Reorganization of extracellular matrix by hepatic stellate cells (Ito cells) : Possible amelioration of liver fibrosis

肝星状细胞（HSC）在细胞外基质（ECM）成分的作用下表现出明显的形态学变化，并通过胶原凝胶培养，使HSC发生了许多细胞突起的伸长。本研究的结果表明，细胞表面整合素与ECM成分结合，随后是各种细胞内信号传导，最后是细胞骨架，特别是微管组装，诱导了突起的延伸。正如Senoo等人先前报道的那样，HSC不仅在形态上发生改变，而且在增殖和胶原合成/分泌方面也发生改变。明胶酶谱分析结果进一步证实，HSC在Ⅰ型胶原凝胶上培养的条件培养液中含有活化的基质金属蛋白酶（MMP）-2，而在聚苯乙烯和Matrigel上培养的条件培养液中没有MMP-2的表达，而在Ⅰ型胶原凝胶上培养的条件培养液中也有TIMP-2的表达，如通过逆转录酶-聚合酶链反应（RT-PCR）检测的，表明MMP-2的活化形式受到抑制。荧光免疫染色和原位酶谱进一步表明MMP- 1蛋白的存在和它们在I型胶原凝胶上的培养物中的活性。两者合计，这些结果表明，在转录水平或在翻译/翻译后阶段的ECM成分，如基底膜含有IV型胶原蛋白或间质基质成分，包括I型胶原蛋白的重组的复杂的调节机制。因此，使用各种ECM组分的HSC培养系统可用于研究肝组织中ECM重组的机制，并进一步探索通过控制ECM组分（包括胶原）的合成和分泌以及MMP表达来改善肝纤维化的可能性。

Hepatic stellate cells (HSCs) exhibited a marked change in morphology depending on the extracellular matrix (ECM) component as a substratum, and HSCs were altered to elongate many cellular processes, as seen in vivo, by the culturing using interstirial collagen gel. The results from this study revealed that the process elongation was induced by cell surface integrin binding to ECM components, followed by various intracellular signaling and finally cytoskeleton, particularly, microtubule assembly. As previously reported by Senoo et al., HSCs displayed an alteration not only in morphology, but also in proliferation and collagen synthesis/secretion. The results from gelatin zymography in this study further demonstrated that the conditioned medium from HSC culture on type I collagen gel but not on polystyrene surface and on Matrigel contained an activated form of matrix metalloproteinase (MMP)-2, However, the culture on type I collagen gel also indicated the expression of tissue inhibitor of matrix metalloprotease (TIMP)-2, as detected by reverse transcriptase-polymerase chain reaction (RT-PCR), suggesting the suppression of the activated form of MMP-2. Fluorescence immunostaining and in situ zymography further indicated the presence of MMP- 1 proteins and their activities in the culture on type I collagen gel. Taken together, these results suggest a complicated regulatory mechanism at transcriptional level or at translational/posttranslational stages for reorganization of ECM components such as the basement membrane containing type IV collagen or interstitial matrix components including type I collagen. HSC culture system using various ECM components is, therefore, useful to investigate the mechanism of reorganization of ECM in the liver tissue, and further to explore the possibility to ameliorate liver fibrosis by the control of synthesis and secretion of ECM components including collagen and of MMP expression.