软肝饮对HBV转基因小鼠HBV-DNA、肝纤维化四项指标及炎症细胞因子表达的影响

Effect of Ruangan Decoction on HBV-DNA,Four Markers of Liver Fibrosis,and Expression of Inflammatory Cytokines in HBV Transgenic Mice

目的探讨中药软肝饮对乙型肝炎病毒(hepatitis B virus,HBV)转基因小鼠HBV-DNA、肝纤维化四项指标及炎症细胞因子表达的影响。方法将50只雌雄各半的HBV转基因小鼠随机分为中药软肝饮治疗组(高剂量组、中剂量组、低剂量组)、恩替卡韦组、模型组和空白对照组,各组分别连续给药4周后取材。采用实时荧光定量PCR法检测外周血及肝组织中HBV-DNA表达水平,ELISA法检测血清中肝纤维化四项指标层粘连蛋白(laminin,LN)、透明质酸(haluronic acid,HA)、Ⅳ型胶原(collage typeⅣ,Ⅳ-C)、Ⅲ型前胶原(procollagen typeⅢ,PCⅢ)、炎症细胞因子白细胞介素2(interleukin-2,IL-2)、白细胞介素4(interleukin-4,IL-4)、白细胞介素10(interleukin-10,IL-10)、干扰素γ(interferonγ,IFN-γ)]水平。结果与模型组比较,软肝饮高、中、低剂量组和恩替卡韦组小鼠血清和肝组织中HBV-DNA表达水平明显降低,差异有统计学意义(P<0.05);软肝饮的作用呈明显的剂量依赖性(P<0.05)。与空白对照组比较,模型组小鼠血清LN、HA、Ⅳ-C、PCⅢ表达水平显著升高(P<0.05);与模型组比较,软肝饮高剂量组和恩替卡韦组LN、HA、Ⅳ-C、PCⅢ表达水平显著降低(P<0.05);软肝饮降低血清LN、HA、Ⅳ-C的作用呈剂量依赖性(P<0.05)。与空白对照组比较,模型组IL-2、IL-10、IFN-γ水平显著降低(P<0.05),IL-4水平显著升高(P<0.05);与模型组比较,软肝饮高、中剂量组和恩替卡韦组IL-4水平显著降低(P<0.05),IL-2、IL-10、IFN-γ水平显著升高(P<0.05);与软肝饮低剂量组比较,软肝饮高、中剂量组IL-4水平显著降低(P<0.05),IL-10水平显著升高(P<0.05)。结论软肝饮治疗HBV转基因小鼠发挥抗病毒、抗纤维化作用机制可能与其调控炎症细胞因子水平有关。

Objective To investigate the effect of the traditional Chinese medicine Ruangan Decoction on hepatitis B virus(HBV)-DNA,four markers of liver fibrosis,and expression of inflammatory cytokines in HBV transgenic mice.Methods A total of 50 HBV transgenic mice(25 male mice and 25 female mice)were randomly divided into high-,middle-,and low-dose Ruangan Decoction treatment groups,entecavir group,model group,and blank control group,and related samples were collected after 4 consecutive weeks of administration.Quantitative real-time PCR was used to measure the level of HBV-DNA in peripheral blood and liver tissue;ELISA was used to measure the serum levels of the four markers of liver fibrosis[laminin(LN),hyaluronic acid(HA),collage typeⅣ(Ⅳ-C),and procollagen typeⅢ(PCⅢ)]and inflammatory cytokines[interleukin-2(IL-2),interleukin-4(IL-4),interleukin-10(IL-10),and interferon-γ(IFN-γ)].Results Compared with the model group,the high-,middle-,and low-dose Ruangan Decoction treatment groups and the entecavir group had a significant reduction in the level of HBV-DNA in serum and liver tissue(P<0.05),and the effect of Ruangan Decoction was dose-dependent(P<0.05).Compared with the blank control group,the model group had significant increases in the serum levels of LN,HA,Ⅳ-C,and PCⅢ(P<0.05);compared with the model group,the high-dose Ruangan Decoction treatment group and the entecavir group had significant reductions in the levels of LN,HA,Ⅳ-C,and PCⅢ(P<0.05);Ruangan decoction reduced the serum levels of LN,HA,andⅣ-C in a dose-dependent manner(P<0.05).Compared with the blank control group,the model group had significant reductions in the levels of IL-2,IL-10,and IFN-γ(P<0.05)and a significant increase in the level of IL-4(P<0.05);compared with the model group,the high-and middle-dose Ruangan Decoction treatment groups and the entecavir group had a significant reduction in the level of IL-4(P<0.05)and significant increases in the levels of IL-2,IL-10,and IFN-γ(P<0.05);compared with the low-dose Ruangan Decoction treatment group,the high-and middle-dose Ruangan Decoction treatment groups had a significant reduction in the level of IL-4(P<0.05)and a significant increase in the level of IL-10(P<0.05).Conclusion Ruangan Decoction exerts antiviral and anti-fibrotic effects on HBV transgenic mice possibly by regulating the levels of inflammatory cytokines.