褪黑素对缺血再灌注损伤大鼠肾脏的保护作用及其作用机制研究

Protective effect of melatonin on renal ischemia reperfusion injury and its mechanism in rats

目的探讨褪黑素对肾缺血再灌注损伤大鼠肾脏的保护作用及其机制。方法通过无创血管夹夹闭双侧肾动静脉建立大鼠肾缺血再灌注模型,将实验大鼠随机分为假手术组、缺血再灌注组、褪黑素5mg组、褪黑素10mg组和VC组,检测再灌注后6h和24h各组大鼠血清尿素氮(serum urea nitrogen,SUN)和肌酐(serum creatinine,Scr),肾脏线粒体活力、超氧化物歧化酶(superoxide dismutase,SOD)活性、总三磷酸腺苷(ATP)酶活性、丙二醛(malondialdehyde,MDA)含量、Bax、Bcl-2水平,一氧化氮(nitric oxide,NO)含量及一氧化氮合酶(endothelial nitric oxide synthase,eNOS)表达。结果褪黑素5mg、褪黑素10mg及VC组再灌注后6h及24h的血清SUN与缺血再灌注组相比明显降低(t值分别为6h后3.154,3.268,3.415,24h后3.864,4.327,4.586;P值分别为6h后0.032,0.029,0.014;24 h后0.006,0.005,<0.001),Scr与缺血再灌注组相比亦明显降低(t值分别为6h后3.021,3.215,3.534,24h后3.856,4.026,721;P值分别为6h后0.037,0.026,0.012;24h 0.007,0.003,<0.001),肾脏线粒体MDA含量明显降低(t值分别为6h后4.241,5.386,6.352,24h后5.436,6.174,6.053;P值分别为6h后0.004,0.001,<0.001,24h后<0.001,<0.001,<0.001);褪黑素5mg、褪黑素10mg及VC组再灌注后6及24h与缺血再灌注组相比的线粒体活力明显升高(t值分别为6h后1.894,3.762,3.758,24h后0.864,4.143,3.972;P值分别为6h后0.097,0.016,0.017,24h后0.247,0.008,0.011)、SOD活性明显升高(t值分别为6h后16.154,19.637,18.026,24h后15.587,18.596,P值分别为6h后<0.001,<0.001,<0.001,24h后<0.001,<0.001,<0.001)、总ATP酶活性与缺血再灌注组相比均明显升高(t值分别为6h后4.361,5.397,5.403,24h后3.327,4.832,4.851;P值分别为6h后0.004,<0.001,<0.001,24h后0.031,<0.001,<0.001)。上述差异均有统计学意义。结论缺血前和再灌注早期给予褪黑素,可有效减轻肾缺血/再灌注损伤(renal ischemia/reperfusion injury,RIRI)大鼠肾损伤,改善肾功能。其机制可能与其保护线粒体功能,增加抗氧化酶活性,增加保护性NO的含量,减少细胞凋亡等作用有关。

Objective To investigate the protective effect of melatonin （MT） on renal ischemia reperfusion injury and its mechanism in rats. Methods The rat renal ischemia reperfusion model was established by clamping bilateral renal arteries and veins with non-invasive vascular clamp. The rats were then randomly divided into sham operation group, ischemia reperfusion （IR） group, MT 5mg group, MT 10mg group and VC group. Serum BUN and Cr, mitochondrial activity, SOD activity, total ATPase activity, malondialdehyde （MDA） content, nitric oxide （NO） content and eNOS expression in renal tissue, and Bax and Bcl-2 expression levels in renal cortex were detected after reperfusion for 6h and 24h. Results After reperfusion for 6h and 24h and compare to IR group, serum BUN and Cr were significantly decreased in MT 5mg group, MT 10mg group and VC group （for MT 5mg group： t=3.154, P=0.032; t=3.864, P=0.006; t=3.021, P=0.037; t=3.856, P=0.007; for MT 10mg group： t=3.268, P=0.029; t=4.327, P=0.005; t=3.215, P=0.026; t=4.026, P=0.003; for VC group： t=3.415, P=0.014; t=4.586, P〈0.001; t=3.534, P=0.012; t=721, P〈0.001）; kidney mitochondrial activity, SOD activity and total ATP activity were significantly increased in MT 5mg group, MT 10mg group and VC group （MT 5mg group： t=1.894, P=0.097; t=0.864, P=0.247; t=16.154, P〈0.001; t=15.587, P〈0.001; t=4.361, P=0.004; t=3.327, P=0.031; MT 10mg group： t=3.762, P=0.016; t=4.143, P=0.008; t=19.637, P〈0.001; t=18.596, P〈0.001; t=5.397, P〈0.001; t=4.832, P〈0.001; VC group： t=3.758, P=0.017; t=3.972, P=0.011; t=18.026, P〈0.001; t=17.923, P〈0.001; t=5.403, P〈0.001; t=4.851, P〈0.001）; MDA decreased significantly in MT 5mg group, MT 10mg group and VC group （MT 5mg group： t=4.241, P=0.004; t=5.436, P〈0.001; MT 10mg group： t=5.386, P=0.001; t=6.174, P〈0.001; VC group： t=6.352, P〈0.001; t=6.053, P〈0.001）. NO increased significantly in MT 5mg group, MT 10mg group and VC group after reperfusion for 6h as compared that in IR group （MT 5mg group： t=5.821, P〈0.001; MT 10mg group： t=9.107, P〈0.001; VC group： t=8.728, P〈0.001）. Conclusions Injection of MT at the early stage of ischemia reperfusion can effectively reduce the mild renal injury and improve renal function in renal ischemia reperfusion injury rats. Its mechanism may relate to the protection of mitochondrial function, the increase of antioxidant enzyme activity, the increase of protective NO content, and the reduction of apoptosis.