应用小鼠主动脉移植模型探讨天然抗体在慢性移植物血管病中发挥功效的研究

发布时间：2018-03-02 13:02

本文选题：移植物血管病　切入点：天然抗体　出处：《吉林大学》2017年博士论文　论文类型：学位论文

【摘要】：背景:T细胞靶向主流免疫抑制剂的应用已大大改善了器官移植的早期预后,然而远期移植物存活率却维持原样。主要原因是不可避免的慢性排斥反应。所有血管化实体移植器官最终都可能因为慢性移植物血管病而功能衰竭。以供体特异性抗体为主的抗体介导性排斥反应通常被主流观点认为与移植物血管病有关。我们系列研究的早期研究结果发现,虽然作为固有免疫系统的一部分,固有B细胞(小鼠的B1 B细胞)所分泌的天然抗体(Nab),独立于供体特异性抗体以外,影响了肾移植物的远期预后。为了直接揭示移植物血管病与Nab的相互影响,我们选用小鼠主动脉移植模型来探索Nabs在慢性移植物血管病发病机制中的作用。材料与方法:1、诱导内生性天然抗体:紫外线照射新鲜小鼠(C57BL/6)胸腺细胞悬液诱导凋亡后,尾静脉注射给受体小鼠(C57BL/6),重复4次,每次间隔一周,流式细胞分析证实小鼠血清天然抗体含量是否随之提高;2、建立小鼠主动脉移植模型:应用“套管”技术,用供体小鼠的胸主动脉置换受体小鼠的肾动脉下腹主动脉,用恰当的方法评估移植物血管病的严重程度;3、设立4个实验组分别为:(1)全错配(BALB/c→C57BL/6)/内生性高Nab,(2)全错配/基线水平Nab,(3)无错配(C57BL/6→C57BL/6)/内生性高Nab,(4)无错配/基线水平Nab。按照实验设计执行手术,术后6周获取移植物做组织学分析。结果:1、流式细胞分析确认小鼠血清Nab含量会随着注射次数和时间的推移而增长,并且可以维持到实验终点(4-6周),内生性Nab小鼠模型建立成功;2、“套管”法小鼠主动脉移植被证明是一种优于传统缝线血管吻合的手术方式,数字型变量内膜增生指数(I/M)、血管狭窄度(%)和血管中膜单位面积细胞数,可以准确评价移植物血管病的严重程度;3、在内生性Nab影响下的小鼠,接受异源(BALB/c→C57BL/6)主动脉移植后,移植物血管病的病程进展相比全错配/未诱导Nab组小鼠有大幅延缓(全错配/Nab组:I/M=0.96±0.13,Occlusion%=24.8±3.9%;全错配/未诱导Nab组I/M=0.13±0.04,Occlusion%=4.4±2.0%;PI/M=0.004,Pocclusion=0.0108),内膜完整度及管腔通畅程度几乎与同源(无错配)移植对照组没有区别(全错配/Nab组:I/M=0.13±0.04,Occlusion%=4.4±2.0%;无错配/未诱导Nab组:I/M=0.07±0.04,Occlusion%=5.8±0.2%;PI/M=0.3170,Pocclusion=0.6352)。结论:内生性Nab的存在延缓了同种异体移植物血管管腔狭窄的进程,同时没有影响受体自身免疫系统的功能。
[Abstract]:Background the early prognosis of organ transplantation has been greatly improved by the use of major immunosuppressants targeting at: t cells. However, the long-term graft survival rate remains the same. The main reason is the inevitable chronic rejection. All vascularized solid transplant organs may eventually fail due to chronic graft angiopathy. Antibody-mediated rejection is commonly thought to be associated with graft angiopathy. Early findings in our series of studies have shown that. Although as part of the innate immune system, natural antibodies secreted by innate B cells (B 1 B cells in mice) are independent of donor-specific antibodies. In order to reveal the interaction between graft angiopathy and Nab, We selected mouse aortic transplantation model to explore the role of Nabs in the pathogenesis of chronic graft angiopathy. Materials and methods: 1, induced endogenous natural antibody: ultraviolet irradiation of fresh mouse C57BL / 6 thymocyte suspension induced apoptosis. Caudal vein was injected into the recipient mice C57BL / 6 and repeated four times, with a one-week interval. Flow cytometry confirmed whether the content of natural antibodies in the serum of mice was increased or not, and a mouse aortic transplantation model was established: the "cannula" technique was used. The abdominal aorta of the recipient mice was replaced by thoracic aorta of donor mice. The severity of graft angiopathy was evaluated by the appropriate method. 鈫扖57BL / 6 / endogenous high Nablite 2) Total mismatch / baseline level Nabo 3) no mismatch C57BL / 6. 鈫扤o mismatch / baseline Nab. according to the experimental design, the grafts were obtained 6 weeks after operation for histological analysis. Results: 1, flow cytometry confirmed that the serum Nab content of mice would increase with the number and time of injection. And it can be maintained until the experimental end point of 4-6 weeks, and the endogenous Nab mouse model was established successfully. The "cannula" method has been proved to be a better surgical method than traditional suture vascular anastomosis. The index of intimal hyperplasia (I / M) and the number of cells per unit area of vascular media can be used to accurately evaluate the severity of graft angiopathy in mice under the influence of endogenous Nab. 鈫扖57BL / 6) after aorta transplantation, The progression of grafts angiopathy was significantly delayed than that in the total mismatched / uninduced Nab group (total mismatch / nab group: 0.96 卤0.13 Occlusion.24.8 卤3.9T; I / M = 0.13 卤0.04) in the Nab group, 4.4 卤2.0PIP / M0.004Pocclusion 0.0108A, the degree of intimal integrity and patency of the lumen was almost homologous (no mismatch). There was no difference in control group (total mismatch / nab group: I / M 0.13 卤0.04Occlusion = 4.4 卤2.0; no mismatch / uninduced Nab group: I / M 0.07 卤0.04Occlusion = 5.8 卤0.2T = 0.3170Pocclusion 0.6352). Conclusion: the existence of endogenous Nab can delay the process of allograft vascular stenosis. At the same time, it does not affect the function of the receptor autoimmune system.
【学位授予单位】：吉林大学
【学位级别】：博士
【学位授予年份】：2017
【分类号】：R543

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