Extracellular matrix microfibrils are composed of core proteins coated with fibronectin.
Extracellular matrix microfibrils are composed of core proteins coated with fibronectin.
复制标题
细胞外基质微纤维由纤连蛋白包被的核心蛋白组成。
DOI：
--
复制
发表时间：
1985
影响因子：
3.2
通讯作者：
O. Blumenfeld
中科院分区：
文献类型：
--
作者：
E. Schwartz;S. Goldfischer;B. Coltoff;O. Blumenfeld
培养的小牛主动脉平滑肌细胞的细胞外蛋白主要由直径为10-20 nm的微纤维组成，这是许多组织中描述的“弹性蛋白相关”微纤维的典型特征。化学和免疫化学证据表明，微纤维至少包括两种蛋白质：核心蛋白和纤连蛋白。微纤维的不溶性蛋白质以颗粒的形式获得，并且在兔中产生针对这些组分的抗体。抗血清与不溶性微纤维蛋白和可溶性纤维连接蛋白在酶联免疫吸附试验中反应，并免疫染色培养细胞中的细胞外微纤维。制备免疫球蛋白（IG）级分并用纤连蛋白吸收。被吸收的IgG保留其与微纤维蛋白的反应性，但不再与可溶性纤连蛋白反应。使用吸附的IgG和IgG对可溶性纤连蛋白进行免疫荧光研究。两种抗体均在对数期细胞的细胞外基质中显示免疫反应性微纤维。然而，随着培养时间的增加，随着细胞达到汇合，与吸收的IgG反应的微纤维的免疫荧光变得不那么强烈，而与IgG反应的微纤维纤连蛋白增加;在汇合的细胞中，基本上没有染色被检测到与吸收的IgG，和密集的网络强烈染色的微纤维被认为与IgG纤连蛋白。用尿素处理这些培养物导致纤连蛋白的部分解离，并增加了具有吸收的IgG的微纤维的可视化;双标记免疫荧光显示这两种蛋白质发生在相同的微纤维上。通过免疫电镜证实了免疫反应位点定位于细胞外微纤维。几乎定量裂解与溴化氰未能解离的纤连蛋白的抗原活性片段的核心蛋白的微纤维的溴化氰片段。得出的结论是，微纤维含有核心蛋白和纤连蛋白，它们共同分布在不溶性的，可能是共价交联的聚集体中。核心蛋白首先由细胞沉积，并且作为培养时间的函数，纤连蛋白逐渐覆盖其表面。
Extracellular proteins of cultured calf aortic smooth muscle cells consist predominantly of microfibrils 10-20 nm in diameter typical of "elastin-associated" microfibrils described in many tissues. Chemical and immunochemical evidence is presented that microfibrils consist of at least two proteins: core protein and fibronectin. Insoluble proteins of the microfibrils were obtained in the form of a pellet and antibodies raised in rabbits against these components. The antisera reacted with the insoluble microfibrillar proteins and with soluble fibronectin in enzyme-linked immunosorbent assay, and immunostained the extracellular microfibrils in cultured cells. An immunoglobulin (Ig) fraction was prepared and absorbed with fibronectin. The absorbed IgG retained its reactivity with the microfibrillar proteins but was no longer reactive with soluble fibronectin. Immunofluorescence studies were carried out using the absorbed IgG and IgG to soluble fibronectin. Both antibodies showed immunoreactive microfibrils in the extracellular matrix of cells in log phase. However, with increasing time in culture, as the cells reached confluence, the immunofluorescence of microfibrils reacting with the absorbed IgG became less intense, whereas that of microfibrils reacting with IgG to fibronectin increased; in confluent cells, essentially no staining was detected with the absorbed IgG, and a dense network of intensely stained microfibrils was seen with IgG to fibronectin. Treatment of these cultures with urea led to partial dissociation of the fibronectin and increased visualization of the microfibrils with the absorbed IgG; double-label immunofluorescence showed that both proteins occurred on the same microfibrils. The localization of immunoreactive sites to the extracellular microfibrils was confirmed by immunoelectron microscopy. Nearly quantitative cleavage with CNBr failed to dissociate the antigenically active fragments of fibronectin from the CNBr fragments of the core proteins of the microfibrils. It was concluded that microfibrils contain core proteins and fibronectin that are codistributed in insoluble, possibly covalently cross-linked, aggregates. The core proteins are first deposited by the cell and, as a function of time in culture, fibronectin gradually coats their surface.