Research about mechanism of delayd xenograft rejection (DXR).

目的：不协调异种移植发生延迟异种移植排斥反应（DXR），补体活化、天然抗体和凝血被CVF （cobra venom factor）等药物抑制。虽然DXR的心脏病理显示NK（自然杀伤细胞）和巨噬细胞浸润，但这些细胞如何诱导DXR的机制尚不清楚。在这项研究中，我们利用来自DA大鼠（DA）的先天性NK活性缺乏大鼠，Beige大鼠（Bg）来研究NK细胞对DXR的贡献。材料与方法：以Yac-1细胞为靶细胞，DA和Bg的脾细胞为效应细胞，测定NK活性（^<51>Cr释放法）。用流式细胞术比较DA和Bg两种细胞表面抗原的表达。采用RT-PCR方法检测DA和Bg脾细胞中穿孔素、颗粒酶A、颗粒酶B等效应分子的表达。CVF对补体抑制作用（1,0天）。30 u /公斤。iv.)，将豚鼠心脏移植到DA和Bg，并比较其移植物的存活时间。结果:1。与DA相比，BG中NK活性严重受损。（在E/T=100时，DA: 62.8%, BG: 18.2%）DA组与BG组NK细胞数量无明显差异，而BG组CD8阳性和IL2R阳性细胞数量较DA组减少。与da相比，Bg中穿孔素、颗粒酶A和颗粒酶B mRNA的表达受到抑制。Bg组移植物平均存活时间（28(]SY.+-[)1.5小时，n=9）明显长于DA组（20(]SY.+-[)2.1小时，n=9）。结论：NK细胞和CD8阳性细胞在DXR过程中起重要作用。因此，抑制这些细胞的功能可以延长异种移植物的存活时间。

Purpose : Delayd xenograft rejection (DXR) takes place in discordant xenotransplantation, even complement activation, natural antibody and blood coagulation are inhibited by drugs such as CVF (cobra venom factor). Although pathology of the heart underlying DXR shows infiltration of NK (natural killer) cells and macrophages, the mechanism of how these cells can induce DXR is unknown. In this study, we addressed the contribution of NK cells to DXR using congenitally NK activity deficient rat, Beige rat (Bg) that is derived from DA rat (DA).Materials and methods :1. NK activity (^<51>Cr release assay) was measured using Yac-1 cells as targets and splenocytes of DA and Bg as effector cells.2. Expression of cell surface antigens in DA and Bg splenocytes was compared by flowcytemetry.3. Using RT-PCR,the expression of effector molecules such as perforin, granzyme A and granzyme B was detected in DA and Bg splenocytes.4. Under complement inhibition by CVF (-1,0 day. 30U/kg. iv.), guinea pig hearts were transplanted into DA and Bg, and their grafts survival time were compared.Result :1. NK activity in BG was severely impaired in comparison with that in DA.(DA : 62.8%, BG : 18.2% at E/T=100)2. There was no difference in NK cells number between DA and BG,whereas cell numbers of CD8 positive and IL2R positive cells were decreased in Bg as compared with those in DA.3. The expressions of perforin, granzyme A and granzyme B mRNA were inhibited in Bg as compared with those in DA.4. Grafts mean survival time in Bg (28(]SY.+-。[)1.5hr, n=9) was significantly longer than that in DA (20(]SY.+-。[)2.1hr, n=9).Conclusion : NK cells and CD8 positive cells play an important role in the process of DXR.Therefore, the inhibition of these cells function would lead to longer xenografts survival.