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The study elucidating the molecular mechanisms underlying transplant vasculopathy

该研究阐明了移植血管病变的分子机制

基本信息

批准号：
08457349
负责人：
SHIRAKURA Ryota
金额：
$ 4.99万
依托单位：
Osaka University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (B)
财政年份：
1996
资助国家：
日本
起止时间：
1996 至 1997
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-08457349/
关键词：
rat heart graft retransplantation technique Tranplant vasculopathy PCR ラット SRY遺伝子 In Situ PCR法

项目摘要

To determine the origin of infiltrating cells in a retransplnted heart in our rat chronic rejection model, we have tried to establish the PCR in situ hybridization technique using a male specific gene, SRY,and a male dominant repetitive DNA sequence, RN91ES8, as targets. Despite the meticulous titration of fixation time, conditions of protease treatment, PCR conditions or the selection of ptimers and probes, it as not possible for us to detect a specific signal in the tissue. Then, we moved to the establishment of semi-quantitation of rat cytokine mRNA by RT-PCR techniques. We have now almost succeeded to quantitate about 40 different rat cytokine mRNAs expressed in transplanted heart. We are studying the molecular mechanisms underlying transplant vasculopathy in our rat heart-retransplantation model right now.Trying to establish the molecular techniques on the one hand, we have studied the pathophysiology of the disease using conventional techniques on the other. The studies listed in the publication table revealed that the critical immune responses within the first 5 days after transplantation determined the occurrence of transplant vasculopathy and showed that either CD4+ or CD8+ T cells and macrophages were essential for the critical immune responses. Moreover, the experiments retransplantting an allograft into a F1 animal or a nude rat indicated that the alloimmune responses by T cells after the initial period from the transplantation are dispensable for the progression of the disease. We are applying the molecular techniques established in this study on this unique model of transplant vasculopathy to elucidate the molecular mechanisms of the disease.

为探讨慢性排斥反应大鼠再移植心脏中浸润细胞的来源，我们尝试建立了以雄性特异基因SRY和雄性显性重复DNA序列RN 91 ES 8为靶基因的PCR原位杂交技术。尽管对固定时间、蛋白酶处理条件、PCR条件或ptimers和探针的选择进行了细致的滴定，但我们不可能检测到组织中的特异性信号。然后，我们转移到建立半定量的大鼠细胞因子mRNA的RT-PCR技术。我们现在几乎成功地定量了大约40种不同的大鼠细胞因子mRNA在移植心脏中的表达。我们目前正在大鼠心脏再移植模型中研究移植血管病变的分子机制，一方面试图建立分子技术，另一方面我们已经用常规技术研究了疾病的病理生理学。出版物表格中列出的研究表明，移植后前5天内的关键免疫应答决定了移植血管病变的发生，并表明CD 4+或CD 8 + T细胞和巨噬细胞对关键免疫应答至关重要。此外，将同种异体移植物再移植到F1动物或裸大鼠中的实验表明，在移植后的初始阶段之后，T细胞的同种免疫应答对于疾病的进展是不利的。我们正在将本研究中建立的分子技术应用于这种独特的移植血管病变模型，以阐明这种疾病的分子机制。