[S]Proteoglycan metabolism of arterial smooth muscle cells cultured from normotensive and hypertensive rats (original) (raw)
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Hypertension, 1999
Extracellular matrix (ECM) modifications in the vascular wall contribute to the narrowing of arteries in hypertension. Because direct evidence for the role of proteoglycans (PGs) in the pathological process of resistance-sized arteries has not already been demonstrated, we examined the effect of growth factors on secreted and membrane-bound PG synthesis by cultured mesenteric vascular smooth muscle cells (VSMC) from spontaneously hypertensive rats (SHR) and Wistar rats. After 48 hours of stimulation with angiotensin II (Ang II), platelet-derived growth factor (PDGF-BB), and 10% fetal calf serum (FCS) or 0.1% FCS as control, PG synthesis (in dpm/ng DNA) was evaluated in the medium (M-ECM) and in the cell layer (P-ECM) by a double-isotopic label method with both [ 3 H]-glucosamine and [ 35 S]-sodium sulfate, which are incorporated into all complex carbohydrates or only into sulfated disaccharides, respectively. VSMC from SHR displayed a significantly lower level of synthesis of M-ECM [ 3 H]-PGs than those of Wistar rats in all the experimental groups, including the control group (0.1% FCS), but no differences in M-ECM [ 35 S] uptake were found in any case. In the P-ECM, Ang II was the only factor that produced a lesser effect on [ 3 H]-glucosamine and a greater effect on [ 35 S]-sodium sulfate uptakes in VSMC from SHR than from Wistar rats. The most prominent change seen in VSMC from SHR was an increased sulfation, assessed by [ 35 S]/[ 3 H] ratio, in nonstimulated cells and in response to 10% FCS and Ang II but not to PDGF-BB compared with VSMC from Wistar rats. These data indicate the existence of changes in PG modulation in the resistance vessels of SHR, which suggests that PGs may contribute to the development of structural and functional modifications in hypertensive states. (Hypertension. 1999;34[part 2]:893-896.
The Journal of cell biology, 1983
Near confluent monolayers of arterial smooth muscle cells derived from Macaca nemestrina were labeled with Na2[35S]O4 and the newly synthesized proteoglycans present in the culture medium and cell layer were extracted with either 4 M guanidine HCl (dissociative solvent) or 0.5 M guanidine HCl (associative solvent) in the presence of protease inhibitors. The proteoglycans in both compartments were further purified by cesium chloride density gradient ultracentrifugation. Two size classes of proteoglycans were observed in the medium as determined by chromatography on Sepharose CL-2B. The large population (Kav = 0.31) contained predominantly chondroitin sulfate chains with Mr = approximately 40,000. The smaller population (Kav = 0.61) contained dermatan sulfate chains of similar Mr (approximately 40,000). When tested for their ability to aggregate, only proteoglycans in the large-sized population were able to aggregate. A chondroitin sulfate containing proteoglycan with identical proper...
Atherosclerosis, 1987
, or in EC-conditioned medium, show increased synthesis of glycosaminoglycans (GAG). We have found that both the amount and type of GAG produced by the SMC are dependent on the density of the EC. EC (porcine) at a low density (0.1-0.5 x IO6 cells/25 cm2), or their conditioned media, where the most active per cell in stimulating GAG. All GAG were stimulated but the increase was due mostly to hyaluronic acid (HA). At intermediate densities (1.0 X 106/25 cm2) stimulation was markedly reduced, but still present, and both HA and sulphated GAG were similarly increased. At high densities (1.5-3 x 106/25 cm2) where EC were confluent there was very little stimulation of HA but continued stimulation of sulphated GAG synthesis. The shift in stimulation from HA to sulphated GAG with increasing density was most clearly demonstrated by the decrease in the HA to the chondroitin sulphate ratio. These findings provide support for the general concept that SMC metabolism may be affected by changes in the state of the endothelium.
