were increased in COVID-19 patients versus controls (P < 0.001), except for ADAMTS-13 activity that was normal in both groups. The increased expression of sVCAM-1, VWF, sTNFRI, and HS was related to COVID-19 disease severity (P < 0.05). Several differences in these parameters were found between ICU groups: SS patients showed significantly higher levels of VWF, TM, sTNFRI, and NETS compared with critical COVID-19 patients and ADAMTS-13 activity was significantly lover in SS, S, and NI-SIRS versus critical COVID-19 (P < 0.001). Furthermore, α2-antiplasmin activity was higher in critical COVID-19 versus NI-SIRS (P < 0.01) and SS (P < 0.001), whereas PAI-1 levels were significantly lower in COVID-19 patients compared with NI-SIRS, S, and SS patients (P < 0.01).. COVID-19 patients present with increased circulating endothelial stress products, complement activation, and fibrinolytic dysregulation, associated with disease severity. COVID-19 endotheliopathy differs from SS, in which endothelial damage is also a critical feature of pathobiology. These biomarkers could help to stratify the severity of COVID-19 disease and may also provide information to guide specific therapeutic strategies to mitigate endotheliopathy progression.

Topics: ADAMTS13 Protein; Aged; alpha-2-Antiplasmin; Biomarkers; Complement Membrane Attack Complex; COVID-19; DNA; Female; Heparitin Sulfate; Humans; Male; Middle Aged; Patient Acuity; Plasminogen Activator Inhibitor 1; Prospective Studies; Receptors, Tumor Necrosis Factor, Type I; Sepsis; Thrombomodulin; Vascular Cell Adhesion Molecule-1; von Willebrand Factor

Topics: Animals; Disease Models, Animal; Endothelial Cells; Female; Glycocalyx; Heparitin Sulfate; Liver; Lung; Male; Mice; Mice, Inbred C57BL; Neutrophil Activation; Neutrophils; Sepsis; Staphylococcus aureus

Topics: Anticoagulants; COVID-19 Drug Treatment; Critical Illness; Glycosaminoglycans; Heparin; Heparitin Sulfate; Humans; Inflammation; Protein Binding; Pulmonary Gas Exchange; Sepsis

The endothelial glycocalyx (eGC), a carbohydrate-rich layer lining the luminal surface of the endothelium, provides a first vasoprotective barrier against vascular leakage in sepsis. We hypothesized that angiopoietin-2 (Angpt-2), antagonist of the endothelium-stabilizing receptor Tie2, induces a rapid loss of the eGC in human sepsis. Using intravital microscopy, we measured the perfused boundary region (PBR), an inverse parameter of eGC dimensions in sublingual microvessels, in patients with sepsis and age-matched nonseptic subjects. Median PBR values were significantly higher in patients compared with controls and correlated with serum Angpt-2 levels. To transfer and further explore these findings in a cell culture system, we exposed endothelial cells (ECs) to serum (5%) from a subgroup of septic patients and nonseptic controls. Confocal and atomic force microscopy revealed that sepsis serum, but not control serum, induced thinning of the eGC on human ECs in vitro, which correlated with paired PBR values obtained in vivo (

Topics: Adult; Aged; Angiopoietin-2; Case-Control Studies; Cell Line; Endothelial Cells; Female; Glucuronidase; Glycocalyx; Heparitin Sulfate; Humans; Intravital Microscopy; Male; Microscopy, Atomic Force; Middle Aged; Phosphorylation; Prospective Studies; Receptor, TIE-2; Sepsis; Signal Transduction

