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1.
BACKGROUND: Previous studies in our laboratory have demonstrated the importance of the thymus for rapid and stable tolerance induction in an allotransplant model. The focus of the present study was to explore the feasibility of autologous thymic transplantation to produce a new transplantable organ (thymokidney) and to examine the function of subsequent vascularized thymokidney transplants in T cell development. MATERIALS AND METHODS: Eight juvenile swine received autologous thymic grafts under the renal capsule. Thymic tissue was obtained through a partial (n=6) or complete (n=2) thymectomy, and growth of the autologous thymic graft was compared between partially and completely thymectomized animals. Two of the partially thymectomized animals received irradiated (1000 cGy) as well as non-irradiated autologous thymic grafts. Graft survival, growth and evidence of thymocyte development was determined by (a) macroscopic examination of the implanted tissue, (b) histological examination, and (c) flow cytometry. Naive CD4 SP T cells were identified by CD45RA-expression. RESULTS: Growth of transplanted thymic tissue was demonstrated in all thymic graft recipients. No difference was seen between partially and completely thymectomized animals. By POD 60, the thymic grafts exhibited normal macroscopic and microscopic structure, and normal thymocyte composition. Irradiated thymic tissue displayed a similar pattern of development, but growth was markedly delayed. To evaluate thymic function of the graft, a composite thymokidney was transplanted into a recipient which had previously been thymectomized, had few circulating CD4-single positive cells and had lost MLR reactivity. The number of CD4+/CD45RA+ cells in this animal increased steadily from POD 30 to POD 150, indicating that the thymus of the composite thymokidney allograft was functional; in addition, MLR assays demonstrated that the recipient recovered immunocompetence. CONCLUSIONS: The establishment of a thymokidney by thymic autografting to the renal subcapsular space results in normal thymic growth and function, and may provide a valuable tool for studying the role of the thymus in tolerance induction. As far as we are aware, we provide the first evidence of functional vascularized thymic graft reconstituting T cells and leading to a return of a immunocompetence in a large animal model.  相似文献   

2.
BACKGROUND: The potential of xenotransplantation for clinical application will require overcoming barriers of humoral and cellular rejection, through strategies using immune suppression or tolerance induction. This laboratory has previously reported the induction of tolerance in the discordant xenogeneic model of pig-to-rodent thymic transplantation. We also have described a miniature swine model of fully mismatched allogeneic composite vascularized thymokidney transplantation that induced transplantation tolerance. We tested a combination of these approaches in a clinically relevant pig-to-primate model of xenotransplantation. METHODS: Composite thymokidney grafts were prepared 40 to 80 days before transplantation by the autologous implantation of thymic tissue under the renal capsule of human decay-accelerating factor transgenic swine. Baboons received xenotransplants of both human decay-accelerating factor composite thymokidneys and omental implants of thymic tissue. Recipients were treated with an immunosuppressive-conditioning regimen including thymectomy or thymic irradiation, extracorporeal immunoadsorption of anti-alphaGal antibodies and T-cell depletion. Recipients were followed for indicators of xenograft rejection, T-cell depletion and reconstitution, anti-alphaGal antibody levels, and mixed lymphocyte responses. Immunologic responses were studied in those animals that survived for more than 3 weeks. RESULTS: Thymokidney xenografts survived for up to 30 days, with evidence of viable thymic epithelium and Hassall's corpuscles under the renal capsule and in the omental implants, and with evidence of few host lymphocytes. Three animals demonstrated donor-specific unresponsiveness, while maintaining normal alloresponses, in mixed-lymphocyte-response assays performed after immunosuppression had been stopped. Rejected grafts demonstrated humoral damage without evidence of cellular infiltrates. After graftectomy, one animal maintained donor-specific cellular unresponsiveness and stable anti-alphaGal antibody levels for more than 2 months. CONCLUSIONS: We concluded that composite thymokidney and thymic-tissue xenotransplantation from swine to baboons can induce donor-specific cellular unresponsiveness and stable anti-alphaGal antibody levels, suggesting avoidance of sensitization after xenotransplantation. The presence of viable donor-swine thymic epithelium could have a role in the development of donor-specific T-cell tolerance. Further strategies to address humoral rejection could prolong graft survival and result in long-term tolerance to xenografts.  相似文献   

