健脾化瘀方通过调节RASSF1A激活p38 MAPK信号通路治疗肝细胞癌的机制研究  

Mechanism of Jianpi Huayu Decoction Treating Hepatocellular Carcinoma by Regulating Ras-associated Domain Family 1 Subtype A and Activating the p38 Mitogen-activated Protein Kinase Signaling Pathway

作　　者：吴慧 王紫莹[1,2] 陈小凯 陈嘉毅 陈思儒 罗晓琦 徐恩玲 王昌俊 WU Hui;WANG Ziying;CHEN Xiaokai;CHEN Jiayi;CHEN Siru;LUO Xiaoqi;XU Enling;WANG Changjun(School of Medicine,South China University of Technology,Guangdong Guangzhou 510006,China;Department of Traditional Chinese Medicine,Guangdong Provincial People’s Hospital Affiliated to Southern Medical University,Guangdong Guangzhou 510080,China;Department of Pharmacy,Guangdong Provincial People’s Hospital Affiliated to Southern Medical University,Guangdong Guangzhou 510080,China;Department of Traditional Chinese Medicine,Guangzhou University of Chinese Medicine,Guangdong Guangzhou 510006,China;Department of Traditional Chinese Medicine,Southern Medical University,Guangdong Guangzhou 510515,China)

机构地区：[1]华南理工大学医学院,广东广州510006 [2]南方医科大学附属广东省人民医院中医科,广东广州510080 [3]南方医科大学附属广东省人民医院药学部,广东广州510080 [4]广州中医药大学中医系,广东广州510006 [5]南方医科大学中医系,广东广州510515

出　　处：《安徽中医药大学学报》2025年第1期80-89,共10页Journal of Anhui University of Chinese Medicine

基　　金：广东省基础与应用基础研究基金项目(2022A1515011260);广东省中医药局科研项目(20221004)。

摘　　要：目的探讨健脾化瘀方治疗肝细胞癌的作用机制。方法将SD大鼠随机分为对照组,健脾化瘀方含药血清低、中、高剂量组,每组10只;健脾化瘀方低、中、高剂量组大鼠分别灌胃健脾化瘀方4.32、8.64、17.28 g/kg,对照组灌胃等容积生理盐水。观察健脾化瘀方含药血清对肝癌细胞株HepG2和Huh7中Ras相关结构域家族1亚型A(Ras association domain family 1 isoform A,RASSF1A)、p38丝裂原激活的蛋白激酶(p38 mitogen-activated protein kinase,p38 MAPK)、磷酸化p38 MAPK(p-p38 MAPK)、细胞凋亡率、细胞增殖相关蛋白(p21、cyclin D1)、细胞凋亡相关蛋白[半胱天冬酶(cysteinyl aspartate specific proteinase,caspase)-3、caspase-8、caspase-9]的影响。采用RASSF1A小干扰RNA(RASSF1A siRNA)干扰RASSF1A的表达,将细胞分为对照组、阴性血清组、含药血清组、含药血清+对照siRNA组、含药血清+RASSF1A siRNA组,通过CCK-8、EdU、TUNEL、Western blot实验验证健脾化瘀方是否通过RASSF1A发挥抑制肝癌细胞增殖、促进肝癌细胞凋亡的作用;采用p38 MAPK信号通路抑制剂SB203580抑制p38 MAPK信号通路,将细胞分为对照组、阴性血清组、含药血清组、含药血清+SB203580组,通过CCK8、EdU、TUNEL、Western blot实验验证健脾化瘀方是否通过p38 MAPK信号通路发挥抑制肝癌细胞增殖、促进肝癌细胞凋亡的作用。结果健脾化瘀方含药血清干预48 h后,RASSF1A mRNA和蛋白表达水平随着含药血清浓度的增加而升高,细胞凋亡率及p-p38、p21、caspase-3、caspase-8、caspase-9蛋白表达水平显著升高(P<0.05),细胞存活率及cyclin D1蛋白表达水平显著降低(P<0.05)。与含药血清组比较,含药血清+RASSF1A siRNA组细胞中RASSF1A、p-p38、p21、caspase-3、caspase-8、caspase-9蛋白表达水平和细胞凋亡率显著降低(P<0.05),细胞存活率及cyclin D1蛋白表达水平显著升高(P<0.05)。与含药血清组比较,含药血清+SB203580组细胞中p-p38、p21、caspase-3、cObjective To investigate the mechanism of action of Jianpi Huayu Decoction(JPD)in the treatment of hepatocellular carcinoma(HCC).Methods Sprague-Dawley rats were randomly divided into control group and low-,middle-,and high-dose JPD-containing serum groups,with 10 rats in each group.The rats in the low-,middle-,and high-dose JPD-containing serum groups were given JPD by gavage at a dose of 4.32,8.64,and 17.28 g/kg,respectively,and those in the control group were given an equal volume of normal saline by gavage.Related tests were conducted to observe the effect of JPD-containing serum on Ras-associated domain family 1 subtype A(RASSF1A),p38 mitogen-activated protein kinase(p38 MAPK),phosphorylated p38 MAPK(p-p38 MAPK),apoptosis rate,cell proliferation-related proteins(p21 and cyclin D1),and cell apoptosis-related proteins(caspase-3,caspase-8,and caspase-9)in HCC cells lines HepG2 and Huh7.RASSF1A small interfering RNA(RASSF1A siRNA)was used to interfere with the expression of RASSF1A,and the cells were divided into control group,negative serum group,drug-containing serum group,drug-containing serum+NC siRNA group,and drug-containing serum+RASSF1A siRNA group;CCK8 assay,EdU,TUNEL,and Western blot were used to investigate whether JPD could inhibit the proliferation of HCC cells and promote their apoptosis through RASSF1A.The p38 MAPK signaling pathway inhibitor SB203580 was used to inhibit the p38 MAPK signaling pathway,and the cells were divided into control group,negative serum group,drug-containing serum group,and drug-containing serum+SB203580 group;CCK8 assay,EdU,TUNEL,and Western blot were used to investigate whether JPD could inhibit the proliferation of HCC cells and promote their apoptosis through the p38 MAPK signaling pathway.Results After intervention with JPD-containing serum for 48 hours,the mRNA and protein expression levels of RASSF1A increased with the increase in the concentration of JPD-containing serum,and there were significant increases in cell apoptosis rate and the protein expression levels o

关 键 词：健脾化瘀方 肝细胞癌 RASSF1A MAPK 细胞增殖 细胞凋亡 

分 类 号：R285[医药卫生—中药学]