Anti-RANTES联合环孢素A诱导小鼠二次心脏移植长期免疫耐受
发布时间：2018-12-10 08:38
【摘要】：目的探讨Anti-RANTES联合环孢素A在小鼠心脏二次移植急性排斥反应中的作用。方法构建小鼠心脏二次移植模型:初次移植,以Balb/c小鼠为供鼠,C57BL/6小鼠为受鼠,进行腹部心脏移植;二次移植,以同种Balb/c小鼠为供鼠,初次移植后存活的C57BL/6小鼠为受鼠,进行二次颈部心脏移植。然后经不同步骤处理,分为四组进行实验:对照组:初次腹部心脏移植存活小鼠,于初次腹部心脏移植2w后行二次颈部心脏移植,同时予腹腔注射与其他组同剂量的生理盐水处理;实验组A,初次腹部心脏移植存活小鼠,于初次腹部心脏移植2w后行心脏二次颈部移植,同时予腹腔注射与其他组同剂量的环孢素A处理(n=6);实验组B,初次腹部心脏移植存活小鼠,于初次腹部心脏移植2w后行心脏二次颈部移植,同时予腹腔注射与其他组同剂量的Anti-RANTES处理(n=6);实验组C,初次腹部心脏移植存活小鼠,于初次腹部心脏移植2w后行心脏二次颈部移植,同时予腹腔混合注射与其他组同剂量的环孢素A和Anti-RANTES处理(n=6)。观察四组二次移植心脏存活时间;HE染色组织病理学改变观察各组异位心脏急性排斥反应的程度;Q-PCR检测二次心脏移植物中RANTES、IFN-γ、IL-2、IL-10和TGF-β基因的相对表达量;ELISA检测RANTES、IFN-γ、IL-2、IL-10和TGF-β在受鼠血清中的浓度。结果在Anti-RANTES联合环孢素A实验组,移植心脏的存活时间可达8.58±0.24天,而在对照组、Anti-RANTES组和应用环孢素A组,移植心脏的存活时间分别为(3.1±0.43)、(5.2±0.26)和(5.58±0.20)天(P0.01);心脏移植物HE染色可见,在Anti-RANTES联合环孢素A实验组,其炎症细胞的浸润较对照组、单用Anti-RANTES实验组A、单环孢素A实验组B明显减少(P0.01);Q-PCR的结果提示,移植心脏内RANTES、IL-2、INF-γ的m RNA表达在Anti-RANTES联合环孢素A实验组较其他组明显减少,而移植心脏内IL-10、TGF-βm RNA的表达在Anti-RANTES联合环孢素A实验组较其他组明显增加(P0.01);ELISA的结果提示,移植心脏内RANTES、IL-2、INF-γ的血清浓度在Anti-RANTES联合环孢素A实验组较其他组明显减少,而移植心脏内IL-10、TGF-β血清浓度在Anti-RANTES联合环孢素A实验组较其他组明显增加(P0.01)。结论CC族趋化因子配体5(CCL5)RANTES,在小鼠二次心脏移植免疫排斥的诱发和发展过程中,起着至关重要的作用。单用可以Anti-RANTES或者环孢素A都可以有效抑制二次心脏移植的急性排斥反应,减轻炎症细胞向移植物聚集、减轻移植物中炎症细胞因子RANTES、IL-2、INF-γ的分泌而增加抗炎细胞因子IL-10、TGF-β的分泌,从而延长二次移植物的存活时间。但联合应用Anti-RANTES和环孢素A,以上抗急性免疫排斥效果更加明显,因而可以诱导小鼠对二次心脏移植较长期耐受。
[Abstract]:Objective to investigate the role of Anti-RANTES combined with cyclosporine A in acute rejection of second heart transplantation in mice. Methods the second heart transplantation model of mice was established. The abdominal heart transplantation was carried out with Balb/c mice as donor and C57BL/6 mice as recipient mice for the first time. The second transplantation was carried out with the allogeneic Balb/c mice as donor mice and the C57BL/6 mice survived after the first transplantation as recipient mice. Then, after different steps, they were divided into four groups: the control group: the first abdominal heart transplantation survival mice, after the first abdominal heart transplantation 2 weeks after the second cervical heart transplantation, At the same time, intraperitoneal injection of the same dose of normal saline treatment as other groups; Experimental group A, the first abdominal heart transplantation survival mice, after the first abdominal heart transplantation 2 weeks after the second cervical heart transplantation, at the same time intraperitoneal injection of the same dose of cyclosporine A treatment (nong6); The experimental group B, the first abdominal heart transplantation survival mice, after the first abdominal heart transplantation 2 weeks after the second cervical heart transplantation, at the same time intraperitoneal injection of the same dose of Anti-RANTES as other groups (nong6); Experimental group C, the first