绞股蓝总皂苷对非酒精性脂肪肝大鼠Treg/Th17免疫功能的影响

Beneficial Effect of Stevenleaf on the Imbalance between Th17 and Treg in Nonalcoholic Fatty Liver Disease Model Rats

关键词:  绞股蓝总皂苷  非酒精性脂肪肝  Treg/Th17细胞  炎症因子

Key words:  stevenleaf  nonalcoholic fatty liver  Treg/Th17 cells  inflammatory factors

摘要:

目的 研究绞股蓝总皂苷对非酒精性脂肪肝（nonalcoholic fatty liver disease，NAFLD）大鼠肝脏保护作用及免疫调节作用机制。方法 48只SD大鼠随机分为正常对照组，模型对照组，多烯磷脂酰胆碱胶囊组（阳性对照组，150 mg·kg^-1）及绞股蓝总皂苷高、中、低（240，120，60 mg·kg^-1）剂量组，除正常对照组外，其余各组连续饲喂高脂饲料10周，制备NAFLD模型；模型成功后，各组连续给药8周。实验期间，分别于给药0，4，8周眼眶取血检测血清AST、ALT；末次给药后，采用流式细胞技术检测外周血Th17和Treg细胞含量；酶联免疫法检测血清IL-17、IL-10及TNF-α含量。HE染色检查肝组织病理变化和免疫组化法检查肝组织IL-17和Foxp3表达。结果 给药4周，绞股蓝总皂苷高剂量显著降低大鼠血清ALT、AST（P<0.01）；给药8周，绞股蓝总皂苷各剂量均能显著降低血清ALT、AST（P<0.05，0.01）。绞股蓝总皂苷高、中剂量能改善肝组织病变，降低TNF-α水平；高剂量显著降低IL-17、升高IL-10水平（P<0.05，0.01），降低淋巴细胞IL-17含量，升高CD4⁺CD25⁺Treg含量（P<0.05），明显减少炎症因子IL-17的表达和增加Foxp3表达。结论 绞股蓝总皂苷能改善NAFLD大鼠肝脏病变，其作用机制可能与调节肝脏Treg/Th17细胞平衡，减少促炎症因子和增加抗炎因子产生相关。

Abstract:

OBJECTIVE To study the effect of stevenleaf on improving non-alcoholic fatty liver disease in rat model and clarify mechanism from the point of immune-regulation. METHODS Forty-eight of rats were divided into six groups:normal group, model group, Essentiale (150 mg·kg^-1), high-dose (240 mg·kg^-1), moderate-dose (120 mg·kg^-1) and low-dose of stevenleaf (60 mg·kg^-1) groups. Rats were given high fat food for continuous 10 weeks, then given correspond drugs for 8 weeks. The levels of serum AST and ALT were measured at 0, 4 and 8 weeks experimental session. The rats were anesthetized after administration 8 weeks. The peripheral blood lymphocytes and serum were separated to measure the contents of Th17 and Treg cells in peripheral blood by flow cytometry and the levels of IL-17, IL-10 and TNF-α in serum by enzyme-linked immunosorbent assay (ELISA). HE staining was used to observe the pathological changes of liver tissue and immunohistochemistry to observe the positive expression of IL-17 and Foxp3 in liver tissue. RESULTS The 240 mg·kg^-1 of stevenleaf could reduce levels of serum ALT and AST after administration for 4 weeks (P<0.01). stevenleaf 240 mg·kg^-1, 120 mg·kg^-1 and 60 mg·kg^-1could reduce levels of serum ALT and AST after administration for 8 weeks (P<0.05, 0.01). Stevenleaf 240 mg·kg^-1 and 120 mg·kg^-1 could improve liver damage and reduce the level of TNF-α (P<0.05). Stevenleaf 240 mg·kg^-1 could reduce the content of IL-17 and increase IL-10 in serum (P<0.05, 0.01), reduce content of Th17 cell (CD4⁺IL-17⁺) and increase CD4⁺CD25⁺ Treg in peripheral blood lymphocytes (P<0.05), also significantly reduced the expression of inflammatory factor IL-17 and increased Foxp3 expression. CONCLUSION Stevenleaf could protect liver tissue damages, its mechanism may be related to regulating Treg/Th17 cell balance, reducing pro-inflammatory factors and increasing anti-inflammatory factors.