The endothelial glycocalyx and its regulated shedding are important to vascular health. Endo-β-D-glucuronidase heparanase-1 (HPSE1) is the only enzyme that can shed heparan sulfate. However, the mechanisms are not well understood. We show that HPSE1 activity aggravated Toll-like receptor 4 (TLR4)-mediated response of endothelial cells to LPS. On the contrary, overexpression of its endogenous inhibitor, heparanase-2 (HPSE2) was protective. The microfluidic chip flow model confirmed that HPSE2 prevented heparan sulfate shedding by HPSE1. Furthermore, heparan sulfate did not interfere with cluster of differentiation-14 (CD14)-dependent LPS binding, but instead reduced the presentation of the LPS to TLR4. HPSE2 reduced LPS-mediated TLR4 activation, subsequent cell signalling, and cytokine expression. HPSE2-overexpressing endothelial cells remained protected against LPS-mediated loss of cell-cell contacts. In vivo, expression of HPSE2 in plasma and kidney medullary capillaries was decreased in mouse sepsis model. We next applied purified HPSE2 in mice and observed decreases in TNFα and IL-6 plasma concentrations after intravenous LPS injections. Our data demonstrate the important role of heparan sulfate and the glycocalyx in endothelial cell activation and suggest a protective role of HPSE2 in microvascular inflammation. HPSE2 offers new options for protection against HPSE1-mediated endothelial damage and preventing microvascular disease.

Topics: Animals; Disease Models, Animal; Endothelial Cells; Glucuronidase; Glycocalyx; Heparitin Sulfate; Humans; Lipopolysaccharides; Male; Mice; Microfluidic Analytical Techniques; Sepsis; Signal Transduction; Toll-Like Receptor 4

Septic patients frequently develop cognitive impairment that persists beyond hospital discharge. The impact of sepsis on electrophysiological and molecular determinants of learning is underexplored. We observed that mice that survived sepsis or endotoxemia experienced loss of hippocampal long-term potentiation (LTP), a brain-derived neurotrophic factor-mediated (BDNF-mediated) process responsible for spatial memory formation. Memory impairment occurred despite preserved hippocampal BDNF content and could be reversed by stimulation of BDNF signaling, suggesting the presence of a local BDNF inhibitor. Sepsis is associated with degradation of the endothelial glycocalyx, releasing heparan sulfate fragments (of sufficient size and sulfation to bind BDNF) into the circulation. Heparan sulfate fragments penetrated the hippocampal blood-brain barrier during sepsis and inhibited BDNF-mediated LTP. Glycoarray approaches demonstrated that the avidity of heparan sulfate for BDNF increased with sulfation at the 2-O position of iduronic acid and the N position of glucosamine. Circulating heparan sulfate in endotoxemic mice and septic humans was enriched in 2-O- and N-sulfated disaccharides; furthermore, the presence of these sulfation patterns in the plasma of septic patients at intensive care unit (ICU) admission predicted persistent cognitive impairment 14 days after ICU discharge or at hospital discharge. Our findings indicate that circulating 2-O- and N-sulfated heparan sulfate fragments contribute to septic cognitive impairment.

Topics: Animals; Brain-Derived Neurotrophic Factor; Cognitive Dysfunction; Female; Heparitin Sulfate; Hippocampus; Long-Term Potentiation; Male; Memory Disorders; Mice; Sepsis

Survivors of sepsis and other forms of critical illness frequently experience significant and disabling cognitive and affective disorders. Inflammation, ischemia, and glial cell dysfunction contribute to this persistent brain injury. In this issue of the JCI, Hippensteel et al. show that endothelial injury in animal models of sepsis or endotoxemia leads to shedding of heparan fragments from the endothelial glycocalyx. These fragments directly sequester brain-derived neurotrophic factor and impair hippocampal long-term potentiation, an electrophysiologic correlate of memory. The authors further explore the specific characteristics of heparan fragments that bind neurotrophins and the presence of these fragments in the circulation of patients who survive sepsis. This study highlights an important mechanism by which vascular injury can impair brain function.

