heparitin-sulfate and Hyperplasia

Thrombogenicity and neointimal hyperplasia are major causes for synthetic vascular graft failure. Bioactive coatings like heparin have improved patency by reducing thrombogenicity, but neointimal hyperplasia still remains an unsolved problem. Surface coatings with heparan sulfate (HS), the major component of the glycocalyx of endothelial cells, have shown reduced platelet and cell adhesion in vitro. The aim of the study was to evaluate the in vivo surface properties of expanded ePTFE vascular grafts with a semisynthetic HS-like coating (SSHS).. ePTFE vascular grafts (n = 16, diameter 3.5 mm) covalently coated with SSHS were compared with uncoated grafts (n = 16) of the same diameter in a carotid interposition model in 16 sheep. The grafts were harvested at 20 wk for histological and morphometric analysis.. SSHS-coated grafts showed less neointima formation than uncoated grafts (P < 0.001). There was no evidence for cell or protein adhesion to SSHS-coated grafts, whereas the surface of uncoated ePTFE grafts was covered with a confluent circular layer of neointima. No difference was found concerning reactions at the anastomotic site of the genuine carotid vessel, both groups displayed neointimal hyperplasia.. ePTFE grafts covalently coated with a semisynthetic SSHS-glycosaminoglycan successfully mimicked the endothelial glycocalyx. They displayed excellent antiadhesive properties preventing neointimal formation on the graft surface. The results indicate that a biomimetic SSHS coating may be a useful component of bioengineered grafts and an alternative to synthetic surfaces and endothelial seeding.

Topics: Animals; Biomimetic Materials; Blood Vessel Prosthesis; Blood Vessel Prosthesis Implantation; Carotid Arteries; Cell Adhesion; Cell Proliferation; Coated Materials, Biocompatible; Endothelial Cells; Female; Graft Occlusion, Vascular; Heparitin Sulfate; Hyperplasia; Materials Testing; Models, Animal; Neointima; Polytetrafluoroethylene; Prosthesis Design; Sheep, Domestic; Time Factors

Coronary microvascular hyperplasia is a cause of microvessel angina, although the underlying cellular mechanisms remain unclear. We examined how mononuclear cells expressing β-actin (β-MNCs), which were identified in coronary vessels, induce coronary microvascular hyperplasia.The presence of β-MNCs in coronary hyperplastic arterial (HAM) and venous microvessels (HVM) was examined by endomyocardial biopsy in 25 patients with suspected microvessel angina. β-MNCs were identified in 14 HAMs obtained from 11 patients. Basic fibroblast growth factor and heparin sulfate were injected into the infarcted myocardium to induce HAM and HVM in 28 beagles, and then we examined the role of β-MNCs in the onset of HAM and HVM. The following changes were observed after infarction induction in beagles: (a) migration of β-MNCs from the existing microvessels into the interstitial space at 1-2 weeks; (b) those traversing the adventitia into the media, but not intima, of microvessels; (c) their transformation to smooth muscle cells (SMCs) and/or connective tissues (collagen and elastin fibers); (d) and medial hyperplasia without intimal hyperplasia. Medial hyperplasia was classified into SMC-proliferative and both SMC- and connective tissue-proliferative types. β-MNCs expressed CD(34) but did not express other major vessel-related cell markers.β-MNCs are a vascular progenitor, and migrate out of the adventitia into media, and participate in the etiology of coronary microvascular medial hyperplasia.

Topics: Actins; Animals; Cell Movement; Connective Tissue; Coronary Vessels; Dogs; Female; Fibroblast Growth Factor 2; Heparitin Sulfate; Humans; Hyperplasia; Male; Microvascular Angina; Microvessels; Middle Aged; Tunica Media

To clarify the difference between cutaneous responses to single and repeated barrier disruption, changes of epidermal gene expression were examined by using RT-PCR. In repeatedly barrier-disrupted skin, heparanase was specifically up-regulated in epidermis. In addition, there was a marked decrease in heparan sulfate (HS) chains of perlecan in basement membrane at the dermal-epidermal junction (DEJ) compared with singly disrupted skin. HS chains form a reservoir for heparan sulfate-binding growth factors. In repeatedly barrier-disrupted skin, expression of vascular endothelial growth factor-A (VEGF-A), an angiogenic factor, was induced in epidermis, whereas thrombospondin-1 (TSP-1), an angiogenesis inhibitor, was down-regulated, and concomitantly blood vessels were elongated and enlarged in dermis. Expression of VEGF-C, a lymphangiogenesis factor, was augmented in epidermis of repeatedly barrier-disrupted skin, concomitantly with an increase in the number and size of lymphatic vessels. Topical application of a synthetic heparanase inhibitor, 1-[4-(1H-benzoimidazol-2-yl)phenyl]-3-[4-(1H-benzoimidazol-2-yl)phenyl]urea, to skin after barrier disruption significantly suppressed wrinkle formation, degradation of HS chains in the basement membrane, epidermal hyperplasia and the changes of blood and lymphatic vessels. These results suggest that chronic barrier disruption activates heparanase and induces gene expression changes, leading to increased growth factor interaction between epidermis and dermis, and facilitating various cutaneous changes, including wrinkle formation.

