NITRIC OXIDE AND INSULIN IN ISLET TRANSPLANTATION

本研究的目的是提高临床胰岛移植的成功率。 移植 提出了两个相互关联的假设， 这个目标 首先，一氧化氮（NO）被假定为是一种主要的 胰岛功能障碍的介质。 第二，一个以前不认识的 胰岛素的能力，以减少NO的生产提出解释其 胰岛保护活性。 这两个假设都将得到检验 直接通过将本研究中的实验集中在使用 诱导型一氧化氮合酶“敲除”小鼠（iNOS-/-）。 具体 目的探讨巨噬细胞NO在胰岛移植中的作用 排斥反应 根据初步和已发表的结果， 在实验室中，提出NO以2个“波”抑制胰岛： 发生在同基因或同种异体胰岛移植后1-2天， 导致早期胰岛功能障碍。 第二波NO介导的 胰岛功能障碍被认为发生在经典的同种异体移植过程中， 排斥反应 早期波将通过比较同基因的 糖尿病iNOS-/-和iNOS-/+小鼠的胰岛功能;第二波 将在这些宿主中使用同种异体胰岛进行研究。 的抑制剂 然后，NO将用于确定如何最好地增加胰岛功能。 该实验室的初步和已发表的证据表明， 以临床适用的方式抑制NO确实增加了 胰岛移植 将胰岛植入腹膜内 由于移植物内NO的产生和 在该部位可以容易地跟踪同种异体移植物排斥反应。 小鼠 将是主要使用的物种，因为，像人类一样，他们是“低” NO的生产者;大鼠将用于选定的实验，因为 更大的尺寸。 具体目标II将通过以下方式确定机制： 胰岛素促进胰岛功能。 一个占主导地位的范例是 胰岛素可以让移植的β细胞“休息”。 然而，在这方面， 初步证据表明，糖尿病患者使用胰岛素 小鼠或大鼠降低巨噬细胞iNOS mRNA表达和NO 生产 因此，具体的实验将再次比较iNOS- /-和iNOS-/+小鼠直接测试胰岛- 胰岛素的保护活性来自于减少NO的产生。 最后，将采用高血糖钳夹来维持 高血糖症，以确定是否胰岛保护活性的 胰岛素依赖于其降低血糖的能力。 的结果 这项调查应该提供重要的新信息， 提高临床胰岛移植的成功率， NO在细胞排斥反应中的作用的基本信息 移植，以及如何通过胰岛素调节NO的产生。

The goal of this investigation is to increase the success of clinical islet transplantation. Two interrelated hypotheses are put forth to achieve this goal. First, Nitric Oxide (NO) is postulated to be a prime mediator of islet dysfunction. Second, a previously unrecognized ability of insulin to decrease NO production is proposed to explain its islet-protective activity. Both of these hypotheses will be tested directly by focusing the experiments in this investigation on the use of inducible Nitric Oxide Synthase "knockout" mice (iNOS-/-). Specific Aim I will determine the role of macrophage NO in islet graft rejection. Based on preliminary and published results from this laboratory, NO is proposed to inhibit islets in 2 "waves": an early wave that occurs 1-2 days posttransplant of syngeneic or allogeneic islets is responsible for early islet dysfunction. A second wave NO-mediated islet dysfunction is proposed to occur during classical allograft rejection. The early wave will be investigated by comparing syngeneic islet function in diabetic iNOS-/- and iNOS-/+ mice; the second wave will be investigated using allogeneic islets in these hosts. Inhibitors of NO will then be used to determine how to best increase islet function. Preliminary and published evidence from this laboratory suggests that inhibiting NO in a clinically applicable way does increase the success of islet transplantation. Islets will be implanted in the intraperitoneal cavity in this investigation because intragraft NO production and allograft rejection responses can readily be followed at this site. Mice will be the primary species used because, like humans, they are "low" producers of NO; rats will be used in selected experiments because of their larger size. Specific Aim II will determine the mechanism by which insulin promotes islet function. A dominant paradigm has been that insulin allows transplanted beta-cells to "rest". However, preliminary evidence indicates that insulin administration to diabetic mice or rats decreases macrophage iNOS mRNA expression and NO production. Therefore, specific experiments will again compare iNOS- /- and iNOS-/+ mice to directly test the hypothesis that the islet- protective activity of insulin results from decreasing NO production. Finally, hyperglycemic clamps will be employed to maintain hyperglycemia in order to determine if the islet-protective activity of insulin depends on its ability to lower blood glucose. The results of this investigation should provide important new information that will increase the success of clinical islet transplantation as well as providing basic information on what role NO plays in rejection of cell transplants, and how NO production is regulated by insulin.