Topics: Animals; Cognitive Dysfunction; Glycocalyx; Heparitin Sulfate; Humans; Poisons; Sepsis

Intravenous fluids, an essential component of sepsis resuscitation, may paradoxically worsen outcomes by exacerbating endothelial injury. Preclinical models suggest that fluid resuscitation degrades the endothelial glycocalyx, a heparan sulfate-enriched structure necessary for vascular homeostasis. We hypothesized that endothelial glycocalyx degradation is associated with the volume of intravenous fluids administered during early sepsis resuscitation.. We used mass spectrometry to measure plasma heparan sulfate (a highly sensitive and specific index of systemic endothelial glycocalyx degradation) after 6 h of intravenous fluids in 56 septic shock patients, at presentation and after 24 h of intravenous fluids in 100 sepsis patients, and in two groups of non-infected patients. We compared plasma heparan sulfate concentrations between sepsis and non-sepsis patients, as well as between sepsis survivors and sepsis non-survivors. We used multivariable linear regression to model the association between volume of intravenous fluids and changes in plasma heparan sulfate.. Consistent with previous studies, median plasma heparan sulfate was elevated in septic shock patients (118 [IQR, 113-341] ng/ml 6 h after presentation) compared to non-infected controls (61 [45-79] ng/ml), as well as in a second cohort of sepsis patients (283 [155-584] ng/ml) at emergency department presentation) compared to controls (177 [144-262] ng/ml). In the larger sepsis cohort, heparan sulfate predicted in-hospital mortality. In both cohorts, multivariable linear regression adjusting for age and severity of illness demonstrated a significant association between volume of intravenous fluids administered during resuscitation and plasma heparan sulfate. In the second cohort, independent of disease severity and age, each 1 l of intravenous fluids administered was associated with a 200 ng/ml increase in circulating heparan sulfate (p = 0.006) at 24 h after enrollment.. Glycocalyx degradation occurs in sepsis and septic shock and is associated with in-hospital mortality. The volume of intravenous fluids administered during sepsis resuscitation is independently associated with the degree of glycocalyx degradation. These findings suggest a potential mechanism by which intravenous fluid resuscitation strategies may induce iatrogenic endothelial injury.

Topics: Administration, Intravenous; Adult; Aged; Angiopoietin-2; Atrial Natriuretic Factor; Biomarkers; Endothelium; Female; Fluid Therapy; Glycocalyx; Heparitin Sulfate; Humans; Male; Mass Spectrometry; Middle Aged; Natriuretic Peptide, Brain; Resuscitation; Sepsis; Syndecan-1; Thrombomodulin; Tissue Plasminogen Activator; Vascular Endothelial Growth Factor Receptor-1

Life-threatening cardiomyopathy is a severe, but common, complication associated with severe trauma or sepsis. Several signaling pathways involved in apoptosis and necroptosis are linked to trauma- or sepsis-associated cardiomyopathy. However, the underling causative factors are still debatable. Heparan sulfate (HS) fragments belong to the class of danger/damage-associated molecular patterns liberated from endothelial-bound proteoglycans by heparanase during tissue injury associated with trauma or sepsis. We hypothesized that HS induces apoptosis or necroptosis in murine cardiomyocytes. By using a novel Medical-

Topics: Algorithms; Animals; Apoptosis; Cardiomyopathies; Caspase 3; Cell Culture Techniques; Cells, Cultured; Cytochromes c; Heparitin Sulfate; Humans; Machine Learning; Mice; Myocytes, Cardiac; Necrosis; Receptor-Interacting Protein Serine-Threonine Kinases; Sepsis; Signal Transduction; Wounds and Injuries

The endothelial glycocalyx is a glycosaminoglycan-enriched endovascular layer that, with the development of novel fixation and in vivo microscopy techniques, has been increasingly recognized as a major contributor to vascular homeostasis. Sepsis-associated degradation of the endothelial glycocalyx mediates the onset of the alveolar microvascular dysfunction characteristic of sepsis-induced lung injury (such as the Acute Respiratory Distress Syndrome, ARDS). Emerging evidence indicates that processes of glycocalyx reconstitution are necessary for endothelial repair and, as such, are promising therapeutic targets to accelerate lung injury recovery. This review discusses what has been learned about the homeostatic and pathophysiologic role of the pulmonary endothelial glycocalyx during lung health and injury, with the goal to identify promising new areas for future mechanistic investigation.