3.
Xenogeneic thymus transplantation in a pig-to-baboon model   总被引:4,自引:0,他引:4  
BACKGROUND: We have tested whether fetal porcine thymic tissue transplantation can lead to tolerance across a discordant (pig-to-baboon) xenogeneic barrier. METHODS: Six baboons underwent a conditioning regimen with thymectomy, splenectomy, and anti-monkey CD3 antibody conjugated to a diphtheria toxin binding site mutant (FN18-CRM9). Porcine fetal or neonatal thymic tissue was transplanted into three baboons. Three control baboons received either no transplanted pig tissue (n=1) or adult pig lymph node (n=2). Cellular responses and skin xenografts were used to test for tolerance. RESULTS: After T-cell depletion and thymic transplantation, recovery of thymus-dependent na?ve-type CD4 cells (CD4/CD45RA ) and in vitro xenogeneic hyporesponsiveness were observed. No sensitization of alpha-galactosyl antibody responses was observed. The thymic grafts survived up to 48 days. Porcine skin xenografts were performed in two of these animals with survival of 22 and 24 days. Only two of these animals were completely T-cell depleted, and both failed to recover thymus-dependent T cells (CD4/CD45RA ). In one animal, general in vitro hyporesponsiveness was observed, with subsequent death from infection. The second animal demonstrated delayed recovery of T cells and prolonged general hyporesponsiveness in vitro. Neither animal demonstrated prolongation of porcine skin grafts compared with allografts (both rejected by day 13). CONCLUSIONS: Porcine thymic tissue is able to induce xenogeneic hyporesponsiveness. More efficient thymic engraftment may allow this approach to induce xenograft tolerance.  相似文献   

4.
目的建立并利用人源化小鼠模型,研究组织工程材料的体内免疫原性。方法利用NOD/SCID小鼠(非肥胖型糖尿病鼠与重症联合免疫缺陷鼠反交鼠模型).通过腹腔注射人外周血单个核细胞(PBMC)(100×10^6)建立人源化小鼠,在人源化小鼠皮下植入异体人皮肤(阳性对照)、人体脂肪干细胞与脱钙骨构建的组织工程骨(实验组)、单纯脱钙骨材料(阴性对照)。在植入后1周、2周、3周、4周,取小鼠尾静脉血,流式细胞仪检测人CD4^+和CD8^+淋巴细胞比例。4周后,取小鼠的移植局部皮肤进行HE染色分析,同时检测脾细胞人CD4^+和CD8^+淋巴细胞比例。结果NOD/SCID小鼠腹腔注射人PBMC4周内,在外周血和脾中均可测到人CD4^+和CD8^+淋巴细胞。病理分析结果显示:在人源化小鼠皮下植入的组织工程材料及成体干细胞构建的组织工程骨组未见炎性细胞浸润;而单独植入人皮肤组可见明显的炎性细胞浸润。结论人源化小鼠可作为组织工程材料体内免疫原性的研究的良好模型。  相似文献   

5.
Effect of cyclosporin on the thymus and spleen in rats   总被引:1,自引:0,他引:1  
Cyclosporin is a potent immunosuppressive agent. When cyclosporin is administered at a dose of 5 mg/kg/day i.p. for 2 weeks and 8 weeks, 15 mg/kg/day i.p. for 2 weeks, and 30 mg/kg/day i.p. for 1 week to normal Fischer rats, all thymuses were atrophic and their weight and number of thymocytes decreased dose dependently. However, no change was observed in spleen size. Histologically, thymic medulla was severely atrophic and the cellularity of thymic cortex, and splenic periarterial sheath and splenic marginal zone were decreased. Thymic cortex was more atrophic in the high dose group. All components recovered to near normal morphology 2 to 3 weeks after drug withdrawal, but recovery of splenic change was delayed in many rats. A reduction of T lymphocytes, particularly, helper T lymphocytes was detected by avidin-biotin peroxidase complex method using the monoclonal antibodies, W3/13, W3/25, OX8 and OX6. Many of the rats administered cyclosporin developed infection. Because of these changes in the immune response, precaution must be taken to prevent infection.  相似文献   

