基于权重配方法的降脂颗粒改善非酒精性脂肪性肝病的有效成分配伍优化研究

Optimization of active components compatibility of Jiangzhi Granule to improve NAFLD based on weighed modification method

目的基于改善非酒精性脂肪性肝病(NAFLD)的作用,筛选中药复方降脂颗粒有效成分的最优配伍。方法(1)用脂肪酸诱导HepG2和L02细胞24 h,建立肝细胞脂肪变性模型(模型组,M)。用降脂颗粒的3种有效成分(原人参二醇、丹参酮ⅡA和大黄素)根据权重配方法组成的6组剂量配伍(A、B、C、D、E、F)干预脂肪变性细胞24 h。正常组用常规培养基培养。通过检测油红O染色、细胞三酰甘油(TG)和总胆固醇(TC)含量,观察药物对细胞脂质积聚的影响,筛选出最优配伍组,并与3种有效成分的单独效应进行比较。(2)SPF级雄性C57BL/6J小鼠按体质量随机分为正常组、模型组、配伍F高剂量组和配伍F低剂量组,每组8只。正常组用普通饲料喂养,其余给予高脂饲料,诱导小鼠非酒精性脂肪性肝病模型,造模22周后,予以相应药物干预8周。检测小鼠体质量、肝重、血清丙氨酸转氨酶(ALT)、血脂和肝脏组织病理变化,验证优选配伍的药效。结果(1)细胞油红O染色显示模型组细胞出现大量脂滴积聚,各配伍组药物脂滴含量均不同程度地减少。细胞TG含量检测显示M组中TG含量升高,配伍组C、配伍组F明显降低TG水平,其中配伍组F作用最显著。且配伍组F减轻脂质积聚的作用优于原人参二醇、丹参酮ⅡA、大黄素单独的作用。(2)非酒精性脂肪性肝病小鼠模型组体质量、肝重和血清ALT较对照组明显升高,配伍F高剂量组干预可显著降低肝重和ALT的水平。配伍F高剂量组和配伍F低剂量组均可降低非酒精性脂肪性肝病小鼠血清TC水平,配伍F高剂量组可明显下调高密度脂蛋白胆固醇(HDL-c)和低密度脂蛋白胆固醇(LDL-c)的水平。HE染色和油红O染色表明,配伍F高剂量组和配伍F低剂量组的肝细胞脂肪变性均不同程度地改善,并具有剂量依赖性。结论通过体内体外实验,优选出由原人参二醇、丹参酮ⅡA、大黄素组成的药物配伍F组,可改善非酒精性脂肪性肝病小鼠血脂、肝脂及肝功能,为研发质量可控、疗效稳定的用于治疗非酒精性脂肪性肝病的新型药物建立了实验基础。

Objective To select out the optimal compatibility of Jiangzhi Granule components for improving non-alcoholic fatty liver disease(NAFLD).Methods(1)HepG2 and L02 cells were induced by fatty acids for 24 hours to establish a model of hepatosteatosis(model group,M).Six compatibility groups(named A,B,C,D,E,F)combined by 3 major components of Jiangzhi Granule(including protopanaxadiol,tanshinoneⅡA and emodin)according to the weighed modification method were used to intervene the model cells for 24 h.Cells cultured in the normal medium were used as normal control.The optimal proportion group was selected according to the effect of drugs on alleviating lipid accumulation observed by oil red O staining and intracellular triglyceride(TG)、total cholesterol(TC)content.The effect of the optimal group was also compared with the individual effect of the three active components.(2)SPF male C57 BL/6 J mice were randomly divided into normal group,model group,compatible F high-dose group and compatible F low-dose group according to their body weight,with 8 mice in each group.The mice in the normal group was fed with ordinary feed,and the rest were given high-fat feed.After 22 weeks of inducting NAFLD modeling,the mice were treated with corresponding drugs for 8 weeks.The body weight,liver weight,serum alanine aminotransferase(ALT),serum lipid and hepatic histopathological changes were detected to verify the efficacy of the optimal compatibility.Results(1)Cell oil red O staining showed that a large number of lipid droplets accumulated in the cells of the model group,which were alleviated in different degrees in all of the 6 compatibility groups.The detection of cellular TG content showed that the TG content in the group M increased,while the level of TG decreased significantly in the group C and the group F,among which group F had the most significant effect.Moreover,the effect of group F on reducing lipid accumulation was better than protopanaxadiol,tanshinoneⅡA and emodin alone.(2)The body weight,liver weight and serum ALT of NAFLD mice in model group were significantly higher than those in the control group,and the intervention of compatible F high-dose group could significantly reduce the level of liver weight and ALT.Both compatible F high-dose group and compatible F low-dose group could reduce the level of serum TC in NAFLD mice,and compatible F high-dose group could significantly down-regulate the levels of high density lipoprotein cholesterol(HDL-c)and low density lipoprotein cholesterol(LDL-c).HE staining and oil red O staining showed that compatible F high-dose group and compatible F low-dose group improved hepatocyte steatosis in a dose-dependent manner.Conclusion Through the experiment in vivo and in vitro,the drug compatibility group F composed of propanaxadiol,tanshinoneⅡA and emodin is selected out,which can improve serum lipid,hepatosteatosis and liver function of NAFLD model and helps to establish the experimental basis for the development of new type of drugs for NAFLD with controllable quality and stable therapeutic effects.