人脐带来源间充质干细胞向HepG2肝癌细胞原位移植瘤的归巢及分化

发布时间：2018-03-01 10:26

本文关键词： 间充质干细胞肝癌归巢分化　出处：《中国肿瘤生物治疗杂志》2017年06期 　论文类型：期刊论文

【摘要】：目的:观察人脐带来源的间充质干细胞(human umbilical cord derived mesenchymal stem cell,HUMSC)向HepG2肝癌细胞原位移植瘤的归巢及分化。方法:二步法体外诱导HUMSC向肝细胞分化,通过Real-time PCR和Western blotting检测不同时间点HUMSC中肝细胞特异性基因甲胎蛋白(alpha-fetoprotein,AFP)及白蛋白(albumin,ALB)mRNA和蛋白水平的变化;Transwell实验观察HUMSC在HepG2肝癌细胞条件培养基作用下的趋化现象;采用皮下-原位移植的方法建立BALB/c裸鼠的HepG2肝癌细胞原位移植瘤模型,利用慢病毒感染的方法将HUMSC标记CMV或AFP启动子驱动的f Luc,制备成MSC.CMVLuc或MSC.AFPLuc,并将其注射入荷瘤肝组织,IVIS成像系统观察MSC.CMVLuc向肿瘤部位的迁移归巢及MSC.AFPLuc在肝癌微环境中向肝细胞分化。结果:HUMSC在体外诱导培养过程中出现细胞形态学变化证明其向肝细胞分化,同时诱导培养后AFP、ALB mRNA和蛋白的表达明显升高(均P0.05);HUMSC向HepG2肝癌细胞的趋化呈瘤细胞密度依赖性(P0.01);MSC.CMVLuc注射后2 d迁移并聚集于肝脏,注射后5 d肝脏内发光信号进一步增强;MSC.AFPLuc肝内注射后24 h已向肝细胞分化,注射第7天AFP启动子活性最强,注射第9天活性消失。结论:在小鼠体内外证明了HUMSC具有向肝脏归巢及分化的能力,为今后MSC应用于肝癌基因靶向治疗提供了一种新的策略。
[Abstract]:Objective: to observe the homing and differentiation of human umbilical cord derived mesenchymal stem cells derived from human umbilical cord into orthotopic transplantation of HepG2 hepatoma cells. Methods: two step method was used to induce HUMSC to differentiate into hepatocytes. Real-time PCR and Western blotting were used to detect the changes of the levels of alpha-fetoprotein (alpha-fetoprotein), alpha-fetoprotein (AFP) and albumin albumin (Alb) in HUMSC at different time points. The chemotaxis of HUMSC in the conditioned medium of HepG2 hepatocarcinoma cells was observed by Transwell experiment. The model of orthotopic transplantation of HepG2 hepatoma cells in BALB/c nude mice was established by subcutaneous orthotopic transplantation. Using lentivirus infection, HUMSC labeled CMV or f Luc-driven by AFP promoter was prepared into MSC.CMVLuc or MSC.AFPLuc. and injected into liver tissue of liver bearing tumor to observe the migration and homing of MSC.CMVLuc to tumor site and MSC.AFPLuc orientation in the microenvironment of liver cancer. Results the morphological changes of HUMSC in vitro showed that HUMSC differentiated into hepatocytes. At the same time, the expression of mRNA and protein of AFPG-ALB was significantly increased (both P0.05 and HUMSC were chemotaxis to HepG2 hepatoma cells in a density-dependent manner. P0.01P0.01MC.CMVLuc migrated and gathered in liver 2 days after injection. After 5 days of injection, the luminescence signal in the liver further enhanced the differentiation into hepatocytes 24 hours after injection of MSC.AFP Luc, and the activity of AFP promoter was the highest on the 7th day of injection. Conclusion: HUMSC has the ability to homing and differentiating into the liver in vivo and in vitro, which provides a new strategy for the future application of MSC in gene targeting therapy of liver cancer.
【作者单位】：中国医学科学院北京协和医学院血液病医院血液学研究所实验血液学国家重点实验室;天津医科大学基础医学院;
【基金】：国家自然科学基金资助项目(No.81400176,No.81402121) 天津市应用基础与前沿技术研究计划基金资助项目(No.14JCYBJC23700)~~
【分类号】：R735.7

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