Lark M W, Wight T N
Arteriosclerosis. 1986 Nov-Dec;6(6):638-50. doi: 10.1161/01.atv.6.6.638.
The nature of the extracellular matrix may influence the types and amounts of proteoglycans synthesized by arterial smooth muscle cells. To determine if collagen modulates proteoglycan metabolism by these cells, arterial smooth muscle cells derived from aortic explants of the pigtail monkey (Macaca nemestrina) were cultured on both tissue culture plastic and hydrated type I collagen gels for 7 days. Cells were radiolabeled with Na2[35S]O4 during the final 48 hours of the growth period, and proteoglycans were extracted from the culture medium and cell layer by using 4 M guanidine hydrochloride in the presence of protease inhibitors. Cultures on collagen accumulated approximately 40% less [35S]O4 = -radiolabeled proteoglycan when compared to cultures on plastic. However, cells on collagen accumulated approximately 50% of their newly synthesized proteoglycan within the cell layer, while cells on plastic deposited less than 20% of their total radiolabeled proteoglycan in this culture compartment. Pulse-chase analysis indicated that the elevated accumulation of proteoglycan observed in the cell layer of collagen cultures was due, at least in part, to inhibition of turnover of these molecules. Cells on both collagen and plastic synthesized a large chondroitin sulfate proteoglycan which was secreted into the medium and deposited within the cell layer. On the other hand, cells on collagen synthesized a smaller iduronic acid-rich dermatan sulfate proteoglycan which was deposited only within the collagen gel and not secreted into the medium. By comparison, cells grown on plastic synthesized both a small glucuronic acid-rich dermatan sulfate proteoglycan which was secreted into the medium as well as an iduronic acid-rich dermatan sulfate which was present in the cell layer. Unlike the cell layer-derived dermatan sulfate proteoglycan isolated from the collagen gels, the majority of the cell layer-derived dermatan sulfate from cells on plastic was insensitive to papain treatment and thus identified as free glycosaminoglycan chains. Analysis of the total radiolabeled proteoglycans isolated under the two culture conditions indicated that cultures grown on collagen accumulate less chondroitin sulfate proteoglycan and heparan sulfate proteoglycan but over twice as much iduronic acid-rich dermatan sulfate proteoglycan. This culture system is offered as a model to determine the mechanisms by which collagen may in part regulate the metabolism of proteoglycans by arterial smooth muscle cells.
细胞外基质的性质可能会影响动脉平滑肌细胞合成的蛋白聚糖的类型和数量。为了确定胶原蛋白是否调节这些细胞的蛋白聚糖代谢，将来自猪尾猴（食蟹猴）主动脉外植体的动脉平滑肌细胞在组织培养塑料和水合I型胶原蛋白凝胶上培养7天。在生长周期的最后48小时用Na2[35S]O4对细胞进行放射性标记，并在蛋白酶抑制剂存在的情况下，用4M盐酸胍从培养基和细胞层中提取蛋白聚糖。与在塑料上培养的细胞相比，在胶原蛋白上培养的细胞积累的[35S]O4=-放射性标记的蛋白聚糖减少了约40%。然而，在胶原蛋白上的细胞在细胞层内积累了约50%的新合成蛋白聚糖，而在塑料上的细胞在该培养隔室中沉积的总放射性标记蛋白聚糖不到20%。脉冲追踪分析表明，在胶原蛋白培养物的细胞层中观察到的蛋白聚糖积累增加至少部分是由于这些分子的周转受到抑制。在胶原蛋白和塑料上的细胞都合成了一种大的硫酸软骨素蛋白聚糖，该蛋白聚糖被分泌到培养基中并沉积在细胞层内。另一方面，在胶原蛋白上的细胞合成了一种较小的富含艾杜糖醛酸的硫酸皮肤素蛋白聚糖，该蛋白聚糖仅沉积在胶原蛋白凝胶内，而不分泌到培养基中。相比之下，在塑料上生长的细胞合成了一种小的富含葡萄糖醛酸的硫酸皮肤素蛋白聚糖，该蛋白聚糖被分泌到培养基中，以及一种存在于细胞层中的富含艾杜糖醛酸的硫酸皮肤素。与从胶原蛋白凝胶中分离的细胞层衍生的硫酸皮肤素蛋白聚糖不同，从塑料上的细胞中分离的细胞层衍生的硫酸皮肤素的大部分对木瓜蛋白酶处理不敏感，因此被鉴定为游离糖胺聚糖链。对在两种培养条件下分离的总放射性标记蛋白聚糖的分析表明，在胶原蛋白上生长的培养物积累的硫酸软骨素蛋白聚糖和硫酸乙酰肝素蛋白聚糖较少，但富含艾杜糖醛酸的硫酸皮肤素蛋白聚糖是其两倍多。该培养系统作为一个模型，用于确定胶原蛋白可能部分调节动脉平滑肌细胞蛋白聚糖代谢的机制。
