Proteoglycans in primate arteries. III. Characterization of the proteoglycans synthesized by arterial smooth muscle cells in culture.
Proteoglycans in primate arteries. III. Characterization of the proteoglycans synthesized by arterial smooth muscle cells in culture.
复制标题
灵长类动脉中的蛋白聚糖。 iii。在培养中由动脉平滑肌细胞合成的蛋白聚糖的表征。
DOI：
10.1083/jcb.96.1.167
复制
发表时间：
1983-01
期刊：
影响因子：
--
通讯作者：
Hascall VC
中科院分区：
文献类型：
--
作者：
Wight TN;Hascall VC
用Na 2 [35 S] O 4标记来自Macaca nemestrina的动脉平滑肌细胞的近汇合单层，并在蛋白酶抑制剂存在下用4 M盐酸胍（解离溶剂）或0.5 M盐酸胍（缔合溶剂）提取培养基和细胞层中存在的新合成的蛋白聚糖。通过氯化铯密度梯度超离心进一步纯化两个隔室中的蛋白聚糖。通过Sepharose CL-2B色谱法测定，在培养基中观察到两种大小的蛋白聚糖。大群体（Kav = 0.31）主要含有硫酸软骨素链，Mr =约40，000。较小的群体（Kav = 0.61）含有相似Mr（约40，000）的硫酸皮肤素链。当测试它们的聚集能力时，只有大规模群体中的蛋白聚糖能够聚集。从细胞层中分离出具有相同性质的含有蛋白聚糖的硫酸软骨素。此外，细胞层含有硫酸皮肤素成分，其在Sepharose CL-2B上的洗脱时间比培养基中存在的硫酸皮肤素蛋白聚糖晚（Kav = 0.78）。纯化的蛋白聚糖的电子显微镜显示了瓶刷结构，其包含平均140 nm长的中心核，平均具有8至10个侧突起。侧突起的长度变化，但平均在70和75 nm之间。用番红0染色后，在平滑肌细胞培养物的细胞间基质中观察到类似的瓶刷结构。该培养系统提供了一个模型，以调查参与动脉蛋白多糖的合成和降解的调节参数。
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 properties was isolated from the cell layer. In addition, the cell layer contained a dermatan sulfate component which eluted later on Sepharose CL-2B (Kav = 0.78) than the dermatan sulfate proteoglycan present in the medium. Electron microscopy of the purified proteoglycans revealed a bottlebrush structure containing a central core averaging 140 nm in length with an average of 8 to 10 side projections. The length of the side projections varied but averaged between 70 and 75 nm. Similar bottlebrush structures were observed in the intercellular matrix of the smooth muscle cell cultures after staining with Safranin 0. This culture system provides a model to investigate parameters involved in the regulation of synthesis and degradation of arterial proteoglycans.