Topics: Endothelium, Vascular; Glucuronidase; Glycocalyx; Heparitin Sulfate; Humans; Lung; Lung Injury; Respiratory Distress Syndrome; Sepsis

Topics: Animals; Glycocalyx; Heparitin Sulfate; Humans; Lung; Respiratory Distress Syndrome; Sepsis; Syndecan-1

Heparanase, a heparan sulfate (HS)-specific endoglycosidase, plays an important role in inflammation and mediates acute pulmonary and renal injuries during sepsis. To explore its role in septic intestinal injury, a non-anticoagulant heparanase inhibitor, N-desulfated/re- N-acetylated heparin (NAH), was administrated to a mouse sepsis model induced by cecal ligation and puncture (CLP). Immunohistochemical staining revealed massive shedding of HS from the intestinal mucosal surfaces after CLP, and effective inhibition of heparanase by NAH was confirmed by markedly reduced HS shedding. Following CLP, intestinal expression of heparanase was increased, whereas pretreatment with NAH reduced the sepsis-induced upregulation of heparanase expression. Meanwhile, CLP led to shedding of syndecan-1 and upregulated expression of proteases such as matrix metalloprotease-9 and urokinase-type plasminogen activator in the intestine, whereas NAH markedly suppressed syndecan-1 shedding and protease upregulation following CLP. In addition, pretreatment with NAH attenuated intestinal injury, inhibited neutrophil infiltration and suppressed the production of inflammatory cytokines (tumor necrosis factor-α, interleukin-1β, and interleukin-6) in the intestine during sepsis, and it also significantly reduced the elevation of inflammatory cytokines in the serum 24 hr after CLP. Our findings demonstrate that the activation of intestinal heparanase contributes to intestinal injury during early sepsis by facilitating the destruction of mucosal epithelial glycocalyx and promoting inflammatory responses.

Topics: Animals; Cytokines; Glucuronidase; Heparin; Heparitin Sulfate; Interleukin-1beta; Interleukin-6; Intestinal Mucosa; Intestines; Male; Mice, Inbred C57BL; Neutrophil Infiltration; Sepsis; Syndecan-1; Tumor Necrosis Factor-alpha; Up-Regulation

The endothelial glycocalyx is a heparan sulfate (HS)-rich endovascular structure critical to endothelial function. Accordingly, endothelial glycocalyx degradation during sepsis contributes to tissue edema and organ injury. We determined the endogenous mechanisms governing pulmonary endothelial glycocalyx reconstitution, and if these reparative mechanisms are impaired during sepsis. We performed intravital microscopy of wild-type and transgenic mice to determine the rapidity of pulmonary endothelial glycocalyx reconstitution after nonseptic (heparinase-III mediated) or septic (cecal ligation and puncture mediated) endothelial glycocalyx degradation. We used mass spectrometry, surface plasmon resonance, and in vitro studies of human and mouse samples to determine the structure of HS fragments released during glycocalyx degradation and their impact on fibroblast growth factor receptor (FGFR) 1 signaling, a mediator of endothelial repair. Homeostatic pulmonary endothelial glycocalyx reconstitution occurred rapidly after nonseptic degradation and was associated with induction of the HS biosynthetic enzyme, exostosin (EXT)-1. In contrast, sepsis was characterized by loss of pulmonary EXT1 expression and delayed glycocalyx reconstitution. Rapid glycocalyx recovery after nonseptic degradation was dependent upon induction of FGFR1 expression and was augmented by FGF-promoting effects of circulating HS fragments released during glycocalyx degradation. Although sepsis-released HS fragments maintained this ability to activate FGFR1, sepsis was associated with the downstream absence of reparative pulmonary endothelial FGFR1 induction. Sepsis may cause vascular injury not only via glycocalyx degradation, but also by impairing FGFR1/EXT1-mediated glycocalyx reconstitution.

