Department of Nephrology, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430022, Peoples Republic of China.
Cell Biol Int. 2011 Aug;35(8):827-34. doi: 10.1042/CBI20100411.
Proteinuria is a well-established exacerbating factor of chronic kidney diseases. However, the harmful effects of protein overload on podocytes and the underlying mechanisms are still poorly understood. In the present study, we examined the effects of high concentrations of albumin on podocytes and investigated the role of CD2AP (CD2-associated protein) in albumin overload-induced podocyte apoptosis. Conditionally immortalized mouse podocytes were cultured in vitro and treated with different concentrations of BSA. In addition, CD2AP eukaryotic expression vector or siRNA (small interfering RNA) was transfected into podocytes before they were exposed to BSA. Podocyte apoptosis, expressions of active caspase-3 (p17) and CD2AP, and the distribution of F-actin cytoskeleton were detected by flow cytometry, Western-blot analysis and fluorescent staining respectively. It was found that exposure of podocytes to BSA induced podocyte apoptosis in a concentration-dependent manner that was accompanied by up-regulation of active caspase-3, the disruption of F-actin cytoskeleton, and decreased expression of CD2AP. Transfection of CD2AP eukaryotic expression vector into podocytes increased CD2AP expression, partially restored F-actin distribution, blocked active caspase-3 expression and inhibited podocyte apoptosis. In contrast, transfection of CD2AP siRNA deteriorated the above changes induced by BSA. It is concluded that protein overload induces podocyte apoptosis via the down-regulation of CD2AP and subsequent disruption of cytoskeleton of podocytes, and CD2AP may play an important role in protein overload-induced podocyte injury.
蛋白尿是慢性肾脏病的一个明确的恶化因素。然而，蛋白过载对足细胞的有害影响及其潜在机制仍知之甚少。在本研究中，我们研究了高浓度白蛋白对足细胞的影响，并探讨了 CD2AP（CD2 相关蛋白）在白蛋白过载诱导的足细胞凋亡中的作用。将条件永生化的小鼠足细胞进行体外培养，并以不同浓度的 BSA 处理。此外，在 BSA 处理之前，将 CD2AP 真核表达载体或 siRNA（小干扰 RNA）转染到足细胞中。通过流式细胞术、Western blot 分析和荧光染色分别检测足细胞凋亡、活性 caspase-3（p17）和 CD2AP 的表达以及 F- 肌动蛋白细胞骨架的分布。结果发现，BSA 暴露于足细胞中，诱导足细胞凋亡呈浓度依赖性，同时伴有活性 caspase-3 的上调、F-肌动蛋白细胞骨架的破坏和 CD2AP 的表达下调。将 CD2AP 真核表达载体转染到足细胞中，增加了 CD2AP 的表达，部分恢复了 F-肌动蛋白的分布，阻断了活性 caspase-3 的表达，抑制了足细胞凋亡。相反，CD2AP siRNA 的转染恶化了 BSA 诱导的上述变化。结论是，蛋白过载通过下调 CD2AP 并随后破坏足细胞的细胞骨架诱导足细胞凋亡，CD2AP 可能在蛋白过载诱导的足细胞损伤中起重要作用。
