摘要
目的了解TGF-β1刺激HSC后SB-525334对I型胶原分泌、miR-19b和miR-29b表达的影响。方法用MTT法检测TGF-β1与SB-525334干预LX-2细胞的最适药物浓度、最佳作用时间。LX-2细胞干预分为4组,对照组、TGF-β1干预组、SB-525334干预组、TGF-β1和SB-525334共干预组。通过蛋白质印迹法分析I型胶原蛋白的表达。反转录实时定量聚合酶链反应(RT-qRCR)分析各组miR-19b和miR-29b的表达。结果TGF-β1浓度为10 ng/mL时,HSC生存率最高,为132.0%。当SB-525334浓度为10μmol/l时,其抑制率最强,为38.4%。TGF-β1干预组LX-2细胞I型胶原蛋白的表达量(82.80±4.39)%较对照组(17.89±4.27)%显著增高(P<0.01),TGF-β1和SB-525334共干预组LX-2细胞I型胶原蛋白表达量为(62.62±3.72)%低于TGF-β1组(82.80±4.39)%(P<0.05)。RT-qPCR结果显示SB-525334可下调miR-19b的表达,TGF-β1干预组、SB-525334干预组、以及TGF-β1和SB-525334共干预组LX-2细胞miR-19b相对表达量分别为0.62±0.07、0.28±0.06、0.64±0.20,较对照组明显降低(P<0.01),TGF-β1和SB-525334共干预组LX-2细胞miR-19b的相对表达量较SB-525334组高(P<0.05),TGF-β1和SB-525334共干预组与TGF-β1干预组LX-2细胞miR-19b的相对表达量比较无明显差异(P>0.05)。SB-525334可下调miR-29b的表达,TGF-β1干预组、SB-525334干预组、以及TGF-β1和SB-525334共干预组LX-2细胞miR-29b相对表达量分别为0.77±0.05、0.44±0.04、0.61±0.06,较对照组降低(P<0.05),TGF-β1和SB-525334共干预组LX-2细胞miR-29b的相对表达量较TGF-β1干预组降低(P<0.05),TGF-β1和SB-525334共干预组LX-2细胞miR-29b的相对表达量较SB-525334干预组升高(P<0.05)。结论TGF-β1抑制剂SB-525334可抑制HSC分泌I型胶原,并下调其miR-19b和miR-29b的表达。
Objective The hepatic stellate cell line LX-2 was selected to study the effect of SB-525334 on the secretion of type I collagen and the expression of micro-ribonucleic acid(miR)-19b and miR-29b after stimulation by transforming growth factor beta1(TGF-β1),so as to evaluate the value of SB-525334 in the prevention and treatment of hepatic fibrosis.Methods Methyl thiazolyl tetrazolium(MTT)method was used to detect the optimal concentration and time of TGF-β1 and SB-525334 interfering with LX-2 cells.Then the LX-2 cells were divided into 4 groups,the control group,the TGF-β1 intervention group,the SB-525334 intervention group,and the co-intervention group.The expression of type I collagen,miR-19b and miR-29b was analyzed by western blot and reverse transcription real-time quantitative polymerase chain reaction(RT-qPCR).Results Hepatic stellate cell MTT test showed that the highest survival rate was 132.0%when the concentration of TGF-β1 was 10 ng/mL.When the concentration of SB-525334 was 10μmol/L,the inhibition rate was 38.4%.Western blot showed that SB-525334 inhibited type I collagen expression in LX-2 cells.The type I collagen expression of LX-2 cells in TGF-β1 intervention group was significantly higher than that in the control group([82.80±4.39]%vs[17.89±4.27]%,P<0.01).The type I collagen expression of LX-2 cells in co-intervention group was lower than that in TGF-β1 intervention group([62.62±3.72)%vs[82.80±4.39]%,P<0.05).RT-qPCR showed that SB-525334 downregulated miR-19b expression in LX-2 cells.The relative miR-19b expression of LX-2 cells in TGF-β1 intervention group,SB-525334 intervention group,and co-intervention group(0.62±0.07,0.28±0.06,0.64±0.20)was significantly lower than that in the control group(P<0.01).The relative miR-19b expression of LX-2 cells in co-intervention group was higher than that in SB-525334 group(P<0.05).There was no significant difference in the relative miR-19b expression of LX-2 cells between co-intervention and TGF-β1 intervention groups(P>0.05).Besides,SB-52533
作者
吴江华
马俊骥
郭昱
郭平
WU Jiang-hua;MA Jun-Ji;GUO Yu;GUO Ping(Department of Gastroenterology,the Second Hospital of Hebei Medical University,Hebei Key Laboratory of Gastroenterology,Hebei Institute of Gastroenterology,Shijiazhuang 050000,Hebei Province,China)
出处
《肝脏》
2019年第9期1011-1014,共4页
Chinese Hepatology
基金
国家自然科学基金项目(81200311)
河北省自然科学基金资助项目(H2015206431)
河北省医学重点科技研究计划项目(20130184)