Topics: Animals; Cecum; Endothelium, Vascular; Fibroblast Growth Factor 2; Glycocalyx; Heparitin Sulfate; Homeostasis; Ligation; Lung; Male; Mice, Inbred C57BL; N-Acetylglucosaminyltransferases; Polysaccharide-Lyases; Punctures; Receptor, Fibroblast Growth Factor, Type 1; Sepsis; Signal Transduction

Malfunctioning of the intestinal microcirculation secondary to severe acute pancreatitis (SAP) can cause injuries to the intestinal mucosal barrier, translocation of gut flora, and sepsis. The glycocalyx on the vascular endothelium helps maintain its normal function through multiple mechanisms, including regulation of vascular permeability and inhibition of intercellular adhesion. It is unknown that whether pancreatitis inflicts injuries to the intestinal mucosal barrier through damaging glycocalyx or stabilizing glycocalyx can be a potential therapeutic target in maintaining the integrity of the intestinal mucosal barrier during SAP. Injecting sodium taurocholate into the pancreatic duct of Sprague-Dawley rats induced SAP. Intestinal perfusion, changes in endothelial glycocalyx, and the associated molecular mechanisms were assessed by laser Doppler velocimetry, electron microscopy, and the levels of heparan sulfate, syndacan-1, and tumor necrosis factor-α (TNF-α) in the superior mesenteric vein. Protective effects of hydrocortisone treatment in the intestinal microcirculation during SAP were evaluated. Degradation of the glycocalyx in intestinal vascular endothelium developed 3 h after the onset of SAP in rats. By 12 h, significant reduction of intestinal perfusion was observed. The concomitant elevated levels of TNF-α in the superior mesenteric vein suggest that TNF-α is involved in the degradation of the glycocalyx. With the use of hydrocortisone, intestinal perfusion was improved and the degradation of glycocalyx was reduced. The degradation of glycocalyx is involved in the malfunction of the intestinal microcirculation. The massive release of TNF-α participates in this process and leads to glycocalyx degradation. Hydrocortisone may be a good therapy to prevent this process.

Topics: Animals; Cell Adhesion; Disease Models, Animal; Endothelium, Vascular; Glycocalyx; Heparitin Sulfate; Hydrocortisone; Intestines; Male; Mesenteric Veins; Microcirculation; Pancreatitis; Perfusion; Permeability; Rats; Rats, Sprague-Dawley; Sepsis; Syndecan-1; Taurocholic Acid; Time Factors; Tumor Necrosis Factor-alpha

The heart is one of the most frequently affected organs in sepsis. Recent studies focused on lipopolysaccharide-induced mitochondrial dysfunction; however myocardial dysfunction is not restricted to gram-negative bacterial sepsis. The purpose of this study was to investigate circulating heparan sulfate (HS) as an endogenous danger associated molecule causing cardiac mitochondrial dysfunction in sepsis. We used an in vitro model with native sera (SsP) and sera eliminated from HS (HS-free), both of septic shock patients, to stimulate murine cardiomyocytes. As determined by extracellular flux analyzing, SsP increased basal mitochondrial respiration, but reduced maximum mitochondrial respiration, compared with unstimulated cells (P < 0.0001 and P < 0.0001, respectively). Cells stimulated with HS-free serum revealed unaltered basal and maximum mitochondrial respiration, compared with unstimulated cells (P = 0.1174 and P = 0.8992, respectively). Cellular ATP-level were decreased in SsP-stimulated cells but unaltered in cells stimulated with HS-free serum compared with unstimulated cells (P < 0.0001 and P = 0.1593, respectively). Live-cell imaging revealed an increased production of mitochondrial reactive oxygen species in cells stimulated with SsP compared with cells stimulated with HS-free serum (P < 0.0001). Expression of peroxisome proliferator-activated receptors (PPARα and PPARγ) and their co-activators PGC-1α, which regulate mitochondrial function, were studied using PCR. Cells stimulated with SsP showed downregulated PPARs and PGC-1α mRNA-levels compared with HS-free serum (P = 0.0082, P = 0.0128, and P = 0.0185, respectively). Blocking Toll-like receptor 4 revealed an inhibition of HS-dependent downregulation of PPARs and PGC-1α (all P < 0.0001). In conclusion, circulating HS in serum of septic shock patients cause cardiac mitochondrial dysfunction, suggesting that HS may be targets of therapeutics in septic cardiomyopathy.

