摘要
目的:研究采用人脐带间充质干细胞条件培养基(MSC-CM)治疗肝细胞氧化应激损伤中微小RNA(miRNA)的差异性表达并探讨其调控机制。方法:制备人正常肝细胞株L02氧化应激损伤模型,体外分离培养健康人脐带间充质干细胞并制备MSC-CM。损伤模型经MSC-CM治疗,用凋亡、细胞活力、细胞周期及线粒体膜电位等实验验证其治疗效果。定量RT-q PCR筛选差异性表达miRNA。应用生物信息学预测其靶蛋白后使用Western blot实验验证。结果:MSC-CM治疗可以显著减少H2O2氧化应激损伤所致的细胞凋亡比率,增加细胞活力,调节细胞周期。经RT-q PCR筛选出显著差异表达的miR-143、miR-145、miR-301a与let-7a,均在损伤后表达升高,而在MSC-CM作用后表达降低。Western blot实验结果显示miR-143的预测靶蛋白HK2和ADRB1的表达在肝细胞损伤后降低,MSC-CM作用后升高,与miR-143调控趋势一致。结论:MSC-CM治疗可能通过调节相关miRNA及其靶蛋白逆转H_2O_2所致肝细胞氧化应激损伤。
AIM： To evaluate the changes of microRNA（ miRNA） in hepatocytes during hydrogen peroxideinduced oxidative stress injury,and to observe the alleviating effect of mesenchymal stem cell-conditioned medium（ MSCCM） in this progress. METHODS： The hepatocyte oxidative stress injury model was established using hydrogen peroxide and human normal liver cell line L02. MSC-CM was prepared using centrifugation and filter. The effects of MSC-CM on hepatocyte injury were evaluated by apoptosis analysis,cell viability detection,cell cycle,and mitochondrial membrane potential（ MMP）. Twenty-one differentially expressed miRNAs were selected by gene chip hybridization,in which miR-143,miR-145,miR-301 a and let-7a were confirmed by RT-q PCR. Bioinformatics software was utilized to predict target proteins of these miRNAs,and then the proteins were verified by Western blot. RESULTS： MSC-CM markedly attenuated hydrogen peroxide-induced oxidative stress injury by reducing apoptosis,promoting cell viability and regulating cell cycle. The expression of miR-143,miR-145,miR-301 a and let-7a,indentified by RT-q PCR,increased under the condition of oxidative stress injury,while decreased after MSC-CM treatment. The expression of miR-143 predicted target proteins,HK2 and ADRB1,decreased under the hydrogen peroxide-exposure,while increased after MSC-CM treatment,which is consistent with the regulatory trend of miR-143. CONCLUSION： MSC-CM might attenuate hydrogen peroxide induced oxidative stress injury via inhibiting apoptosis and regulating some miRNA expression.
出处
《中国病理生理杂志》
CAS
CSCD
北大核心
2016年第9期1670-1676,共7页
Chinese Journal of Pathophysiology
基金
深圳市科技研发基金知识创新计划(No.JCYJ20140418115449178)
山东省科技发展计划(No.2014GSF118131&2013GSF11812)
山东大学基本科研业务费资助项目(No.2014QLKY02&2014QY003-11)
