Direct involvement of the receptor-mediated apoptotic pathways in cisplatin-induced renal tubular cell death
Direct involvement of the receptor-mediated apoptotic pathways in cisplatin-induced renal tubular cell death
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DOI：
10.1046/j.1523-1755.2003.00709.x
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发表时间：
2003-01-01
影响因子：
19.6
通讯作者：
Iida, M
中科院分区：
文献类型：
--
作者：
Tsuruya, K;Ninomiya, T;Iida, M
背景肿瘤坏死因子（TNF）受体家族成员，如Fas和TNF受体1（TNFR 1），被认为在多种细胞和器官中诱导凋亡。虽然已经假设了一些潜在的情况下，这些受体参与急性肾功能衰竭（ARF）的发病机制，直接证据表明，他们参与肾小管细胞（RTC）的死亡和肾功能不全是不确定的。本研究在两个系统中检测了这些受体在RTC死亡中的作用：（1）顺铂诱导的ARF的体内小鼠和大鼠模型;（2）从C57 BL/6（B6）小鼠、Fas突变型B6-lpr/lpr小鼠和TNFR 1缺陷型小鼠以及正常大鼠肾（NRK 52 E）细胞中分离的小鼠近端肾小管细胞（PTC）。逆转录-聚合酶链反应表明，顺铂诱导的上调Fas，Fas配体和TNF-α的mRNA在肾脏在体内和RTCs在体外，在小鼠和大鼠。相反，TNFR 1 mRNA的水平是相当大的，但没有改变顺铂的反应。在B6 PTC中与顺铂孵育后，通过酶联免疫吸附试验（ELISA）测定的细胞培养基中TNF-α的产生和通过荧光激活细胞分选仪（FACS）分析测定的Fas表达增加。为了研究Fas和TNFR 1是否直接参与RTC死亡和肾功能不全，我们使用细胞活力测定和流式细胞术分析与荧光素异硫氰酸酯结合的膜联蛋白V和碘化丙啶染色比较细胞对顺铂的耐药性。B6 PTC中细胞活力丧失和细胞死亡（凋亡和坏死）的比率高于其他细胞，而Fas突变型PTC和TNFR 1缺陷型PTC之间的比率相当。Caspase-8活性在B6 PTC中增加，但在Fas突变型PTC和TNFR 1缺陷型PTC中不增加. B6小鼠肾功能不全和RTC死亡（凋亡和坏死）均较B6小鼠严重。基于这些数据，我们得出结论，Fas和TNFR 1介导的凋亡途径直接参与顺铂诱导RTC死亡过程的发病机制。
Background. Tumor necrosis factor (TNF) receptor family members, such as Fas and TNF receptor 1 (TNFR1), are thought to induce apoptosis in a variety of cells and organs. Although a number of potential scenarios have been postulated for the involvement of these receptors in the pathogenesis of acute renal failure (ARF), direct evidence for their involvement in death of renal tubular cells (RTCs) and renal dysfunction is preliminary.Methods. This study examined the roles of these receptors in RTC death in two systems: (1 ) in vivo murine and rat models of cisplatin-induced ARF, and (2 ) murine proximal tubular cells (PTCs), which were isolated from C57BL/6 (B6) mice, Fas -mutant B6-lpr/lpr mice and TNFR1 -deficient mice, and normal rat kidney (NRK52E) cells in vitro.Results. Reverse transcription-polymerase chain reaction indicated cisplatin-induced up-regulation of Fas, Fas ligand and TNF-alpha mRNAs in the kidney in vivo and in RTCs in vitro, both in mice and rats. In contrast, the level of TNFR1 mRNA was substantial but did not change in response to cisplatin. TNF-alpha production in cell culture medium determined by enzyme-linked immunosorbent assay (ELISA) and Fas expression determined by fluorescence-activated cell sorter (FACS) analysis increased following incubation with cisplatin in B6 PTCs. In order to examine whether Fas and TNFR1 are directly involved in RTC death and renal dysfunction, we compared cell resistance to cisplatin using a cell viability assay and FACS analysis with fluorescein isothiocyanate-conjugated annexin V and propidium iodide staining. The ratios of cell viability loss and cell death, both from apoptosis and necrosis, were higher in B6 PTCs than in other cells, while the ratios were comparable between Fas -mutant PTCs and TNFR1 -deficient PTCs. Caspase-8 activity was increased in B6 PTCs, but not in Fas -mutant PTCs and TNFR1 -deficient PTCs. Furthermore, the renal dysfunction and RTC death, both apoptosis and necrosis, induced by cisplatin were more severe in B6 mice in vivo.Conclusion. Based on these data, we conclude that the Fas- and TNFR1-mediated apoptotic pathways are directly involved in the pathogenesis of cisplatin-induced RTC death process.