Topics: Animals; Basement Membrane; Dermis; Enzyme Inhibitors; Epidermis; Fibroblast Growth Factor 2; Fibroblast Growth Factor 7; Gene Expression; Glucuronidase; Heparan Sulfate Proteoglycans; Heparitin Sulfate; Hyperplasia; Lymphangiogenesis; Male; Mice; Mice, Hairless; Neovascularization, Pathologic; Skin; Skin Aging; Thrombospondin 1; Vascular Endothelial Growth Factor A; Vascular Endothelial Growth Factor C

Smooth muscle cell (SMC) proliferation is a critical process in vascular disease. Heparan sulfate (HS) proteoglycans inhibit SMC growth, but the role of endogenous counterparts in the vessel wall in control of SMC function is not known in detail. Perlecan is the major HS proteoglycans in SMC basement membranes and in vessel wall extracellular matrix (ECM). In this study, transgenic mice with HS-deficient perlecan were analyzed with respect to vascular phenotype and intimal lesion formation. Furthermore, SMC cultures were established and characterized with respect to morphology, immunocytochemical features, proteoglycan synthesis, proliferative capacity, and ECM binding of basic fibroblast growth factor (FGF-2). In vitro, mutant SMCs formed basement membranes with perlecan core protein, but with decreased levels of HS, they showed diminished secretion of HS-containing perlecan into the medium and a defective ECM-binding capacity of FGF-2. In vitro, mutant SMCs showed increased proliferation compared with wild-type cells, and in vivo, enhanced SMC proliferation and intimal hyperplasia were observed after flow cessation of the carotid artery in mutant mice. The results indicate that the endogenous HS side-chains of perlecan contribute to SMC growth control both in vitro and during intimal hyperplasia, possibly by sequestering heparin-binding mitogens such as FGF-2.

Topics: Animals; Cell Division; Cells, Cultured; Crosses, Genetic; Exons; Fibroblast Growth Factor 2; Heparan Sulfate Proteoglycans; Heparitin Sulfate; Hyperplasia; Mice; Mice, Inbred C57BL; Mice, Transgenic; Muscle, Smooth, Vascular; Protein Binding; Sequence Deletion; Tunica Intima

Perlecan, a heparan sulfate proteoglycan, has been suggested to be critical for regulation of vascular repair. We generated clones of endothelial cells expressing an antisense vector targeting domain III of perlecan. Transfected cells produced significantly less perlecan than parent cells and showed a reduced ability to inhibit the binding and mitogenic activity of fibroblast growth factor-2 in vascular smooth muscle cells. Endothelial cells were seeded onto three-dimensional polymeric matrices and implanted adjacent to porcine carotid arteries subjected to deep injury. Although the parent endothelial cells prevented occlusive thrombosis, perlecan-deficient cells were completely ineffective. The ability of endothelial cells to inhibit intimal hyperplasia, however, was abrogated only in part by perlecan suppression. The differential regulation by perlecan of these different aspects of vascular repair may explain why control of clinical clot formation does not lead to full control of intimal hyperplasia. Thus the use of genetically modified tissue-engineered cells provides a new approach for dissecting the role of specific factors within the complex environment of the blood vessel wall.

Topics: Animals; Antisense Elements (Genetics); Cattle; Cell Division; Cells, Cultured; Endothelium, Vascular; Fibroblast Growth Factor 2; Heparan Sulfate Proteoglycans; Heparitin Sulfate; Humans; Hyperplasia; Male; Muscle, Smooth, Vascular; Proteoglycans; Swine; Thrombosis

Syndecan-1 binds basic fibroblast growth factor (bFGF), modulates neovascularization, plays a role in epithelial differentiation and is up-regulated by WT1. Malignant mesothelioma of the pleura is one of the most aggressive tumours known and expresses high levels of angiogenic growth factors. This study has analysed syndecan-1 expression in mesothelioma tumours and cell lines by immunohistochemistry and immunoblotting, using anti-syndecan-1 antibody directed against the core protein, and has examined its relation to morphology, bFGF, WT1, and intra-tumoural microvascular density (IMD). Shedding of syndecan-1 in the conditioned medium of mesothelioma cell lines was detected in variable amounts. These studies indicate that (1) there is no correlation of syndecan-1 with either bFGF expression or IMD in mesotheliomas in vivo; (2) syndecan-1 is strongly expressed in the epithelial type of mesothelioma and in the epithelial component of biphasic mesotheliomas and the expression is reduced or lost in sarcomatoid differentiation; together with the finding that (3) syndecan-1 correlates with WT1 immuno-expression, this suggests that syndecan-1 might relate to the differentiation state of mesothelial/mesothelioma cells; and (4) syndecan-1-positive tumours are associated with a longer survival (p = 0.02) than mesotheliomas with no or little syndecan-1 expression, on univariate analysis. These findings therefore indicate that syndecan-1 can be an important prognostic indicator in mesotheliomas and its loss may be important in the epithelial-mesenchymal transformation of mesothelioma cells.

