摘要
目的:探讨肿瘤坏死因子-α(TNF-α)对大鼠原代肾小管上皮细胞半胱氨酸天冬氨酸蛋白酶-3(caspase-3)激活与表达的调控以及核因子-κB(NF-κB)在其中的作用。方法:体外分离与培养大鼠肾小管上皮细胞,以TNF-α(10μg/L)按不同时间给予刺激,Western blotting检测caspase-3及其活化裂解片段cleaved caspase-3,以及I-κBα和磷酸化I-κBα(pI-κBα)蛋白水平。分别以TNF-α(10μg/L)处理24h、NF-κB特异性抑制剂Bay11-7082(5μmol/L)处理25h、Bay11-7082(5μmol/L)预处理1h后加入TNF-α(10μg/L)刺激24h,检测caspase-3及cleaved caspase-3的变化。结果:TNF-α刺激6至24h,cleaved caspase-3与caspase-3的相对比值显著高于对照组;而caspase-3蛋白水平在TNF-α刺激后6h无明显增高,12h低于对照组,24h回升;Bay11-7082处理组和Bay11-7082+TNF-α处理组的caspase-3蛋白表达与活化水平显著低于TNF-α处理组和对照组。结论:TNF-α促进大鼠原代肾小管上皮细胞caspase-3的激活与表达,这一过程与NF-κB的激活有关。
AIM： To investigate the production and activation of caspase -3 in primary rat renal proximal tubule cells in response to tumor necrosis factor -α（TNF -α） and the implication of nuclear factor -κB （ NF -κB） in the process. METHODS： Isolated rat renal proximal tubule cells （PTCs） from male adult Sprague Dawley rats were treated with TNF -α according to the indicated time courses. A specific NF -κB inhibitor, Bayl 1 - 7082, was used alone or as a pretreatment for 1 h followed by exposure to TNF -α for 24 h. The protein levels of cleaved caspase - 3, caspase - 3, I -κBα, phosphorylated I -κBot, and GAPDH were detected by Western blotting using specific antibodies. RESULTS： The protein level of cleaved caspase - 5 relative to caspase - 5 was significantly increased in the presence of TNF -α for 6 h, 12 h, and 24 h. Protein levels of caspase - 3 were significantly decreased by 12 h and returned to baseline by 24 h in the presence of TNF -α. Treatment with Bayl 1 - 7082 for 25 h alone or pretreatment with Bayl 1 - 7082 for 1 h followed by addition of TNF -α for 24 h caused a remarkable reduction in both cleaved caspase - 3 and caspase - 3 as compared to control and TNF -α treated groups. An increase in phosphorylated I -κBα was observed from 15 min to 60 min after treatment with TNF-α at a dose of 10 μg/L in PTCs. CONCLUSION： NF-κB is not only associated with the activation of caspase - 3 but also the production of caspase - 3 in primary rat renal proximal tubule cells in response to TNF -α.
出处
《中国病理生理杂志》
CAS
CSCD
北大核心
2010年第1期146-149,共4页
Chinese Journal of Pathophysiology
基金
国家自然科学基金资助项目(No.30671974)
广东省自然科学基金资助项目(No.4009408)