6.
目的:探讨首次尸肾移植受者使用抗Tac单抗诱导治疗后外周血淋巴细胞CD分子的动态变化及其临床意义。方法:对首次尸肾移植的患者,在三联免疫抑制方案(激素+骁悉+新山的明)基础上,给予两剂抗Tac单抗诱导治疗,其中舒莱组30例,赛呢哌组28例。以流式细胞仪检测术前,术后第1天、1周、2周、4周、6周及8周外周血淋巴细胞CD分子的表达。结果:移植受者外周血淋巴细胞CD分子变化明显,其中CD25在术后明显下降(P<0.05),并维持4~6周;CD40亦明显下降(P<0.05),术后2周时开始回升;CD86、CD28、CD80、CD95等均有下降,但各组间差异无统计学意义。舒莱组和赛呢哌组CD分子表达差异亦无统计学意义。结论:抗Tac单抗可以有效地封闭外周血活化淋巴细胞表面的CD25,抑制其增殖和分化;通过某种机制,抗Tac单抗可以降低外周血淋巴细胞CD40的表达,可能起到抑制B淋巴细胞活化的作用。  相似文献   

7.
BACKGROUND: Interferon-gamma, produced by T-helper cells, activates macrophages and increases expression of major histocompatibility complex (MHC) products in acute and chronic rejection. We investigated the role of interferon-gamma in murine heterotopic tracheal allografts. METHODS: Tracheas from BALB/c mice were heterotopically transplanted to BALB/c (12 isografts: 2 weeks [n = 6] and 4 weeks [n = 6], C57BL/6 (12 allografts: 2 weeks [n = 6] and 4 weeks [n = 6]) and C57BL/6 interferon-gamma knockout mice (12 interferon-gamma knockout allografts: 2 weeks [n = 4] and 4 weeks [n = 8]). BALB/c interferon-gamma knockout tracheas were transplanted to C57BL/6 mice (reverse knockout: 4 weeks [n = 6]) and BALB/c interferon-gamma knockout mice (4 weeks [n = 2]). C57BL/6 tracheas were transplanted to Bm12 mice (MHC Class II mismatch allografts: 4 weeks [n = 6]). Conventional histology and immunohistochemistry for CD4, CD8 and CD11b were performed. RESULTS: Minimal (<20%) obliteration was seen at 2 weeks in the allograft groups. No obliteration was seen in the isograft groups. However, all allografts were completely obliterated at 4 weeks. Interferon-gamma knockout allograft combinations displayed severe rejection characterized by intense intra- and extraluminal infiltration by CD4-, CD8- and CD11b-labeled cells. The MHC Class II mismatch allograft group showed normal epithelium and mild sub-epithelial infiltration by CD4+ cells at 4 weeks (CD8-, CD11b-). CONCLUSIONS: Absence of interferon-gamma does not protect the allograft from obliteration. Epithelial destruction by cytotoxic T cells appears to be an important mechanism in the development of obliteration in murine heterotopic tracheal allografts.  相似文献   

