Yan Q, Sage E H
Department of Biological Structure, University of Washington, Seattle 98195-7420, USA.
J Cell Biochem. 1998 Jul 1;70(1):70-83.
Transforming growth factor-beta1 (TGF-beta1) regulates a variety of cellular functions. In several types of cells, for example, it acts as a growth inhibitor and an inducer of apoptotic cell death. Although one of the important modulators in retinal vascular development and retinal neovascularization, the effects of TGF-beta1 on retinal microvascular cells are not fully defined. We have found that proliferation of both bovine retinal endothelial cells (EC) and pericytes was inhibited by TGF-beta1 in a concentration-dependent manner. However, only retinal EC lost viability after exposure to increasing concentrations of TGF-beta1 (up to 10 microg/ml) in the presence of 2% fetal bovine serum. Dying EC exhibited the morphological and biochemical characteristics of apoptosis. Fragmented nuclei and chromatin condensation were apparent after staining with the fluorochrome Hoechst 33258 and the reagent ApopTag; moreover, gel electrophoresis of DNA from TGF-beta1-treated EC demonstrated degradation of chromatin into the discrete fragments typically associated with apoptosis. The addition of anti-TGF-beta1 neutralizing antibody abolished the apoptotic cell death induced by TGF-beta1. Because not all the EC in a given culture died after exposure to TGF-beta1, we separated the apoptosis-sensitive cells from those resistant to TGF-beta1 -mediated apoptosis and determined the expression of several proteins associated with this apoptotic pathway. Apoptosis of EC mediated by TGF-beta1 was associated with a decreased level of the cyclin-dependent kinase inhibitor p21waf1/cip1, compared with that observed in the apoptosis-resistant cells. In contrast, the translation product of the tumor-suppressor gene p53 was increased in the TGF-beta1-treated apoptotic cells. Thus, we propose that p21waf1/cip1 and p53 function in distinct pathways that are protective or permissive, respectively, for the apoptotic signals mediated by TGF-beta1.
转化生长因子-β1（TGF-β1）调节多种细胞功能。例如，在几种类型的细胞中，它作为生长抑制剂和凋亡性细胞死亡的诱导剂。尽管TGF-β1是视网膜血管发育和视网膜新生血管形成的重要调节因子之一，但其对视网膜微血管细胞的影响尚未完全明确。我们发现，TGF-β1以浓度依赖性方式抑制牛视网膜内皮细胞（EC）和周细胞的增殖。然而，在含有2%胎牛血清的情况下，仅视网膜EC在暴露于浓度不断增加的TGF-β1（高达10μg/ml）后丧失活力。即将死亡的EC表现出凋亡的形态学和生化特征。用荧光染料Hoechst 33258和试剂ApopTag染色后，细胞核碎片化和染色质浓缩明显可见；此外，来自TGF-β1处理的EC的DNA凝胶电泳显示染色质降解为通常与凋亡相关的离散片段。添加抗TGF-β1中和抗体可消除TGF-β1诱导的凋亡性细胞死亡。由于给定培养物中的并非所有EC在暴露于TGF-β1后都会死亡，我们将对TGF-β1介导的凋亡敏感的细胞与对其有抗性的细胞分离，并确定了与该凋亡途径相关的几种蛋白质的表达。与在抗凋亡细胞中观察到的情况相比，TGF-β1介导的EC凋亡与细胞周期蛋白依赖性激酶抑制剂p21waf1/cip1水平降低有关。相反，在TGF-β1处理的凋亡细胞中，肿瘤抑制基因p53的翻译产物增加。因此，我们提出p21waf1/cip1和p53分别在不同的途径中发挥作用，这些途径对TGF-β1介导的凋亡信号分别具有保护或允许作用。