abdominal heart transplantation survival mice, after the first abdominal heart transplantation 2 weeks after the second cervical heart transplantation, at the same time intraperitoneal injection of the same dose of cyclosporine A and Anti-RANTES treatment (NN6). The survival time of the cardiac allograft in the four groups and the degree of acute rejection of ectopic heart in each group were observed by HE staining histopathological changes. Q-PCR was used to detect the relative expression of RANTES,IFN- 纬, IL-2,IL-10 and TGF- 尾 in secondary heart grafts, and ELISA was used to detect the concentrations of RANTES,IFN- 纬, IL-2,IL-10 and TGF- 尾 in the serum of recipient mice. Results the survival time of transplanted heart was 8.58 卤0.24 days in Anti-RANTES combined with cyclosporine A group, while in control group, Anti-RANTES group and cyclosporine A group, the survival time was (3.1 卤0.43). (5.2 卤0.26) and (5.58 卤0.20) days (P0.01); HE staining of cardiac grafts showed that the infiltration of inflammatory cells in Anti-RANTES combined with cyclosporine A group was significantly lower than that in control group. The number of inflammatory cells in Anti-RANTES group and single cyclosporine A group B was significantly lower than that in control group (P0.01). The results of Q-PCR showed that the expression of m RNA of RANTES,IL-2,INF- 纬 in the transplanted heart was significantly lower in the experimental group of Anti-RANTES combined with cyclosporine A than in the other groups, but IL-10, in the transplanted heart. The expression of TGF- 尾 m RNA in Anti-RANTES combined with cyclosporine A group was significantly higher than that in other groups (P0.01). The results of ELISA showed that the serum concentration of RANTES,IL-2,INF- 纬 in the transplanted heart was significantly lower in the experimental group of Anti-RANTES combined with cyclosporine A than that in the other groups, but IL-10, in the transplanted heart. The serum concentration of TGF- 尾 in Anti-RANTES combined with cyclosporine A group was significantly higher than that in other groups (P0.01). Conclusion CC chemokine ligand 5 (CCL5) RANTES, plays an important role in the induction and development of immune rejection after secondary heart transplantation in mice. Anti-RANTES or cyclosporine A alone can effectively inhibit the acute rejection of secondary heart transplantation, reduce the accumulation of inflammatory cells to the grafts, and alleviate the inflammatory cytokines RANTES,IL-2, in the grafts. The secretion of INF- 纬 increased the secretion of anti inflammatory cytokine IL-10,TGF- 尾, thus prolonging the survival time of the secondary graft. But the combination of Anti-RANTES and cyclosporine A had more obvious effect on acute immune rejection, so it could induce long-term tolerance to secondary heart transplantation in mice.
