Investigation of transplantation cell therapy for acute renal endothelial cell injury.

在日本，医疗保险的政府资源每年约为30亿日元。ESRD的人口为20万，每位患者每年的费用为500万日元。因此，目前每年用于ESRD的费用为1亿日元。考虑到进入ESRD的主要是2型糖尿病肾病，有必要开发控制进展性肾脏疾病的治疗方法。本项目首次探讨了内皮细胞移植治疗肾脏疾病的疗效。用抗肾小球内皮抗体超选择性注射肾动脉诱导胸腺裸大鼠血栓性微血管病变。内皮损伤在注射后立即开始，其部分恢复过程在3-4天后随着内皮细胞增殖开始。因此，在缺陷部位补充内皮细胞可以增加再生肾小球的数量，加速恢复过程，并保持肾功能。TMA起始3天后，通过左颈动脉导管向动物动脉注射人脐静脉内皮细胞（HUVE）。注射HJVEC一周后采血，取完整肾。24h后检测BUN升高，发现由于肾TMA所致，BUN升高有统计学意义。当HUVEC移植到TMAA肾脏时，这种增加减少。当注射大鼠肾小叶间动脉内皮细胞而不是HUVEC时，这种增加进一步降低。移植后1周仍可在肾小球丛中观察到注射细胞。电子显微镜也证实了这一点。采用活体CCD摄像机检测内皮移植的血流动力学效应，有效地阐明了缺血再灌注损伤时小管周围无血流现象（文献12）。内皮细胞移植后几分钟肾小球红细胞流量增加，最初为低流量。因此，内皮移植在TMA模型中的疗效部分来自于肾小球血流的改善。在此基础上，我们利用绿色荧光大鼠研究骨髓移植对TMA、缺血性急性肾功能衰竭、慢性间质性肾病、shc -肾功能衰竭、月牙状肾小球肾炎等多种肾脏疾病模型的疗效。这些项目目前正在进行中。在这个项目中，至少有一组报告了骨髓移植对缺血性急性肾功能衰竭的疗效，尽管我们可能在这短时间内发现病情恶化。因此，骨髓细胞的排泄因子被认为比移植骨髓细胞更为关键。在研究的因子中，G-CSF是与干细胞移植相关的有前景的因子之一，对缺血-再灌注损伤的恢复过程有效，其机制目前正在研究中。在这些细胞移植实验的基础上，我们阐明了羟基自由基清除剂对缺血再灌注损伤的作用，并于近期获得了论文的认可。该项目成功实施，并在研究期间产生了宝贵的观察结果。少

The governmental resource for medicare is about 3000 million yen per year in Japan. The population of ESRD is 200 thousand and the expense for each patient is 5 million yen per year. Thus 100 million yen is currently spent for ESRD in each year. Given the fact that the entry to ESRD is predominantly increasing from type 2 DM nephropathy, it is necessary to develop the therapeutic approach controlling the progressive renal disease.In this project, the efficacy of endothelial transplantation to renal disease was firstly investigated. The thrombotic micro-angiopathy (TMA) was induced to athymic nude rats injecting anti glomerular endothelial antibody super selectively to renal artery. The endothelial injury is initiated immediately after the injection, while its recovery process partly starts 3-4 days later as the endothelial proliferation. Therefore, the supplementation of endothelial cells to the defective sites may increase the number of regenerating glomeruli, accelerate the recovery … More process, and preserve renal function. Three days after the initiation of TMA, human umbilical venous endothelial cells (HUVE) were arterially injected to animals via catheter thru left jugular artery. One week after the injection of HJVEC, blood sampling was performed and the intact kidney was removed. After 24 h, the increase of BUN was examined and found the statistically significant increase because of the TMA kidney. This increase was reduced when HUVEC was transplanted to TMAA kidney. When rat renal interlobular arterial endothelial cells were injected instead of HUVEC, this increase was further decreased. The injected cells were observed at glomerular tuft even one week after the transplantation. It was also confirmed by electron microscopy. The hemodynamic effect of endothelial transplantation was examined using intravital video CCD camera which efficiently elucidated the peritubular no flow phenomena in ischemia reperfusion injury (ref 12). The glomerular red blood cell flow, initially low flow, was increased several minutes after the endothelial transplantation. Therefore the efficacy of endothelial transplantation in TMA model is partly derived from the improvement of glomerular blood flow.In parallel to these experiments, we developed green fluorescent rats to investigate the efficacy of bone marrow transplantation to several renal disease model, such as TMA, ischemic acute renal failure, chronic interstitial renal disease, SHC-renal failure, and crescentic glomerular nephritis. These projects are currently ongoing. During this project, at least one group reported the efficacy of bone marrow transplantation to ischemic acute renal failure though we could find deterioration during this short period. Therefore, the excretion factors from bone marrow cells were considered rather pivotal instead of transplanted bone marrow cells. Among the factors investigated, G-CSF is one of the promising factors related to stem cell transplantation, that was effective for the recovery process of ischemia-reperfusion injury, and that mechanism is currently investigating. In addition to these cell transplantation experiments, we elucidated the efficacy of hydroxyl radical scavenger to ischemia-reperfusion injury and got the acceptance of publication recently. This project was successfully conducted and generated valuable observations during the research periods. Less