8.
BACKGROUND: Whereas clinical pancreatic transplantation has been highly successful in correcting the hyperglycemia of insulin-dependent diabetes mellitus (type 1), the results of islet transplantation have been disappointing. This discrepancy may be because of, at least in part, nonspecific loss of islets during the time required for revascularization. To test this hypothesis, we have designed composite kidney grafts containing vascularized autologous islets that can be used to compare the engraftment potential of vascularized versus nonvascularized islet tissue. METHODS: (1) Islet-cell isolation: miniature swine underwent either partial pancreatectomy to isolate autologous islets or total pancreatectomy to isolate minor antigen-mismatched islets. Islets were purified from excised pancreatic tissue by enzymatic digestion and discontinuous density gradient purification. Isolated islets were cultured for 3 days before transplant. (2) Creation of vascularized islet kidneys (IK): autologous islets alone (n=6), minor-mismatched islets alone (n=3), and minor-mismatched islets plus simultaneous autologous thymic tissue (n=3) were transplanted beneath the renal capsule of juvenile miniature swine. Minor antigen-mismatched islets were also transplanted into both the vascularized thymic graft of a thymokidney (to produce a thymo-islet kidney [TIK]) and the contralateral native kidney (n=3) and both the host thymus and beneath the renal capsule (n=2). All recipients receiving minor-mismatched islets were treated with a 12-day intravenous (IV) course of either cyclosporine A (CsA) at 10 mg/kg per day or FK506 at 0.15 mg/kg per day. (3) Assessment of Function: to evaluate the function of the transplanted islets, three animals bearing TIK and IK underwent total pancreatectomy 3 months following islet transplantation. RESULTS: (1) Islet-cell yields: an average of 254,960+/-51,879 (4,452+/-932 islet equivalents [IEQ]/gram of pancreas) and 374,410+/-9,548 (4,183+/-721 IEQ/gram of pancreas) viable islets were obtained by partial pancreatectomy and complete pancreatectomy, respectively. (2) Creation of IK: autologous islets engrafted indefinitely, whereas recipients of minor-mismatched islets alone rejected the islets within 2 months. However, when minor-mismatched islets were implanted into both the thymokidney and the contralateral kidney of animals bearing a thymokidney, the islets engrafted indefinitely in both sites (>3 months). Simultaneous implantation of islets into the host thymus and under the renal capsule also led to permanent engraftment of minor-mismatched islets. (3) Function of vascularized islets: three animals with both a TIK and an IK in place for 3 months underwent total pancreatectomy. All three animals maintained normoglycemia thereafter. In two of these animals, the IKs were removed 2 months after the pancreatectomy, and in both cases normoglycemia was maintained thereafter by the TIK. CONCLUSIONS: The implantation of islets beneath the autologous renal capsule permitted the establishment of a vascular supply and thereby supported normal islet-cell growth and function. The presence of thymic tissue beneath the autologous renal capsule facilitated the engraftment of minor-mismatched islets, and such grafts achieved results similar to autologous islet transplants. Therefore, the ability to create vascularized islet grafts may provide a strategy for successful islet transplantation across allogeneic and potentially across xenogeneic barriers.  相似文献   

