肝细胞生长因子转染的MSCs防治小鼠肠道移植物抗宿主病及其机理研究

发布时间：2018-01-12 23:13

本文关键词：肝细胞生长因子转染的MSCs防治小鼠肠道移植物抗宿主病及其机理研究　出处：《中国人民解放军军事医学科学院》2006年博士论文　论文类型：学位论文

【摘要】：异基因造血干细胞移植是治疗恶性血液病最有效的方法之一，但是，由于供受者之间主要组织相容性复合体的不同，移植物抗宿主病(GVHD)的发生难以完全避免。GVHD可以影响多个器官，主要是皮肤、肝脏和肠道。重度肠道GVHD会加重全身GVHD，影响移植的疗效，增加病死率。肠道粘膜损伤能够通过加重GVHD时的炎性细胞因子风暴，因此，粘膜损伤修复和免疫调节是肠道GVHD治疗的关键。肝细胞生长因子(HGF)对许多组织都具有修复作用，间充质干细胞(MSCs)不仅具有损伤修复作用，还具有免疫调节、促进造血恢复的作用，本实验建立小鼠异基因造血干细胞移植肠道GVHD模型；应用高表达HGF的MSCs(MSC-HGF)防治小鼠肠道GVHD，比较其与药物以及单独应用MSC防治GVHD之间的差异。材料和方法Balb／c小鼠经5.5Gy照射后输注C57BL／6小鼠骨髓细胞及脾细胞建立小鼠肠道GVHD模型；分离培养、鉴定C57BL／6小鼠骨髓MSCs，应用脂质体法将质粒pcDNA3.1-HGF瞬时转染入MSCs，检测转染细胞HGF表达情况；实验组根据细胞输注时间(0d，+6d，+10d)和输注形式(MSC或MSC-HGF)的不同分为6组：MSCs-0d，MSC-HGF-0d，MSCs-6d，MSC-HGF-6d，MSCs-10d，MSC-HGF-10d；输注MSCs细胞数量为1×10~6／只：空白对照组未予GVHD预防与治疗，药物对照组采用环胞菌素(CSA，5 mg／kg／d，灌胃)和骁悉(MMF，40 mg／kg／d，灌胃)联合防治GVHD。比较移植后肠道GVHD的发生率和致死率，肠道病理损伤情况以及小鼠存活情况。检测细胞输注后受体小鼠T细胞亚群变化，CD4+CD25+T细胞比例变化，，炎症相关细胞因子表达情况，以及肠道凋亡细胞变化。
[Abstract]:Allogeneic hematopoietic stem cell transplantation (HSCT) is one of the most effective methods for the treatment of malignant hematological diseases, but due to the difference of major histocompatibility complex among donors. Graft-versus-host disease (GVHD) is difficult to avoid completely. GVHD can affect many organs, mainly skin, liver and intestine. Severe intestinal GVHD can aggravate systemic GVHD. Intestinal mucosal injury can be induced by inflammatory cytokines storm in GVHD. Therefore, mucosal damage repair and immunomodulation are the key factors in the treatment of intestinal GVHD. Hepatocyte growth factor (HGF) has repair effect on many tissues, mesenchymal stem cells (MSCs) have not only damage repair, but also immune regulation, promote hematopoietic recovery. The GVHD model of allogeneic hematopoietic stem cell transplantation was established in this experiment. High expression of HGF MSC-HGF-HGFH was used to prevent and treat GVHD in mouse intestine, and the difference was compared with drug and MSC alone in the prevention and treatment of GVHD. Materials and methods Balb/c mice were irradiated with 5.5 Gy and then injected with C57BL / 6 mice bone marrow cells and spleen cells to establish the intestinal GVHD model of mice. C57BL / 6 mouse bone marrow MSCs were isolated and identified. The plasmid pcDNA3.1-HGF was transiently transfected into MSCs by liposome method. The expression of HGF in transfected cells was detected. The experimental group was divided into 6 groups according to the time of cell infusion (0 d, 6 d, 10 d) and the form of infusion (MSC or MSC-HGF) into 6 groups: MSC-HGF-0d. MSCs-6dT MSC-HGF-6dT; MSCs-10d; MSC-HGF-10d; The number of MSCs cells was 1 脳 10 ~ (-6) / mouse: the blank control group was not treated with GVHD, while the drug control group was treated with cyclosporine for 5 mg/kg/d. The incidence and mortality of intestinal GVHD after transplantation were compared with 40 mg / kg / d of mycophenolate mofetil and 40 mg / kg / d of mycophenolate mofetil. The changes of T cell subsets in recipient mice and the percentage of CD4 CD25 T cells and the expression of inflammatory cytokines were detected. And the changes of intestinal apoptotic cells.
【学位授予单位】：中国人民解放军军事医学科学院
【学位级别】：博士
【学位授予年份】：2006
【分类号】：R392

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