以脐带间充质干细胞为载体的靶向基因治疗系统联合5-FU对裸鼠HepG2移植瘤的作用

发布时间：2018-01-15 06:11

本文关键词：以脐带间充质干细胞为载体的靶向基因治疗系统联合5-FU对裸鼠HepG2移植瘤的作用　出处：《山东医药》2017年26期 　论文类型：期刊论文

【摘要】：目的构建以人脐带间充质干细胞(HUMSCs)为载体的靶向基因治疗系统,观察该系统联合低剂量5-FU对裸鼠HepG2移植瘤的治疗作用。方法采用组织块接种法从新鲜无菌新生儿脐带中分离HUMSCs,利用PCR、重叠PCR、酶切、连接等分子生物学技术构建慢病毒表达质粒LentiR.E1A和腺病毒穿梭质粒pAd-hTERTpIL24,并包装慢病毒LentiR.E1A和腺病毒Ad-hTERTp-IL24。采用Transwell实验观察慢病毒和腺病毒共感染对HUMSCs向HepG2细胞趋化的影响。采用皮下接种的方法建立HepG2移植瘤模型,将荷瘤小鼠分成5组各5只,分别给予尾静脉注射PBS、HUMSCs、低剂量5-FU、慢病毒感染HUMSCs联用低剂量5-FU、双病毒共感染HUMSCs联用低剂量5-FU的治疗,比较各组的抑瘤效应。利用BD Annexin-Ⅴ-PE凋亡试剂盒检测腺病毒Ad-hTERTp-IL24与低剂量5-FU联用诱导肝癌细胞凋亡情况。结果成功构建了慢病毒表达载体p LentiR.E1A和腺病毒表达载体pAd-hTERTp-IL24,并成功包装出了慢病毒LentiR.E1A和腺病毒Ad-hTERTp-IL24。Transwell实验显示,慢病毒和腺病毒共感染HUMSCs向肿瘤细胞的迁移能力无明显改变。联合低剂量5-FU治疗裸鼠肝移植瘤中治疗组肿瘤体积较对照组减小(P0.01)。Ad-hTERTp-IL24与低剂量5-FU联用诱导肿瘤微环境内的肿瘤细胞发生凋亡,其荧光强度明显强于Ad-Track+5-Fu组和5-Fu组。结论成功构建了以HUMSCs为载体的靶向基因治疗系统,由HUMSCS.LentiR.E1A运载携带抑瘤基因的腺病毒Ad-hTERTp-IL24联用低剂量5-FU对裸鼠HepG2移植瘤有明显抑制作用。
[Abstract]:Objective to construct a human umbilical cord mesenchymal stem cells (HUMSCs) treatment system as the carrier of targeted gene, to observe the therapeutic effect of the 5-FU system combined with low dose of HepG2 xenografts in nude mice. Methods using tissue explant method HUMSCs isolated from fresh sterile umbilical cord by PCR and overlap PCR, enzyme digestion, connection etc. molecular biological techniques to construct lentiviral plasmid LentiR.E1A and Adenovirus Shuttle Plasmid pAd-hTERTpIL24 expression, and packaging of lentivirus LentiR.E1A and adenovirus Ad-hTERTp-IL24. Transwell to observe the lentivirus and adenovirus infection of HUMSCs to HepG2 cell chemotactic effect. Methods by subcutaneous inoculation to establish HepG2 transplanted tumor model, the mice were divided into 5 groups the 5 rats were given intravenous injection of PBS, HUMSCs, 5-FU combined with low dose, low dose 5-FU HUMSCs lentivirus infection, two virus co infection of HUMSCs combined with low dose 5-FU treatment Treatment effect of tumor were compared. Combined with the induction of apoptosis of hepatoma cells by adenovirus Ad-hTERTp-IL24 detection BD Annexin- V -PE apoptosis kit with low dose of 5-FU. The successful construction of Lentivirus Expression vector p LentiR.E1A and adenovirus expression vector pAd-hTERTp-IL24, and successfully packaged display LentiR.E1A lentivirus and adenovirus Ad-hTERTp-IL24.Transwell experiment lentivirus and adenovirus, CO infection of HUMSCs to the migration of tumor cells had no obvious change. Combined with low dose 5-FU in the treatment of liver tumor in nude mice in the treatment group than the control group reduced tumor volume (P0.01) and.Ad-hTERTp-IL24 combined with low dose of 5-FU within the tumor microenvironment induced apoptosis of tumor cells, the fluorescence intensity was stronger than Ad-Track+5-Fu group and 5-Fu group. Conclusion the successful construction of the carrier of targeted gene therapy system based on HUMSCs, by HUMSCS.LentiR.E1A carrying articles The adenovirus Ad-hTERTp-IL24 with a tumor suppressor gene combined with low dose 5-FU has a significant inhibitory effect on HepG2 xenografts in nude mice.

【作者单位】：中国医学科学院血液病医院血液学研究所;西安交通大学第二附属医院;
【基金】：国家科技重大专项基金资助项目(2012ZX09102301-015)
【分类号】：R450;R735.7
【正文快照】： 肝癌由于其发病隐匿,患者被确诊时往往已达晚期并伴随肝功能不全,多数预后很差。目前,针对肝癌的传统治疗手段包括手术、化疗和放疗等,但疗效均欠佳。随着肿瘤分子生物学和现代细胞分子生物学技术的快速发展,基因治疗成为未来肿瘤治疗的新希望。间充质干细胞具有来源广泛、扩

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