青藤碱减轻小鼠肾脏缺血再灌注损伤

Protective effects of sinomenine on renal ischemia/reperfusion injury in mice

目的研究青藤碱对小鼠肾脏缺血再灌注（IR）损伤的作用及其机制。方法C57BL／6小鼠静脉注射青藤碱200mg／k，对照组注射等体积生理盐水，通过检测血清丙氨酸转氨酶（ALT）和血清肌酐（SCr）分析青藤碱对小鼠的肝、肾毒性。将C57BL／6小鼠分为3组，每组6只。假手术（S0）组小鼠仅接受中线开腹、双侧肾蒂游离及关腹操作；青藤碱处理（SIN处理）组小鼠建立肾脏IR损伤模型，并于肾脏缺血前经尾静脉注射青藤碱200mg／k；生理盐水处理（Saline）组小鼠肾脏缺血前经尾静脉注射等体积生理盐水。再灌注后6h检测各组血清尿素氮（BUN）和SCr水平，观察肾脏组织形态学变化，检测各组肾小管上皮细胞凋亡情况，检测各组肾脏组织内巨噬细胞浸润，检测各组肾脏组织内中性粒细胞的浸润，检测各组肾脏组织内肿瘤坏死因子a（TNF-a）、CXC趋化因子配体10（CXCL-10）、细胞间粘附分子1（ICAM-1）和白细胞介素17（IL-17）mRNA的表达，检测肾脏组织内核因子（NF）-kB亚单位p65磷酸化水平。结果青藤碱对小鼠血清ALT和SCr影响的差异无统计学意义（P〉0．05）。再灌注后6h，SIN处理组小鼠血清BUN和SCr水平明显低于Saline组（P〈0．01，P〈0．05）。与Saline组相比较，SIN处理组小鼠肾小管上皮细胞损伤情况较轻（P〈0．01），肾小管上皮细胞凋亡明显减轻（P〈0．05），肾脏组织内巨噬细胞及中性粒细胞浸润明显减弱（P〈0．05），TNF-a、CXCL-10、ICAM-I和IL-17mRNA表达明显降低（P〈0．05，P〈0．01）。结论静脉应用青藤碱200mg／kg对小鼠无明显肝、肾毒性，可减轻肾脏IR损伤，其机制可能与抑制肾脏再灌注后炎症反应有关。

Objective To evaluate the protective effect of sinomenine （SIN） on renal ischemia/ reperfusion （I/R） in mice. Methods In the experiment one, 12 C57BL/6 mice were randomly divided into 2 groups： SIN group （mice were injected with 200 mg/kg SIN by tail vein） and control group （mice were injected with equal volume of saline）. Six and 24 hs later, the serum was collected and the contents of alanine aminotransferase （ALT） and creatinine （SCr） were determined. In the experiment two, C57BL/6 mice were randomly divided into 3 groups： sham-operated （SO） group, SIN group （mice were injected with 200 mg/kg sinomenine just before ischemia induction） and saline group （mice were injected with equal volume of saline at the same time）. At the 6th h after reperfusion, the sera and renal samples subject to IR injury were collected. The SCr and BUN levels in serum were determined and renal histological changes were also examined. The apoptosis of renal tubular epithelial cells was measured by using terminal deoxynucleotidyl transferase mediated dUTP nick end labeling assay. The infiltration of F4/80 positive macrophages was measured by using immunohistochemistry and that of neutrophils with myeloperoxidase （MPO） kits. The mRNA expression of tumor necrosis factor （TNF）-a, chemokine CXC ligand （CXCL）-10, intercellular adhesion molecule （ICAM）-I and IL-17 was detected by using real-tlme reverse transcription PCR. The activation of transcription factor NF-kB was measured by using Western blotting. Results In the experiment one, there was no significant difference in ALT and SCr between the two groups at 6 or 24 h. In the experiment two,levels of SCr and BUN were lower in SIN group （P〈0. 05 or P〈0. 01 ）, histological damage was milder （P〈0. 01 ）, and apoptosis rate of renal tubular epithelial cells apoptosis was lower than in saline group （P〈0. 05）. The infiltration of macrophages, neutrophils and the mRNA expression of TNF-a, CXCL-10, ICAM-1 and IL-17 in the renal tissue in SIN group were reduced as compared with saline group （P〈0. 05 or P〈0. 01）. The activation of NF-kB in SIN group was significantly downregulated as compared with saline group. Conclusion SIN can ameliorate the renal IR injury without hepatic or renal toxicity, which is associated with inhibition of acute inflammatory response induced by reperfusion.