【学位授予单位】：福建医科大学
【学位级别】：硕士
【学位授予年份】：2015
【分类号】：R654.2
本文编号：2370296
[Abstract]:Objective to investigate the role of Anti-RANTES combined with cyclosporine A in acute rejection of second heart transplantation in mice. Methods the second heart transplantation model of mice was established. The abdominal heart transplantation was carried out with Balb/c mice as donor and C57BL/6 mice as recipient mice for the first time. The second transplantation was carried out with the allogeneic Balb/c mice as donor mice and the C57BL/6 mice survived after the first transplantation as recipient mice. Then, after different steps, they were divided into four groups: the control group: the first abdominal heart transplantation survival mice, after the first abdominal heart transplantation 2 weeks after the second cervical heart transplantation, At the same time, intraperitoneal injection of the same dose of normal saline treatment as other groups; Experimental group A, the first abdominal heart transplantation survival mice, after the first abdominal heart transplantation 2 weeks after the second cervical heart transplantation, at the same time intraperitoneal injection of the same dose of cyclosporine A treatment (nong6); The experimental group B, the first abdominal heart transplantation survival mice, after the first abdominal heart transplantation 2 weeks after the second cervical heart transplantation, at the same time intraperitoneal injection of the same dose of Anti-RANTES as other groups (nong6); Experimental group C, the first abdominal heart transplantation survival mice, after the first abdominal heart transplantation 2 weeks after the second cervical heart transplantation, at the same time intraperitoneal injection of the same dose of cyclosporine A and Anti-RANTES treatment (NN6). The survival time of the cardiac allograft in the four groups and the degree of acute rejection of ectopic heart in each group were observed by HE staining histopathological changes. Q-PCR was used to detect the relative expression of RANTES,IFN- 纬, IL-2,IL-10 and TGF- 尾 in secondary heart grafts, and ELISA was used to detect the concentrations of RANTES,IFN- 纬, IL-2,IL-10 and TGF- 尾 in the serum of recipient mice. Results the survival time of transplanted heart was 8.58 卤0.24 days in Anti-RANTES combined with cyclosporine A group, while in control group, Anti-RANTES group and cyclosporine A group, the survival time was (3.1 卤0.43). (5.2 卤0.26) and (5.58 卤0.20) days (P0.01); HE staining of cardiac grafts showed that the infiltration of inflammatory cells in Anti-RANTES combined with cyclosporine A group was significantly lower than that in control group. The number of inflammatory cells in Anti-RANTES group and single cyclosporine A group B was significantly lower than that in control group (P0.01). The results of Q-PCR showed that the expression of m RNA of RANTES,IL-2,INF- 纬 in the transplanted heart was significantly lower in the experimental group of Anti-RANTES combined with cyclosporine A than in the other groups, but IL-10, in the transplanted heart. The expression of TGF- 尾 m RNA in Anti-RANTES combined with cyclosporine A group was significantly higher than that in other groups (P0.01). The results of ELISA showed that the serum concentration of RANTES,IL-2,INF- 纬 in the transplanted heart was significantly lower in the experimental group of Anti-RANTES combined with cyclosporine A than that in the other groups, but IL-10, in the transplanted heart. The serum concentration of TGF- 尾 in Anti-RANTES combined with cyclosporine A group was significantly higher than that in other groups (P0.01). Conclusion CC chemokine ligand 5 (CCL5) RANTES, plays an important role in the induction and development of immune rejection after secondary heart transplantation in mice. Anti-RANTES or cyclosporine A alone can effectively inhibit the acute rejection of secondary heart transplantation, reduce the accumulation of inflammatory cells to the grafts, and alleviate the inflammatory cytokines RANTES,IL-2, in the grafts. The secretion of INF- 纬 increased the secretion of anti inflammatory cytokine IL-10,TGF- 尾, thus prolonging the survival time of the secondary graft. But the combination of Anti-RANTES and cyclosporine A had more obvious effect on acute immune rejection, so it could induce long-term tolerance to secondary heart transplantation in mice.
【学位授予单位】：福建医科大学
【学位级别】：硕士
【学位授予年份】：2015
【分类号】：R654.2
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1 罗增荣;Anti-RANTES联合环孢素A诱导小鼠二次心脏移植长期免疫耐受[D];福建医科大学;2015年
，本文编号：2370296
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