大麻素受体在肝星状细胞活化中的作用及姜黄素干预效应

目的探究大麻素受体在肝星状细胞(HSCs)活化中的作用及姜黄素的干预效应,为阐明肝纤维化发生发展机理及姜黄素抗肝纤维化机制提供依据。方法 MTT法检测CBR1激动剂NADA与拮抗剂AM251,CBR2激动剂JWH015与拮抗剂AM630对HSCs增殖的影响;Western blot实验检测AM251对HSCs中ERK、JNK、p38蛋白表达及磷酸化水平的影响;Western bolt与细胞免疫荧光检测了姜黄素对HSCs中两种类型大麻素受体CBR1与CBR2蛋白表达的影响;Western bolt检测在姜黄素对CBR1激动剂与拮抗剂的刺激下HSCs产生的细胞外基质(ECM)成分α1(I)collagen和Fibronectin的影响。结果激动CBR1可促进HSCs的增殖,相反拮抗CBR1后则可以抑制HSCs的增殖(P<0.05);而激动CBR2也可抑制HSCs的增殖(P<0.05),但抑制CBR2对HSCs的增殖没有明显的影响(P>0.05)。CBR1拮抗剂AM251能够明显抑制ERK与JNK的磷酸化,并呈剂量依赖性的关系(P<0.05,P<0.01),但对p38蛋白表达的影响较小(P>0.05)。姜黄素可抑制HSCs中CBR1的表达(P<0.05),对CBR2蛋白表达的影响则差异没有统计学意义(P>0.05)。姜黄素可呈剂量依赖性的抑制CBR1激动剂导致的ECM成分表达的上升,并可协同CBR1拮抗剂抑制HSCs表达ECM成分的作用(P<0.05,P<0.01)。结论大麻素受体在HSCs的增殖活化过程中具有重要作用,姜黄素可能通过干预大麻素受体信号通路这一途径达到治疗肝纤维化

Research on the role of cannabinoid receptors in hepatic stellate cell activation and interfering effects of curcumin

Aim To explore the role of cannabinoid receptors in the activation of hepatic stellate cells(HSCs) and the interfering effects of curcumin in the hope of providing basis for elucidating the mechanism of liver fibrosis and curcumin inhibition of liver fibrosis.Methods Influence of CBR1 agonist NADA and antagonist AM630,CBR2 agonist JWH015 and antagonist AM251 on the proliferation of HSCs was evaluated by MTT assay.Western blot assays were used to detect the expression of ERK,JNK and p38 and their phosphorylation levels in HSCs treated with AM251.The effect of curcumin on the expression of two types of cannabinoid receptors CBR1 and CBR2 in HSCs was determined by Western blot and immunofluorescence.The effect of curcumin on extracellular matrix(ECM) components α1(I) collagen and fibronectin in HSCs stimulating by CBR1 agonist and antagonist was also examined by Western blot.Results Activating CBR1 promoted the proliferation of HSCs;on the contrary,CBR1 antagonism inhibited HSCs proliferation(P<0.05).At the same time,activating CBR2 also inhibited the proliferation of HSCs(P<0.05),but CBR2 inhibition had no apparent effect on the proliferation of HSCs(P>0.05).CBR1 antagonist AM251 significantly inhibited the phosphorylation of ERK and JNK in a dose-dependent manner(P<0.05,P<0.01),but p38 was not affected(P>0.05).Curcumin inhibited the expression of CBR1(P<0.05),but at the same time,it had no significant effect on CBR2(P>0.05).Curcumin inhibited the expression of ECM components upregulated by CBR1 agonist dose-dependently in HSCs,and collaboratively inhibited the expression of ECM components in HSCs exposed to CBR1 antagonist(P<0.05,P<0.01).Conclusions Cannabinoid receptors play an important role in the process of HSCs activation.Curcumin may achieve the purpose of treating liver fibrosis through the intervention of cannabinoid receptor pathways.