亚低温对小鼠肾脏缺血再灌注损伤的保护作用

Mild hypothermia protect renal function on ischemia-reperfusion kidney: an experimental study in mice

目的探讨亚低温(MH)对小鼠肾脏缺血再灌注损伤(IRI)的保护作用及相关分子机制。方法取24只8周龄无特定病原体级雄性BALB/c小鼠随机分为缺血再灌注(IR)组、亚低温缺血再灌注(MH+IR)组和正常对照(NC)组,每组各8只。另取15只同种系小鼠单纯行亚低温处理0、2、4、6和8 h,每组各3只。小鼠IR 24 h后,检测IR组、MH+IR组和NC组血清肌酐、冷诱导核糖核酸结合蛋白(CIRP)、硫氧还蛋白(Trx)、B淋巴细胞瘤-2基因蛋白(Bcl-2)、Bcl-2相关X蛋白(Bax)、超氧化物歧化酶(SOD)活性和丙二醛(MDA)含量。采用HE染色法检测肾小管上皮细胞水肿情况。采用末端脱氧核苷酸转移酶介导的三磷酸脱氧尿苷缺口末端标记(TUNEL)法检测肾组织细胞凋亡情况。采用蛋白免疫印迹法检测单纯行MH处理0、2、4、6和8 h的小鼠肾脏CIRP蛋白表达。采用单因素方差分析比较肾脏IR 24 h后3组血清肌酐值、肾组织细胞凋亡比例、SOD活性和MDA含量。P<0.05为差异有统计学意义。结果 IR组和MH+IR组血清肌酐分别为(153.6±21.3)和(87.4±13.8)mg/d L,差异有统计学意义(F=2.38,P<0.05);MH+IR组肾小管上皮细胞水肿明显轻于IR组。MH+IR组和IR组细胞凋亡比例分别为(15.0±2.4)%和(37.0±5.7)%,差异有统计学意义(F=5.41,P<0.05)。MH上调CIRP、Trx蛋白表达,同时提高Bcl-2/Bax比值。MH+IR组和IR组SOD活性分别为(26.9±4.4)和(16.8±2.4)U/mgprot,差异有统计学意义(F=3.36,P<0.05)。MH+IR组和IR组MDA含量分别为(0.76±0.18)和(1.37±0.32)nmol/mgprot,差异有统计学意义(F=3.21,P<0.05)。结论 MH能够显著减轻小鼠肾脏IRI,可能与其降低氧化应激损伤有关。

Objective To investigate the protective effect of mild hypothermia on ischemiareperfusion kidney and the possible mechanisms. Methods Twenty-four male BALB/c mice were randomly divided into three groups： ischemia-reperfusion group（IR Group）, mild hypothermia ischemia-reperfusion group（MH ＋ IR Group） and normal control group（NC Group）,with 8 mice in each group. Another 15 male BALB/c mice were divided into 5 groups, and dealed with mild hypothermia in time gradient（0,2,4,6 and 8 h）. The serum creatinine,cold inducible ribonucleic acid-binding protein（CIRP）,thioredoxin（Trx）,B-cell lymphoma-2（Bcl-2）,Bcl-2 associated X protein（Bax）,superoxide dismutase（SOD） and malondialdehyde（MDA） of kidney tissue of the abovethree groups were detected 24 h later after ischemia-reperfusion. HE staining was used to detected edema of renal tubular epithelial cell. Terminal dexynucleotidyl transferase-mediated deoxyuridine triphosphate nick end labeling was used to detected apoptosis of kidney tissue. CIRP expression of 15 male BALB/c mice which just dealed with mild hypothermia was detected. One-way analysis of variance was used to compared serum creatinine,apoptotic cell proportion of kidney tissue,SOD activity and MDA content of the above three groups 24 h later after ischemia-reperfusion. P〈0. 05 was considered statistically significant. Results The concentration of serum creatinine of IR group and（MH ＋IR） group were（153. 59 ± 21. 29） and（87. 44 ± 13. 81） mg/d L,which has statistical significance（F = 5. 41,P〈0. 05）. The edema of renal tubular epithelial cell in IR group was worse than（MH ＋IR） group. The apoptotic cell proportion of kidney tissue of IR group and（MH ＋ IR） group were（37. 0 ± 5. 7） % and（15. 0 ± 2. 4） %,which has statistical significance（F = 5. 41,P〈0. 05）. MH could promote the expression of CIRP and Trx,and increase the ratio of Bcl-2/Bax. The SOD activity of（MH ＋ IR） group and IR group were（26. 9 ± 4. 4） and（16. 8 ± 2. 4） U/mgprot,which has statistical significance（F = 3. 36,P〈0. 05）. The MDA content of（MH ＋ IR） group and IR group were（0. 76 ± 0. 18） and（1. 37 ± 0. 32） nmol/mgprot,which has statistical significance（F = 3. 21,P〈0. 05）. Conclusion Mild hypothermia protect mice kidney from ischemia-reperfusion damages,which may by reducing oxidative stress injury.