Integrin binding and cell spreading on extracellular matrix act at different points in the cell cycle to promote hepatocyte growth.
Integrin binding and cell spreading on extracellular matrix act at different points in the cell cycle to promote hepatocyte growth.
复制标题
整合素结合和细胞外基质上的细胞扩散在细胞周期的不同点发挥作用，促进肝细胞生长。
DOI：
10.1091/mbc.5.9.967
复制
发表时间：
1994
影响因子：
3.3
通讯作者：
Ingber,DE
中科院分区：
文献类型：
--
作者：
Hansen,LK;Mooney,DJ;Vacanti,JP;Ingber,DE
本研究旨在确定整合素结合和细胞形状变化在细胞外基质（ECM）控制细胞周期进程中的重要性。原代大鼠肝细胞在ecm包被的培养皿中培养于无血清培养基中，培养基中含有饱和量的生长因子（表皮生长因子和胰岛素）。用纤维连接蛋白（FN）包被培养皿的方法可以促进细胞的扩散和整合素的结合。通过在包被人工合成的arg-gly-asp （RGD）肽的培养皿中培养细胞，将整合素结合从细胞形状变化中分离出来，RGD -肽作为整合素配体，但不支持肝细胞扩展。检测早期（junB）和晚期（ras）生长反应基因的表达和DNA合成，以确定这些基质是否诱导g0同步肝细胞重新进入生长周期。在FN上镀的细胞表现出junB和ras基因表达的短暂增加（分别在镀后2和8 h内），并同步进入S期。在rgd包被的培养皿中，观察到junB和ras在相似的时间过程中被诱导，然而，这些圆形细胞没有进入S期。RGD上的圆形细胞在G1中后期被阻断的可能性得到了证实，当胰蛋白酶化并在培养30小时后复制到FN包被的培养皿上时，它们需要相似的时间（12-15小时）才能重新进入S期，因为已经扩散并允许通过FN上的G1进展。我们之前的研究表明，在这些rgd包被的培养皿中培养的肝细胞仍然有活力，并保持高水平的肝脏特异性功能。因此，这些结果表明，ECM在细胞周期的两个不同点起作用来调节肝细胞的生长：第一，通过整合素结合激活G1/ G1过渡；第二，通过与细胞扩散相关的机制促进G1/S过渡并关闭默认分化程序。
This study was undertaken to determine the importance of integrin binding and cell shape changes in the control of cell-cycle progression by extracellular matrix (ECM). Primary rat hepatocytes were cultured on ECM-coated dishes in serum-free medium with saturating amounts of growth factors (epidermal growth factor and insulin). Integrin binding and cell spreading were promoted in parallel by plating cells on dishes coated with fibronectin (FN). Integrin binding was separated from cell shape changes by culturing cells on dishes coated with a synthetic arg-gly-asp (RGD)-peptide that acts as an integrin ligand but does not support hepatocyte extension. Expression of early (junB) and late (ras) growth response genes and DNA synthesis were measured to determine whether these substrata induce G0-synchronized hepatocytes to reenter the growth cycle. Cells plated on FN exhibited transient increases in junB and ras gene expression (within 2 and 8 h after plating, respectively) and synchronous entry into S phase. Induction of junB and ras was observed over a similar time course in cells on RGD-coated dishes, however, these round cells did not enter S phase. The possibility that round cells on RGD were blocked in mid to late G1 was confirmed by the finding that when trypsinized and replated onto FN-coated dishes after 30 h of culture, they required a similar time (12-15 h) to reenter S phase as cells that had been spread and allowed to progress through G1 on FN. We have previously shown that hepatocytes remain viable and maintain high levels of liver-specific functions when cultured on these RGD-coated dishes. Thus, these results suggest that ECM acts at two different points in the cell cycle to regulate hepatocyte growth: first, by activating the G0/G1 transition via integrin binding and second, by promoting the G1/S phase transition and switching off the default differentiation program through mechanisms related to cell spreading.