化痰祛湿活血方对非酒精性脂肪性肝炎大鼠模型ADPN/PI3K/AKT通路关键蛋白的影响

Effect of Huatan Qushi Huoxue prescription on the key proteins of the ADPN/PI3K/Akt signaling pathway in rats with nonalcoholic steatohepatitis

目的探讨化痰祛湿活血方对非酒精性脂肪性肝炎(NASH)大鼠模型ADPN/PI3K/AKT信号通路关键蛋白的影响。方法72只SD雄性大鼠,适应性喂养1周,随机分为空白组、模型组、多烯组、化痰祛湿活血方高、中、低剂量组,共6组,每组12只。空白组予普通饲料,其余各组予高脂饲料造模。除空白组以外,其余各组大鼠均进行灌胃,模型组灌胃生理盐水(1 ml·100 g-1·d-1),多烯对照组灌胃多烯磷脂酰胆碱胶囊混悬液(0.0285 g·100 g-1·d-1),化痰祛湿活血方高、中、低剂量组分别按5.04 g·100 g-1·d-1、2.52 g·100 g-1·d-1、1.26 g·100 g-1·d-1剂量灌胃。10周后取材,化学发光法检测血清ALT、AST、GGT、ALP、TG、TC水平;Western-Blot检测肝组织ADPN、AdipoR2、PI3K、Akt、pAkt、NF-κB、p-NF-κB蛋白表达水平。计量资料多组间比较采用单因素方差分析,进一步两两比较采用LSD-t检验。结果与空白组比较,模型组肝组织呈现脂肪变,血清ALT、AST、ALP、GGT、TG、TC水平均显著升高(P值均<0.05),肝组织ADPN、AdipoR2、PI3K、Akt、p-Akt蛋白表达均降低(P值均<0.05),NF-κB、p-NF-κB蛋白表达均升高(P值均<0.05)。与模型组相比,各治疗组血清ALT、AST、ALP、GGT、TG、TC水平均降低(P值均<0.05),肝组织ADPN、AdipoR2、pAkt蛋白表达均升高(P值均<0.05),NF-κB、p-NF-κB蛋白表达均降低(P值均<0.05)。与多烯组比较,高剂量组血清ALT、AST、ALP、GGT、TG、TC水平均降低(P值均<0.05),肝组织ADPN、AdipoR2、PI3K、pAkt蛋白表达均升高(P值均<0.05),NF-κB、p-NF-κB蛋白表达均降低(P值均<0.05)。结论化痰祛湿活血方可能通过调控ADPN/PI3K/AKT信号通路,上调ADPN、AdipoR2、PI3K、pAkt蛋白表达,下调NF-κB、p-NF-κB蛋白表达,达到治疗NASH的作用。

Objective To investigate the effect of Huatan Qushi Huoxue prescription on the key proteins of the ADPN/PI3K/Akt signaling pathway in a rat model of nonalcoholic steatohepatitis(NASH).Methods After one week of adaptive feeding,72 male Sprague-Dawley rats were randomly divided into blank group,model group,polyene phosphatidylcholine group,and high-,middle-,and low-dose Huatan Qushi Huoxue prescription groups,with 12 rats in each group.The rats in the blank group were given normal diet,and those in the other groups were given high-fat diet to establish a model of NASH.All rats except those in the blank group were given corresponding drug by gavage.The rats in the model group were given normal saline(1 ml·100 g-1·d-1)by gavage,those in the polyene phosphatidylcholine group were given the suspension of polyene phosphatidylcholine capsules(0.0285 g·100 g-1·d-1)by gavage,and those in the high-,middle-,and low-dose Huatan Qushi Huoxue prescription groups were given Huatan Qushi Huoxue prescription at a dose of 5.04 g·100 g-1·d-1,2.52 g·100 g-1·d-1,and 1.26 g·100 g-1·d-1,respectively.Samples were collected 10 weeks later;chemiluminescence was used to measure the serum levels of alanine aminotransferase(ALT),aspartate aminotransferase(AST),gamma-glutamyl transpeptidase(GGT),alkaline phosphatase(ALP),triglyceride(TG),and total cholesterol(TC),and Western blot was used to measure the protein expression of adiponectin(ADPN),adiponectin receptor 2(AdipoR2),phosphoinositide 3-kinase(PI3K),protein kinase B(Akt),phosphorylated Akt(p-Akt),nuclear factor-kappa B(NF-κB),and phosphorylated NF-κB(p-NF-κB)in liver tissue.A one-way analysis of variance was used for comparison of continuous data between multiple groups,and the least significant difference t-test was used for further comparison between two groups.Results Compared with the blank group,the model group showed fatty change of liver tissue and had significant increases in the serum levels of ALT,AST,ALP,GGT,TG,and TC(all P<0.05),as well as significant reductions in the protein expression of ADPN,AdipoR2,PI3K,Akt,and p-Akt(all P<0.05)and significant increases in the protein expression of NF-κB and p-NF-κB(all P<0.05).Compared with the model group,each treatment group had significant reductions in the serum levels of ALT,AST,ALP,GGT,TG,and TC(all P<0.05),significant increases in the protein expression of ADPN,AdipoR2,and p-Akt(all P<0.05),and significant reductions in the protein expression of NF-κB and p-NF-κB(all P<0.05).Compared with the polyene phosphatidylcholine group,the high-dose Huatan Qushi Huoxue prescription group had significant reductions in the serum levels of ALT,AST,ALP,GGT,TG,and TC(all P<0.05),significant increases in the protein expression of ADPN,AdipoR2,PI3K,and p-Akt(all P<0.05),and significant reductions in the protein expression of NF-κB and p-NF-κB(all P<0.05).Conclusion Huatan Qushi Huoxue prescription may exert a therapeutic effect on NASH by regulating the ADPN/PI3K/Akt signaling pathway,upregulating the protein expression of ADPN,AdipoR2,PI3K,and p-Akt,and downregulating the protein expression of NF-κB and p-NF-κB.