中华眼镜蛇毒抑制肾缺血再灌注损伤的实验研究

Evaluation of Chinese cobra venoms in preventing the kidney from ischemia/reperfusion injury

目的探讨中华眼镜蛇毒(CCV)对大鼠肾缺血再灌注(I/R)损伤的肾保护作用及其机制。方法雄性SD大鼠32只,随机分为4组。Ⅰ、Ⅱ组建立肾I/R模型,分别于再灌注前0.5h、12h腹腔注射0.1%CCV(0.4mg/kg);Ⅲ组制备肾I/R模型作病理对照:Ⅳ组为假手术组。应用苦味酸法和二乙酰一肟法分别测定大鼠缺血前、再灌注24h的Scr和BUN值。用免疫比浊法测定再灌注0、0.5、2、24h的血清补体C3水平。HE染色光镜下观察肾组织损伤形态学改变,并利用原位凋亡检测法(TUNEL)检测细胞凋亡情况。结果再灌注后24h的Scr和BUN值在Ⅱ组明显降低,与Ⅰ、Ⅲ组相比差异有统计学意义(P<0.05)。补体C3水平在Ⅱ组于再灌注0h显著降低,与其他组0h时比较,差异均有统计学意义(P<0.05):Ⅰ、Ⅲ组C3于再灌注2h明显下降,与再灌注0h比较,差异有统计学意义(P<0.05)。HE染色可见Ⅰ、Ⅲ组损伤较重,以近端小管变性,坏死为主;Ⅱ组病变明显减轻。在Ⅲ和Ⅰ组均可见大量TUNEL阳性细胞,而在Ⅱ组其数量与上两组相比显著减少(P<0.05)。结论肾I/R可明显引起肾组织损伤和功能损害。再灌注前12h用CCV预处理大鼠,能够明显降低补体C3,从而有效抑制补体介导的肾I/R损伤。

Objective To investigate the renal protection of Chinese cobra venoms （CCV） and its mechanism in renal ischemia/reperfusion（I/R）. Methods Thirty-two rats were divided into four groups. 0.1% CCV was separately infused into abdominal cavity at 0.5 h, 24 h before reperfusionin group Ⅰ and Ⅱ. Group Ⅲ suffered from kidney I/R was served as pathological control. Group Ⅳ was sham operation group. BUN and Scr were measured before ischemia and 24 h after reperfusion. Complement C3 was observed at 0, 0.5, 2, 24 h after reperfusion. The kidney samples were examined by HE stain under light microscopy. Apeptosis was detected by terminal deoxynucleotidyl transferase mediated dUTP-biotin in situ nick-end labeling（TUNEL）. Results Significant histological damage, apeptosis of tubular cell and impaired renal function were found in group Ⅰ and Ⅲ .The above indexes decreased to a less extend in group Ⅱ （P 〈 0.05）. Complement C3 in group Ⅱ decreased markedly at the beginning of reperfusion and remained the same level since then, which is significantly different from that in group Ⅰ and Ⅲ. HE staining indicated severe lesion mainly Iocated in the tubules in group Ⅰ and Ⅲ, whereas the severity was markedly relieved in group Ⅱ. A significant amount of TUNEL positive cells were observed in group Ⅰ and Ⅲ as compared to group Ⅱ （P 〈 0.05）. In group Ⅰ and Ⅲ, the C3 level significantly decreased at 2 h following reperfusion as compared to 0 h following reperfusion （P 〈 0.05）. Conclusions I/R could lead to significant histological damage and renal dysfunction. Pretreating the rats with CCV 12 h before reperfusion could reduce C3 notably and prevent the kidney from c3 mediated I/R injury efficiently.