Topics: Adenosine Triphosphate; Aged; Animals; Down-Regulation; Enzyme-Linked Immunosorbent Assay; Female; Heparitin Sulfate; Humans; Male; Mice; Middle Aged; Mitochondria; Myocytes, Cardiac; Oxygen Consumption; Peroxisome Proliferator-Activated Receptor Gamma Coactivator 1-alpha; PPAR alpha; PPAR gamma; Reactive Oxygen Species; Sepsis; Shock, Septic; Transcription Factors

Heparanase is an endo-β-glucuronidase that cleaves heparan sulfate side chains from their proteoglycans. Thereby, heparanase liberates highly potent circulating heparan sulfate-fragments (HS-fragments) and triggers the fatal and excessive inflammatory response in sepsis. As a potential anti-inflammatory agent for sepsis therapy, peptide 19-2.5 belongs to the class of synthetic anti-lipopolysaccharide peptides; however, its activity is not restricted to Gram-negative bacterial infection. We hypothesized that peptide 19-2.5 interacts with heparanase and/or HS, thereby reducing the levels of circulating HS-fragments in murine and human sepsis. Our data indicate that the treatment of septic mice with peptide 19-2.5 compared to untreated control animals lowers levels of plasma heparanase and circulating HS-fragments and reduces heparanase activity. Additionally, mRNA levels of heparanase in heart, liver, lung, kidney and spleen are downregulated in septic mice treated with peptide 19-2.5 compared to untreated control animals. In humans, plasma heparanase level and activity are elevated in septic shock. The ex vivo addition of peptide 19-2.5 to plasma of septic shock patients decreases heparanase activity but not heparanase level. Isothermal titration calorimetry revealed a strong exothermic reaction between peptide 19-2.5 and heparanase and HS-fragments. However, a saturation character has been identified only in the peptide 19-2.5 and HS interaction. In conclusion, the findings of our current study indicate that peptide 19-2.5 interacts with heparanase, which is elevated in murine and human sepsis and consecutively attenuates the generation of circulating HS-fragments in systemic inflammation. Thus, peptide 19-2.5 seems to be a potential anti-inflammatory agent in sepsis.

Topics: Adult; Animals; Anti-Infective Agents; Enzyme-Linked Immunosorbent Assay; Glucuronidase; Heparitin Sulfate; Humans; Mice; Peptides; Sepsis

Glycosaminoglycans (GAGs) are negatively charged polysaccharides present, e.g., on the luminal face of the blood vessels as heparan sulphate (HS) and hyaluronic acid (HA), in the interstitium as HA, and in neutrofils and plasma as chondroitin sulphate (CS) and HA. Total plasma levels of GAG are increased in human septic shock, but the origin and pathophysiological implications are unclear. In order to determine the source of circulating GAG in sepsis, we compared plasma levels of HS, HA, CS and keratan sulphate (KS) in patients with septic shock and controls.. HS and KS were measured with enzyme-linked immunosorbent assay, and HA and CS disaccharides with liquid chromatography tandem mass spectrometry in plasma obtained from patients admitted to intensive care fulfilling criteria for septic shock as well as from matched control patients scheduled for neurosurgery.. Median levels of HS and HA were fourfold increased in septic shock and were higher in patients that did not survive 90 days (threefold and fivefold for HS and HA, respectively). Median CS levels were unaltered, while KS levels were slightly decreased in sepsis patients. HS and HA levels correlated with levels of interleukin-6 and interleukin-10. Except for HA, GAG levels did not correlate to liver or kidney sequential organ function score.. Median plasma level of HS and HA is increased in septic shock patients, are higher in patients that do not survive, and correlates with inflammatory activation and failing circulation. The increased levels could be due to vascular damage.