Proteoglycans production by aortic vascular smooth muscle cells from hypertensive rats
2000
Remodeling of large and small arteries contributes to the development and complications of hypertension. Artery structural changes in chronic sustained hypertension include vascular smooth muscle cells (VSMC) proliferation and extracellular matrix (ECM) modifications. Extracellular constituents such as proteoglycans (PGs), may modulate vascular stiffness and VSMC growth and differentiation. We examined the effect of growth factors on secreted and membrane-bound PGs
American Journal of Hypertension, 2002
Several functional and structural modifications at the vascular level have been described in spontaneously hypertensive rats (SHR) during the early development of hypertension. In this study, we hypothesize that changes in the extracellular matrix (ECM) could precede the development of hypertension. Synthesis of secreted and membrane-bound sulfated proteoglycans (S-PG) by cultured vascular smooth muscle cells (VSMC) obtained from young spontaneously hypertensive rats (pSHR) mesenteric resistance arteries, during the period preceding the elevation of blood pressure (BP) was tested. After 24 h of stimulation with angiotensin II (Ang II), 10% fetal calf serum (FCS), or 0.1% FCS as control, medium and cell layer S-PG synthesis was evaluated by labeling sulfated disaccharides with [ 35 S]sodium sulfate. To relate this variable with cell proliferation, DNA synthesis was measured by incorporation of [ 3 H]thymidine in the cell lysate. The VSMC from pSHR synthesized more secreted and mem-brane-bound S-PG than age-matched Wistar rat (pW) cells in the nonstimulated (0.1% FCS) and stimulated (Ang II or 10% FCS) experimental groups. When data were expressed as percent of their own control value, both Ang II and 10% FCS lowered basal secreted and cell-associated S-PG content in VSMC from pSHR, whereas in pW rat cells, these agents produced a small increase or no change. An inverse relationship between proliferation and total S-PG production (secreted plus membrane-bound) was found in pSHR cells, but not in pW cells. In conclusion, the present study demonstrates that changes in S-PG synthesis by VSMC of resistance arteries precede the vascular dysfunction associated with the development of hypertension in SHR. Am J Hypertens 2002;15:416 -421
Glycosaminoglycan content in neonatal rat aortic smooth muscle cell cultures
Atherosclerosis, 1988
In the present study the biosynthesis of glycosaminoglycans (GAGS) by neonatal rat aortic smooth muscle cells in culture was studied. Heparan sulfate (HS) was the predominant GAG of the cell layer accounting for 32-49% of the total GAGS depending on the time in culture. The presence of low sulfated chondroitin sulfate (LSC) in aortic smooth muscle cell cultures is reported here for the first time. The effect of ascorbate on the synthesis and accumulation of these macromolecules resulted in a relative increase of C4S and DS in the cell layer. In contrast, the distribution of the GAGS which were secreted into the medium was not significantly effected by the addition of ascorbate. While HS was always found to be a minor component, the other GAGS were present in about equal concentrations. The total GAG accumulation in the medium was much greater (91-97s) than that of the cell layer (3-9s) indicating that the cells are synthesizing relatively large amounts of GAGS, although incorporation of these macromolecules into the extracellular matrix was consistently low.
Biochimica Et Biophysica Acta-general Subjects, 1988
The synthesis of proteoglycans by aorta explants from rabbits with diet-induced atherosclerosis and controls was studied by 3SS-incorporation. Proteoglycans were isolated under dissociative conditions from incubation medium and from arterial explants. Additionally, the tissue proteoglycans that were not extracted by 4 M guanidine-HCI were solubilized by digestion of the tissue by elastase in the presence of proteinase inhibitors. The residual tissue was hydrolyzed by papain and glycosaminoglycans were isolated. The atherosclerotic aorta tissue incorporated twice the amount of 3Ss into proteoglycans than observed for controls; in both groups about 70% of the label incorporated into the tissue was noted in the proteoglycans extracted by guanidine-HCI, while about 30% of the total 35S-labeled proteoglycans synthesized by the explants were found in the media. Atherosclerotic tissue incorporated 3SS predominantly into chondroitin sulfate proteoglycans when compared to control tissue. The chondroitinase ABC-digestable proteoglycans that were extracted by guanidine-HCI from atherosclerotic tissues were of larger molecular size than those from control tissue, but the core proteins from these preparations were similar. The heparan sulfate proteoglycan that was obtained by dissociative extraction from atherosclerotic tissue had greater amounts of N-acetyl and lesser amounts of N-sulfate ester groups than the preparation from control tissue. Digestion of the tissue by elastase yielded heparan sulfate proteoglycan as the major constituent in both groups, although atherosclerotic tissue contained relatively small amounts of this proteoglycan. The residual tissue from both groups contained chondroitin sulfate and heparan sulfate as the major glycosaminoglycans with the latter showing a decrease with atherosderosis. Atherosclerotic tissue secreted into the medium about two-fold more 35Slabeled proteoglycans with larger molecular size than control tissue; proteoglycans of the heparan sulfate and chondroitin sulfate types were the major constituents in the culture medium of both tissues. Thus, proteoglycans undergo both quantitative and qualitative changes in atherosclerosis, reflecting the enhanced smooth muscle cell activity. These changes are potentially important in modulating lipoprotein binding and hemostatic properties, as well as fibrillogenesis of the arterial wall.
Sulfated glycosaminoglycans synthesized by human smooth muscle cells isolated from different organs
Biochimica et Biophysica Acta (BBA) - General Subjects, 1986
The sulfated glycosaminoglycans synthesized by human smooth muscle cells isolated from different organs were identified on the basis of electrophoretic mobility, enzymatic degradation with specific mucopolysaccharidases and by the type of degradation products formed. The results obtained indicated that chondroitin sulfate and beparan sulfate were the main glycosaminoglycans found, that most of the labeled glycosaminoglycans were found in the pericellular pool, and that no marked differences were observed in the sulfated glycosaminoglycan composition of the smooth muscle cells obtained from different organs. 'Liver connective tissue cells', isolated from pathological livers (which had been shown to possess biochemical and physiological features typical of smooth muscle cells) showed a pattern of glycosaminoglycan synthesis similar to that of the smooth muscle cells.
Cell Differentiation, 1983
Monolayer cultures of fibroblast, smooth muscle and endothelium-like cells incorporated 35SO~ into glycosaminoglycans of the extracellular, pericellular and intracellular compartments. These glycosaminoglycans have been identified on the basis of electrophoretic mobility, enzymatic degradation with specific mucopolysaccharidases and by the type of degradation products formed. The sulfated glycosaminoglycans from the extracellular pool of the three cell types show a similar composition, while the intracellular and pericellular pools of the three cells have a different glycosaminoglycans composition. They differ in the relative proportion of heparitin sulfate and chondroitin sulfate and in the structure of isomeric chondroitin sulfate. glycosaminoglycans fibroblasts smooth muscle endothelium-like cells monolayer cultures