Nitric oxide induces apoptosis in renal tubular epithelial cells through activation of caspase-8
Nitric oxide induces apoptosis in renal tubular epithelial cells through activation of caspase-8
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DOI：
10.1152/ajprenal.00341.2005
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发表时间：
2006-05-01
影响因子：
4.2
通讯作者：
Jevnikar, AM
中科院分区：
文献类型：
--
作者：
Du, CG;Guan, QN;Jevnikar, AM
肾小管上皮细胞（TEC）对凋亡的敏感性或抵抗性对于炎症发作（如移植物排斥）后长期维持肾功能至关重要。TEC凋亡可发生与缺血以及与促炎细胞因子和一氧化氮（NO），所产生的浸润单核细胞。TEC在炎症过程中也能产生大量的NO，但NO对TEC损伤的作用和调控机制尚不清楚。用Annexin-V和碘化丙啶染色的流式细胞仪分析体外TEC中的凋亡。Greiss试剂检测培养上清中NO含量，Western blot检测诱导型NO合成酶（NOS 2/iNOS）和caspase-8蛋白表达。在这里，我们表明，小鼠TEC产生大量的NO响应促炎细胞因子（IFN-γ/TNF-α）通过上调NOS 2，和内源性NO的产生抑制L-NMMA减少TEC细胞凋亡在奎宁刺激的文化。外源性NO（硝普钠）诱导TEC凋亡以及caspase-8激活的剂量依赖性的方式。caspase-8在NO诱导的TEC凋亡中的关键作用被证明是通过使用z-IETD-fetrin抑制caspase-8、使用shRNA沉默caspase-8或通过过表达内源性caspase-8抑制剂c-FLIP（细胞Flice-inhibitory protein）来阻断NO诱导的TEC凋亡。结论：内源性NO和外源性NO均可通过TEC凋亡参与肾损伤。caspase-8的激活在NO诱导的细胞凋亡中起着重要作用，抑制caspase-8可能是肾脏炎症的重要治疗靶点。
The susceptibility or resistance of tubular epithelial cells (TEC) to apoptosis is pivotal to the long-term maintenance of kidney function following episodes of inflammation, such as graft rejection. TEC apoptosis can occur with ischemia as well as with proinflammatory cytokines and nitric oxide ( NO), produced by infiltrating mononuclear cells. TEC can also produce abundant amounts of NO during inflammation but the role and regulation of NO-induced injury of TEC are not well understood. Apoptosis in TEC in vitro was determined by FACS analysis with annexin-V and propidium iodide staining. NO in culture supernatants was measured by Greiss reagent, and protein expression of inducible NO synthetase (NOS2/iNOS) and caspase-8 was examined by Western blot analysis. Here, we showed that murine TEC produced abundant amounts of NO in response to proinflammatory cytokines (IFN-gamma/TNF-alpha) through upregulation of NOS2, and inhibition of endogenous NO production by L-NMMA reduced TEC apoptosis in cytokine-stimulated cultures. Addition of exogenous NO ( sodium nitroprusside) induced TEC apoptosis as well as caspase-8 activation in a dose-dependent manner. The key role of caspase-8 in NO-induced TEC apoptosis was demonstrated by that NO-induced TEC apoptosis can be blocked by caspase-8 inhibition using z-IETD-fmk, caspase-8 silencing with shRNA or by overexpressing the endogenous caspase-8 inhibitor c-FLIP ( cellular Flice-inhibitory protein). In conclusion, endogenous NO from NOS2 activity as well as exogenous NO can contribute to renal injury through apoptosis of TEC. Activation of caspase-8 plays a central role in NO-induced apoptosis and caspase-8 inhibition may be an important therapeutic target during renal inflammation.