金属硫蛋白2对脂多糖诱导急性肝损伤的保护作用及可能机制

The Protective Effect of Metallothionein 2 on Lipopolysaccharide-induced Acute Liver Injury and Its Possible Mechanism

目的研究金属硫蛋白2(MT2)对脂多糖(LPS)致小鼠急性肝损伤的保护作用及可能机制。方法选择6～8周龄健康雌性MT2敲基因C57/6小鼠及对应的野生型小鼠各12只,随机分成对照组和LPS组,每组包括敲基因小鼠及野生型小鼠各6只,观察两种基因型小鼠经LPS处理后的基本情况。ELISA法检测每组小鼠血清中MT2、肿瘤坏死因子-α(TNF-α)、白介素6(IL-6)、超氧化物歧化酶(SOD)和丙二醛(MDA)浓度;微板法检测血清中谷草转氨酶(AST)和谷丙转氨酶(ALT)的活性;qPCR检测肝脏MT2、TNF-α和IL-6表达情况,苏木素-伊红染色观察肝组织的病变情况;免疫组化检测F4/80+巨噬细胞数量及F4/80蛋白表达。结果 1)对照组中两种基因型小鼠AST、ALT、TNF-α、IL-6、SOD、MDA血清中含量及肝脏中表达差异无统计学意义(P>0.05);注射LPS后,两种基因型小鼠体内AST、ALT、TNF-α、IL-6、MDA均明显增加(P<0.001),敲基因小鼠较野生型小鼠增加更明显(P<0.05);SOD活性明显降低(P<0.001),敲基因小鼠较野生型小鼠降低更明显(P<0.05);2)病理检测结果显示,敲基因小鼠较野生型小鼠肝脏损伤程度更重,肝脏中F4/80+巨噬细胞数目及F4/80表达较野生型小鼠明显增加(P<0.01或P<0.001)。结论 MT2对LPS诱导的急性肝损伤具有明显的保护作用,其机制可能与影响巨噬细胞迁移活化进而调节炎症反应有关。

Objective To study the protective effect of metallothionein 2 （MT2） on lipopolysaccharide （LPS）-induced acute liver injury in mice and its possible mechanism. Methods 12 healthy female MT2 knockout C57/6 mice aged from 6 to 8 weeks and the other 12 healthy female wild-type mice with the same age were selected and divided randomly into the control group and the LPS group, and each group consisted of 6 knockout mice and 6 wild-type mice. The basic situation of those mice was observed after being treated with LPS. The ELISA method was used to detect the levels of MT2, tumor necrosis factor alpha （TNF-α）, interleukin 6 （IL-6）, superoxide dismutase （SOD） and malondialdehyde （MDA） in each group of mice. The microplate method was adopted to detect the activity of aspartate aminotransferase （AST） and alanine aminotransferase （ALT） in serum. The qPCR essay was used to detect the levels of MT2, TNF-α and IL-6 in liver. The hematoxylin-eosin staining method was used to observe the pathological changes of liver tissues. The immunohistochemistry method was used to detect the number of F4/80＋ macrophages and the expression of F4/80 protein. Results Firstly, there were no significant differences in the levels of AST, ALT, TNF-α, IL-6, SOD and MDA in the serum and liver between the mice of two genotypes in the control group （ P 〉0.05）. After being injected with LPS, the levels of AST, ALT, TNF-α, IL-6 and MDA in the mice of two genotypes were significantly increased （ P 〈0.001） and those in the knockout mice were increased more significantly than those in the wild-type mice （ P 〈0.05）; The SOD activity decreased significantly （ P 〈0.001） and that of the knockout mice decreased more significantly than that of the wild-type mice （ P 〈0.05）. Secondly, the results of the pathological examination showed that the degree of liver injury was more severe in the knockout mice than in the wild-type mice, the number of F4/80＋ macrophages in liver and the expression of F4/80 protein in the knockout mice were significantly higher than those in the wild-type mice （ P 〈0.01 or P 〈0.001）. Conclusion MT2 has an obvious protective effect on LPS-induced acute liver injury, and its mechanism may be related to the regulation of the inflammatory response by influencing the migration and activation of macrophages.