Topics: Adenocarcinoma; Biomarkers, Tumor; Blotting, Western; Cell Differentiation; Chi-Square Distribution; DNA-Binding Proteins; Epithelium; Fibroblast Growth Factor 2; Heparan Sulfate Proteoglycans; Heparitin Sulfate; Humans; Hyperplasia; Immunohistochemistry; Membrane Glycoproteins; Mesothelioma; Neovascularization, Pathologic; Precipitin Tests; Prognosis; Proteoglycans; Survival Rate; Syndecan-1; Syndecans; Transcription Factors; WT1 Proteins

In the present work the activities of GGT and G-6-Pase and the content of Cyt P-450 were determined in surgically removed liver specimens (16 hepatocellular carcinomas, 8 focal nodular hyperplasias and 4 adenomas). The activities were compared to the surrounding seemingly normal liver tissue. In the adenomas neither of the enzymes studied showed alterations, characteristic for hepatocarcinogenesis. Four out of 8 FNHs had the enzyme pattern that was found in experimental liver carcinogenesis. Liver carcinoma specimens proved to be heterogenous. Neither elevated GGT nor reduced G-6-Pase activity was consistent in these samples although the average of G-6-Pase activity decreased to 50 percent. Cytochrome P-450 was significantly reduced in the majority of cases, showing the best agreement with the tendency observed in experimental models. As an other approach, the qualitative and quantitative alterations of proteoglycans (PG) were analized in the same tumor samples. The amount of sugar components of PGs the glycosaminoglycans (GAG) increased by many times in liver tumors. Carcinoma samples were characterized by about twentyfold increase in chondroitin sulfate content, compared to normal liver. The enhancement of GAGs is partly the consequence of a selective alteration in PG expression. The amount of perlecan and decorin was found to be increased, while syndecan disappeared from liver carcinomas. These data suggest that malignant transformation in liver is accompanied by specific alteration in the content, composition and structure of PGs. Presumably, these changes have significance in tumor progression and have also the potential to be used as markers for liver tumors.

Topics: Adenoma; Biomarkers, Tumor; Carcinoma, Hepatocellular; Clinical Enzyme Tests; Cytochrome P-450 Enzyme System; Diagnosis, Differential; gamma-Glutamyltransferase; Glucose-6-Phosphatase; Glycosaminoglycans; Heparan Sulfate Proteoglycans; Heparitin Sulfate; Humans; Hyperplasia; Liver; Liver Neoplasms; Membrane Glycoproteins; Proteoglycans; Syndecans

The efficacies of standard heparin (SH), low molecular weight heparin (LMWH), and a mixture of sulfated glycosaminoglycans (Org 10172) were investigated with respect to their inhibitory effects on intimal thickening after endothelial injury in the common carotid artery of the rat. The injury was induced by air infusion into an isolated segment of the artery; the pharmacologic agents were administered by continuous intravenous infusion. After 2 weeks the animals were killed and the arteries examined. The control animals developed a marked intimal thickening. A dose-dependent decreases in the intima to media area (I-M) ratio was seen after SH, with approximately 50% and 90% inhibition of intimal thickening at doses of 5 and 50 USP U/kg/hr, respectively. At these effective doses, the effect of SH was associated with reendothelialization of the injured area. The effects of LMWH and Org 10172 were similar to that of SH at doses of 50 anti-Xa U/kg/hr, but these agents had only about 40% inhibition at doses of 15 anti-Xa U/kg/hr. The activated partial thromboplastin times were slightly prolonged in the animals treated with 50 USP/anti-Xa U/kg/hr of SH, LMWH, and Org 10172, whereas significant anti-Xa levels were observed at doses higher than 15 USP/anti-Xa U/kg/hr. It is concluded that SH, LMWH, and Org 10172 have significant inhibitory effects of intimal thickening after injury even at nonanticoagulant levels, with SH being the most potent.

Topics: Animals; Blood Coagulation; Carotid Arteries; Cell Division; Chondroitin Sulfates; Dermatan Sulfate; Dose-Response Relationship, Drug; Endothelium, Vascular; Glycosaminoglycans; Heparin; Heparin, Low-Molecular-Weight; Heparinoids; Heparitin Sulfate; Hyperplasia; Male; Rats; Rats, Inbred Strains