聚肌胞对CCl_4诱导肝损伤小鼠模型肝内自然杀伤细胞的调节作用

Regulatory effect of poly I∶C on intrahepatic natural killer cells in mouse model of liver injury induced by carbon tetrachloride

目的观察聚肌胞(Poly I∶C)调节肝脏自然杀伤(NK)细胞活性抗CCl_4诱导肝损伤的作用机制。方法 80只C57BL/6小鼠按体质量分层随机分为正常对照组、Poly I∶C对照组、模型对照组和Poly I∶C组。模型对照组和Poly I∶C组予腹腔注射10%CCl_4(2 ml·kg^(-1)·2 d^(-1))诱导肝损伤模型,正常对照组与Poly I∶C对照组予腹腔注射等剂量橄榄油。Poly I∶C对照组和Poly I∶C组均予Poly I∶C腹腔注射(1 mg·kg^(-1)·2 d^(-1)),正常对照组与模型对照组均予腹腔注射等体积生理盐水。连续用药4周后,观察血清肝功能及肝组织炎症因子水平;HE染色观察肝组织炎症情况;流式细胞术检测肝脏NK细胞及细胞表面激活型受体NKG2D、抑制型受体Ly49A表达。计量资料组间比较采用方差分析,方差齐时采用LSD法,方差不齐时采用Dunnett T3法。结果 Poly I∶C给药1周和4周,正常小鼠肝组织均无明显肝损伤。随CCl_4染毒加重,与正常对照组相比,模型对照组血清ALT、AST和TBil水平逐渐升高(P<0.01),肝内促炎因子水平上调(P<0.05),HE染色提示肝细胞大量坏死,肝脏NK细胞比例及NKG2D表达减少、Ly49A表达增多(P<0.05)。Poly I∶C可改善肝损伤小鼠血清肝功能,降低肝内促炎因子,明显改善肝组织炎症,增加肝脏NK细胞比例、数量及其细胞表面激活型受体NKG2D受体表达(P<0.01)。结论 Poly I∶C具有良好的抗急、慢性肝损伤作用,其抗肝损伤的作用机制与对NK细胞的激活密切相关。

Objective To investigate the effect of polyinosinic- polycytidylic acid（ poly I∶ C） on the activation of natural killer cells（ NK cells）in the mouse model of liver injury induced by carbon tetrachloride（ CCl4）. Methods Eighty C57BL/6 mice were randomly divided into normal control group,poly I∶ C control group,model control group,and poly I∶ C group. The mice in model control group and poly I∶ C group were intraperitoneally injected with 10% CCl4（ 2 ml·kg^-1·2d^-1） to establish the model of liver injury,and the mice in normal control group and poly I∶C control group were intraperitoneally injected with the same dose of olive oil. The mice in poly I∶ C control group and poly I： C group were intraperitoneally injected with poly I∶ C（ 1 mg·kg^-1·2d^-1）,and the mice in normal control group and model control group were intraperitoneally injected with the same volume of normal saline. After continuous administration for 4 weeks,serum levels of liver function indices and liver inflammatory factors were observed; HE staining was applied to observe liver inflammation grade; flow cytometry was applied to measure NK cells in the liver,as well as the expression of the activating receptor NKG2 D and inhibitory receptor Ly49 A on NK cells. Analysis of variance was applied for comparison between groups,with least significant difference（ LSD） test applied for homogeneity of variance and Dunnett＇s T3 method applied for heterogeneity of variance. Results After administration of poly I∶ C for 1 week and 4 weeks,there was no obvious liver injury in normal mice. With aggravation of CCl_4 poisoning,the model control group,compared with the normal control group,had gradually increased serum levels of alanine aminotransferase,aspartate aminotransferase,and total bilirubin（ P〈0. 01） and upregulated levels of intrahepatic proinflammatory factors（ P〈0. 05）; HE staining suggested necrosis of a large number of hepatocytes,a decreased proportion of hepatic NK cells,reduced expression of NKG2 D,and increased expression of Ly49 A in the model control group（ P〈0. 05）. Poly I∶ C improved the serum levels of liver function indices,reduced the levels of intrahepatic proinflammatory factors,significantly improved liver inflammation,and increased the proportion and number of hepatic NK cells and expression of NKG2D（ P〈0. 01）. Conclusion Poly I∶ C can protect the liver from acute and chronic liver injury,and its mechanism of action is closely related to activation of NK cells.