Topics: Adult; Aged; Aged, 80 and over; C-Reactive Protein; Chondroitin Sulfates; Disaccharides; Enzyme-Linked Immunosorbent Assay; Female; Glycosaminoglycans; Heparitin Sulfate; Humans; Hyaluronic Acid; Indicators and Reagents; Interleukin-10; Interleukin-6; Keratan Sulfate; Male; Middle Aged; Multiple Organ Failure; Peroxidase; Sepsis; Shock, Septic; Survival

We recently showed that the heparan sulfate proteoglycan syndecan-1 mediates hepatic clearance of triglyceride-rich lipoproteins in mice based on systemic deletion of syndecan-1 and hepatocyte-specific inactivation of sulfotransferases involved in heparan sulfate biosynthesis. Here, we show that syndecan-1 expressed on primary human hepatocytes and Hep3B human hepatoma cells can mediate binding and uptake of very low density lipoprotein (VLDL). Syndecan-1 also undergoes spontaneous shedding from primary human and murine hepatocytes and Hep3B cells. In human cells, phorbol myristic acid induces syndecan-1 shedding, resulting in accumulation of syndecan-1 ectodomains in the medium. Shedding occurs through a protein kinase C-dependent activation of ADAM17 (a disintegrin and metalloproteinase 17). Phorbol myristic acid stimulation significantly decreases DiD (1,1'-dioctadecyl-3,3,3',3'-tetramethylindodicarbocyanine perchlorate)-VLDL binding to cells, and shed syndecan-1 ectodomains bind to VLDL. Although mouse hepatocytes appear resistant to induced shedding in vitro, injection of lipopolysaccharide into mice results in loss of hepatic syndecan-1, accumulation of ectodomains in the plasma, impaired VLDL catabolism, and hypertriglyceridemia.. These findings suggest that syndecan-1 mediates hepatic VLDL turnover in humans as well as in mice and that shedding might contribute to hypertriglyceridemia in patients with sepsis.

Topics: Animals; Carcinogens; Cell Line, Tumor; Heparitin Sulfate; Hepatocytes; Humans; Hypertriglyceridemia; Lipopolysaccharides; Lipoproteins, VLDL; Liver Neoplasms; Mice; Mice, Inbred C57BL; Mice, Mutant Strains; Protein Kinase C; Sepsis; Syndecan-1; Tetradecanoylphorbol Acetate; Triglycerides

Recent evidence suggests that the endothelial glycocalix plays an important role in lethal outcomes following sepsis. We therefore tested if the endothelial glycocalix is shed in patients with sepsis compared with patients after major abdominal surgery and healthy volunteers.. A total of 150 individuals were tested for levels of inflammatory markers (intercellular adhesion molecule-1 [ICAM-1], vascular cell adhesion molecule-1 [VCAM-1], interleukin-6 [IL-6]) and glycocalix markers (syndecan-1, heparan sulfate). Three groups consisted of patients with severe sepsis or septic shock, patients after major abdominal surgery without systemic inflammatory response syndrome, and healthy volunteers. Blood was drawn, at the time of diagnosis or surgery, and 6, 24, and 48h later. We correlated these markers to each other and to clinically used inflammation markers.. Levels of inflammatory markers were markedly higher in patients with sepsis compared with patients after major abdominal surgery and healthy volunteers. After major abdominal surgery, glycocalix markers in human plasma were at levels comparable to patients with sepsis. In patients with sepsis, levels of IL-6 correlated with syndecan-1, ICAM-1, VCAM-1, and lactate, while ICAM-1 furthermore correlated with CRP and lactate levels.. High levels of glycocalix markers indicated that significant flaking of the endothelial glycocalix occurred in patients with sepsis, and to a lesser extent in patients after major abdominal surgery. This novel finding could explain the nonspecific capillary leaking syndrome of patients with sepsis and after major abdominal surgery, and may identify new targets for treating those patient populations.

Topics: Abdomen; Adult; Aged; Endothelial Cells; Female; Glycocalyx; Heparitin Sulfate; Humans; Intercellular Adhesion Molecule-1; Interleukin-6; Male; Middle Aged; Sepsis; Syndecan-1; Vascular Cell Adhesion Molecule-1