DIFFERENT BETA-1-INTEGRIN COLLAGEN RECEPTORS ON RAT HEPATOCYTES AND CARDIAC FIBROBLASTS
DIFFERENT BETA-1-INTEGRIN COLLAGEN RECEPTORS ON RAT HEPATOCYTES AND CARDIAC FIBROBLASTS
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DOI：
10.1016/0014-4827(90)90194-f
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发表时间：
1990-10-01
影响因子：
3.7
通讯作者：
RUBIN, K
中科院分区：
文献类型：
--
作者：
GULLBERG, D;TURNER, DC;RUBIN, K
在1 mM MnCl 2存在下，将原代大鼠肝细胞和新生儿心脏成纤维细胞的洗涤剂提取物应用于I型胶原-琼脂糖凝胶。用10 mM EDTA洗脱结合的蛋白质，在非还原条件下从肝细胞中产生一种β 1-整联蛋白异二聚体，其Mr为180，000/115，000。可以从表面碘化的成纤维细胞中分离出Mr（非还原）为180，000/115，000和145，000/115，000的两种β 1-整联蛋白。针对人整联蛋白VLA-1的大鼠同源物的单克隆抗体3A 3沉淀亲和纯化的Mr 180，000/115，000异二聚体，建立Mr 180，000亚基与β 1-整联蛋白亚家族的α 1-链的相关性。α 1 β 1-整联蛋白和145，000/β 1-整联蛋白异二聚体都特异性地结合到用缺乏RGD序列的胶原片段α 1（I）CB 3衍生的琼脂糖珠上。使用3A 3单克隆抗体的免疫荧光染色显示，大鼠α 1 β 1-整联蛋白存在于在天然I型胶原上生长的成纤维细胞的粘着斑部位，但不存在于纤连蛋白包被的基质上，尽管两种类型的基质都支持含有β 1-整联蛋白的粘着斑的形成。类似地，在包被有I型胶原蛋白（但不是纤连蛋白）的基质上培养的肝细胞被3A 3 IgG染色成局限于细胞外周的斑片状图案。此外，3A 3 IgG完全抑制肝细胞与I型胶原的附着，而在相同条件下，成纤维细胞与这些基质的附着仅被抑制约40%。肝细胞和心脏成纤维细胞与纤连蛋白的附着不受3A 3抗体存在的影响。总的来说，这些数据表明，大鼠同源物的人VLA-1异源二聚体生化和功能上满足标准的单一胶原受体对大鼠肝细胞。相反，大鼠心脏成纤维细胞利用两种不同的胶原结合整合素粘附胶原，其中之一是人VLA-1异二聚体的大鼠同源物。此外，α 1（I）CB 3含有β 1-整联蛋白的细胞结合位点。
Detergent extracts of primary rat hepatocytes and neonatal cardiac fibroblasts were applied to collagen type I-Sepharose in the presence of 1 mM MnCl2. Elution of bound proteins by 10 mM EDTA yielded one .beta.1-integrin heterodimer from hepatocytes with an Mr of 180,000/115,000 under nonreducing conditions. Two .beta.1-integrins with Mr''s (nonreduced) of 180,000/115,000 and 145,000/115,000 could be isolated from surface-iodinated fibroblasts. A monoclonal antibody, 3A3, directed against the rat homolog of the human integrin VLA-1, precipitated the affinity-purified Mr 180,000/115,000 heterodimer, establishing the relatedness of the Mr 180,000 subunit to the .alpha.1-chain of the .beta.1-integrin subfamily. Both the .alpha.1.beta.1-integrin and the 145,000/.beta.1-integrin heterodimers bound specifically to Sepharose beads derivatized with the collagen fragment .alpha.1(I) CB3, which lacks RGD sequences. Immunofluorescence staining using the 3A3 monoclonal antibody revealed that the rat .alpha.1.beta.1-integrin was present at focal adhesion sites of fibroblasts grown on native collagen type I- but not on fibronectin-coated substrates, although both types of substrates supported the formation of .beta.1-integrin containing focal adhesions. Similarly, hepatocytes cultured on substrata coated with collagen type I (but not fibronectin) were stained in a patchy pattern localized to the cell periphery by 3A3 IgG. Furthermore, 3A3 IgG completely inhibited the attachment of hepatocytes to collagen type I, whereas under identical conditions the attachment of fibroblasts to these substrates was inhibited only by approximately 40%. The attachment of both hepatocytes and cardiac fibroblasts to fibronectin was unaffected by the presence of the 3A3 antibody. Collectively these data show that a rat homolog of the human VLA-1 heterodimer both biochemically and functionally fulfills the criteria of a single collagen receptor on rat hepatocytes. In contrast, rat cardiac fibroblasts utilize two different collagen-binding integrins to adhere to collagen, one of which is the rat homolog of the human VLA-1 heterodimer. Furthermore .alpha.1(I) CB3 contains cell binding sites for .beta.1-integrins.