CAJ | 학술논문

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(10 μg/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(10 μg/L)处理24 h、NF-κB特异性抑制剂Bay11-7082(5 μmol/L)处理25 h、Bay11-7082(5 μmol/L)预处理1 h后加入TNF-α(10 μg/L)刺激24 h,检测caspase-3及cleaved caspase-3的变化.结果: TNF-α刺激6至24 h,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6 h无明显增高,12 h低于对照组,24 h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.
목적: 탐토종류배사인자-α(TNF-α)대대서원대신소관상피세포반광안산천동안산단백매-3(caspase-3)격활여표체적조공이급핵인자-κB(NF-κB)재기중적작용.방법: 체외분리여배양대서신소관상피세포,이TNF-α(10 μg/L)안불동시간급여자격, Western blotting검측caspase-3급기활화렬해편단cleaved caspase-3,이급I-κBα화린산화I-κBα(pI-κBα)단백수평.분별이TNF-α(10 μg/L)처리24 h、NF-κB특이성억제제Bay11-7082(5 μmol/L)처리25 h、Bay11-7082(5 μmol/L)예처리1 h후가입TNF-α(10 μg/L)자격24 h,검측caspase-3급cleaved caspase-3적변화.결과: TNF-α자격6지24 h,cleaved caspase-3여caspase-3적상대비치현저고우대조조;이caspase-3단백수평재TNF-α자격후6 h무명현증고,12 h저우대조조,24 h회승;Bay11-7082처리조화Bay11-7082+TNF-α처리조적caspase-3단백표체여활화수평현저저우TNF-α처리조화대조조.결론: TNF-α촉진대서원대신소관상피세포caspase-3적격활여표체,저일과정여NF-κB적격활유관.
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,Bay11-7082,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-κBα,and GAPDH were detected by Western blotting using specific antibodies. RESULTS:The protein level of cleaved caspase-3 relative to caspase-3 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 Bay11-7082 for 25 h alone or pretreatment with Bay11-7082 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.