赖氨大黄酸通过调控miRNA表达水平抑制HBV增殖  

RHL inhibiting proliferation of HBV in HepG2.2.15 cells through regulating the miRNA expression level

作　　者：张荣花[1] 王密斯 王琳[1] 冯雪研 郝晓方[1] 甄永占[1] 李琳[1] 陈静[1] 章广玲[1,2] 

机构地区：[1]华北理工大学基础医学院病原生物学与免疫学教研室,河北唐山063000 [2]河北省唐山市慢性病临床基础研究重点实验室,063000

出　　处：《检验医学与临床》2015年第19期2817-2819,2822,共4页Laboratory Medicine and Clinic

基　　金：国家自然科学青年基金资助项目(81201281);河北省自然科学基金资助项目(H2013209180;C2012401037;H2013209040);河北省唐山市科技计划资助项目(12140209A-4);华北理工大学研究生创新项目(2015S16;2015S17);国家级;省级大学生创新项目(201410081021)

摘　　要：目的研究赖氨大黄酸在体外对乙型肝炎病毒(HBV)增殖及病毒抗原表达的影响及分子机制,为研究赖氨大黄酸抗HBV作用机制奠定初步的实验依据。方法将HepG2.2.15细胞分为实验组(分别加入5、10、20、40、80μmol/L赖氨大黄酸)及不加赖氨大黄酸的对照组,分别培养24、48h收获上清液,采用四唑氮化合物(MTS)法检测细胞的增殖活性,采用ELISA法检测HBsAg和HBeAg的表达水平,实时定量PCR(RT-PCT)检测其中HBV DNA的拷贝数及miR-210、miR-185、miR-199a-3p、miR-370、miR-196a、miR-184、miR-217的表达水平。miR-210、miR-185、miR-199a-3p、miR-370、miR-196a、miR-184、miR-217反义链(ASO)分别转染HepG2.2.15细胞后,MTS法检测细胞的增殖活性,ELISA法检测HBsAg和HBeAg的表达水平。结果在没有对HepG2.2.15细胞的增殖活性产生影响的情况下,赖氨大黄酸抑制了HBsAg的产生和HBV的增殖。20μmol/L的赖氨大黄酸明显促进了miR-210、miR-185、miR-199a-3p的表达,而抑制了miR-370的表达,与对照组相比较,差异具有统计学意义(P<0.05)。结论赖氨大黄酸可能通过影响细胞内miRNA的表达抑制HepG2.2.15细胞中HBV的增殖和HBsAg的产生。Objective To study the effect and molecular mechanisms of rhein lysinate（RHL） on the prolifera‐tion of hepatitis B virus（HBV） and viral antigen expression to establish the preliminary experimental evidence for re‐searching the anti‐HBV mechanism of RHL .Methods HepG2 .2 .15 cells were divided into the experimental groups （adding 5 ,10 ,20 ,40 ,80 μmol/L RHL） and the control group without adding RHL .After culturing for 24 ,48 h ,the supernatant was obtained .The cell proliferation activity was detected by the non‐radioactive cell proliferation assay （M TS） ,the expression level of HBsAg and HBeAg were detected by adopting the ELISA method ,the HBV DNA copy number and the expression levels of miR‐210 ,miR‐185 ,miR‐199a‐3p ,miR‐370 ,miR‐196a ,miR‐184 and miR‐217 were detected by real time quantitative PCR（RT PCT ） .HepG2 .2 .15 was transfected by miR‐210 ,miR‐185 ,miR‐199a‐3p ,miR‐370 ,miR‐196a ,miR‐184 and miR‐217ASO ,the cell proliferation activity was detected by the M TS method ,the expression levels of HBsAg and HBeAg were detected by rthe enzyme linked immunosorbent assay （ELISA） ,respectively .Results Without generating the effect on the cell proliferation activity ,RHL inhibited the generation of HBsAg and the HBV proliferation .The expression level of miR‐210 ,miR‐185 and miR‐199a‐3p was in‐creased by RHL（20 μmol/L） and the expression level of miR‐370 was inhibited by RHL（20 μmol/L） ,the differences compared with the control group were statistically significant（P 〈 0 .05） .Conclusion RHL is possible to suppress HBV proliferation and HBsAg generation by affecting the miRNA expression in HepG2 .2 .15 cells .

关 键 词：赖氨大黄酸 乙型肝炎病毒 乙型肝炎病毒表面抗原 

分 类 号：R512.6[医药卫生—内科学]