Zeng G, McCue H M, Mastrangelo L, Millis A J
Center for Cellular Differentiation, Department of Biological Sciences, University at Albany, State University of New York, 12222, USA.
Exp Cell Res. 1996 Nov 1;228(2):271-6. doi: 10.1006/excr.1996.0326.
In culture, nontransformed human diploid fibroblasts divide a limited number of times, resulting in a nonproliferating senescent cell culture which exhibits an altered pattern of gene expression. Previously we reported that an early event in the process of replicative senescence was an increase in the synthesis of two connective tissue degrading metalloproteinases, collagenase and stromelysin, and a decrease in the synthesis of the physiological inhibitor of those enzymes, tissue inhibitor of metalloproteinases-1 (TIMP-1). The cytokine TGF-beta1 is known to regulate the expression of each of these three genes and to be synthesized and secreted by cultured human fibroblasts. This suggested the hypothesis that the age-specific modulation of collagenase, stromelysin, and TIMP-1 expression is the result of a change in TGF-beta1 activity during replicative senescence. To test this hypothesis, the responses of early, mid, and late passage (presenescent) fibroblast cell cultures to a TGF-beta neutralizing antibody were evaluated. In early passage cell cultures, exposure to TGF-beta neutralizing antibody resulted in a significant increase in the expression of collagenase and stromelysin and decreased TIMP-1 expression. The antibody did not affect expression of either of those genes by late passage cell cultures, although late passage cultures did respond to added TGF-beta1. Quantification of the levels of active TGF-beta, using a growth inhibition assay, indicates that the level of active TGF-beta1 is decreased during replicative senescence, supporting the conclusion that the modulation of collagenase, stromelysin, and TIMP-1 expression results from diminished TGF-beta activity.
在培养过程中，未转化的人类二倍体成纤维细胞分裂次数有限，导致形成一种不再增殖的衰老细胞培养物，其基因表达模式发生改变。我们之前报道过，复制性衰老过程中的一个早期事件是两种结缔组织降解金属蛋白酶（胶原酶和基质溶素）的合成增加，而这些酶的生理抑制剂金属蛋白酶组织抑制剂 -1（TIMP-1）的合成减少。细胞因子TGF-β1已知可调节这三个基因中每个基因的表达，并且由培养的人类成纤维细胞合成和分泌。这提示了一个假说，即胶原酶、基质溶素和TIMP-1表达的年龄特异性调节是复制性衰老期间TGF-β1活性变化的结果。为了验证这一假说，评估了早代、中代和晚代（衰老前）成纤维细胞培养物对TGF-β中和抗体的反应。在早代细胞培养物中，暴露于TGF-β中和抗体导致胶原酶和基质溶素的表达显著增加，而TIMP-1表达减少。该抗体对晚代细胞培养物中这两种基因的表达均无影响，尽管晚代培养物确实对添加的TGF-β1有反应。使用生长抑制试验对活性TGF-β水平进行定量分析表明，复制性衰老期间活性TGF-β1水平降低，这支持了胶原酶、基质溶素和TIMP-1表达的调节是由于TGF-β活性降低所致的结论。