9.
目的 检验CD103分子是否介导了CD8+T淋巴细胞对同种移植胰岛的免疫损伤.方法 用流式细胞仪检测野生型C57BL/6小鼠外周血CD8+T淋巴细胞表达CD103的情况.以Balb/c小鼠为供者,C57BL/6小鼠为受者,制作同种胰岛移植模型.受者分为3组:M290-SAP组小鼠注射CD103免疫毒素M290-SAP;M290组小鼠注射抗CD103单克隆抗体M290;另以仅接受胰岛移植、不注射任何药物的小鼠为未处理组.检测移植胰岛CD3、CD8、CD44和CD103阳性细胞的表达,检测肠系膜淋巴结中CD3、CD8和CD103阳性细胞的表达.移植物功能丧失或观察期结束时获取移植胰岛,行HE染色和免疫组织化学染色.结果 野生型C57BL/6小鼠外周血的CD8+T淋巴细胞中有44.06%表达CD103.未处理组移植胰岛浸润的细胞成分中有29%的CD8+T淋巴细胞表达CD103.M290-SAP组小鼠淋巴细胞不仅丧失了CD103的表达,而且CD8+T淋巴细胞的绝对数量也减少,该组小鼠血糖稳定时间超过100 d(未处理组为13 d,P<0.05),移植胰岛组织学形态良好.结论 CD8+T淋巴细胞免疫损伤同种移植胰岛必须表达CD103,CD103有可能成为胰岛移植抗排斥反应治疗的新靶点.
Abstract:
Objective To test whether the CD103 molecule mediates CD8+ T lymphocytes on allogeneic islet graft immune injury. Methods By using flow cytometry, the expression of CD103 in peripheral CD8+ T lymphocytes in wild-type C57BL/6 mice was detected. Allogenic islet transplantation models were made using Balb/c donor mice and C57BL/6 recipient mice. Recipients were divided into 3 groups: M290-SAP-treated mice were injected with CD103 immunotoxin M290-SAP; M290-treated mice were injected with CD103 monoclonal antibody M290; untreated mice were only transplanted islet without any drug treatment. CD3, CD8, CD44 and CD103 positive cells were counted in islet allograft infiltrative lymphocytes. CD3, CD8, and CD103 positive cells were measured in the mesenteric lymph node. The islet allografts were removed and subjected to HE staining and immunohistochemical staining at the time of graft loss or the end of the observation period. Results 44. 06% peripheral CD8+ T cells expressed CD103 in wild-type C57BL/6 mice. 29 % CD8+ T cells expressed CD103 in the infiltrative lyrnphocytes of islet allografts in the untreated mice. In M290-SAP-treated mice, the lymphocytes had no CD103 expression and the absolute number of CD8+ lymphocytes was decreased as well The blood glucose was maintained stable for more than 100 days (13 days in untreated group, P<0.05) in the M290-SAP-treated mice. Moreover, the transplanted islets retained intact. Conclusion CD103 expression is required for destruction of pancreatic islet allograft by CD8+ T cells. CD103 might provide a novel target for therapeutic intervention in islet allograft rejection.  相似文献   

10.
Chronic rejection, or cardiac allograft vasculopathy (CAV), remains the leading cause of late death in heart transplant recipients. The precise role and contributions of T lymphocyte subsets to CAV development remains unknown. METHODS: Donor hearts from B6.C-H2bm12 mice were transplanted into T lymphocyte subset knockout recipients and T lymphocyte-reconstituted nude recipients. No immunosuppression was used. Intimal proliferation was measured morphometrically. In vitro studies were performed to analyze the donor-specific activation status of recipient CD8+ lymphocytes by examining cellular proliferation, interleukin-2 secretion, and interleukin-2Ralpha expression. Intracellular cytokine staining assay was performed to determine both the profile and source of intragraft cytokines. RESULTS: Hearts transplanted into wild-type recipients developed severe CAV by 24 days. Intimal lesions were absent in the hearts that were transplanted into nude and CD4-/- knockout mice (containing CD8+ lymphocytes). In contrast, the donor hearts in CD8-/- knockout recipients (containing CD4+ lymphocytes) developed CAV, but significantly less than in wildtype. Adoptive transfer of T lymphocyte subset populations into nude recipients confirmed that CAV was absolutely contingent on CD4+ lymphocytes, and that CD8+ lymphocytes played an additive role in intimal lesion progression in the presence of CD4+ lymphocytes. Although CD8+ lymphocytes alone did not cause CAV in vivo, we demonstrated that MHC class II disparate alloantigens activated CD8+ lymphocytes both in vivo and in vitro. Finally, both CD4+ and CD8+ lymphocytes contributed to the intragraft IL-2 and IFN-gamma production. CONCLUSIONS: In this MHC class II mismatched murine model, CAV is a T lymphocyte dependent event, and absolutely contingent on the presence of CD4+ lymphocytes. Furthermore, CD8+ lymphocytes (1) are activated by MHC class II disparate antigens and (2) play a significant role in the progression of lesion development. Finally, both CD4+ and CD8+ lymphocytes contribute to CAV development via secretion of IFN-gamma, a known mediator of CAV in this model.  相似文献   

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