细胞毒T淋巴细胞相关抗原4免疫球蛋白抑制鼠高危角膜移植免疫排斥反应的实验研究

目的　探讨细胞毒T淋巴细胞相关抗原 4免疫球蛋白 (CTLA4 Ig)对小鼠高危角膜移植免疫排斥反应的抑制作用及局部抗免疫排斥反应机制。方法　建立 5 0只BALB/c小鼠穿透性角膜移植动物模型。治疗组 (2 5只 ) :取C5 7BL/ 6小鼠角膜片 ,放置于 10 μg/mlCTLA4 Ig保存液中浸泡 2 4h后 ,移植到BALB/c小鼠。对照组 (2 5只 ) :植片不行任何处理。术后每 3d用裂隙灯显微镜检查植片情况 ,每周应用组织学和免疫组织化学方法检测植片中各种炎性细胞和淋巴细胞的变化。对出现排斥反应的角膜植片应用逆转录PCR(RT PCR)方法检测部分细胞因子的表达。另外 ,选择经CTLA4 Ig治疗、植片保持透明 6周以上的小鼠作为受体 ,接受来自C5 7BL/ 6小鼠皮肤的移植 ,当移植皮肤发生排斥时 ,进行迟发性超敏反应 (DTH)分析。结果　CTLA4 Ig治疗组角膜植片保持透明 >10 0d。对照组小鼠术后 14d内均发生免疫排斥反应。组织病理学检查显示 ,角膜移植术后 2周 ,CTLA4 Ig治疗组植片保持正常细胞结构 ,无明显炎性细胞和T淋巴细胞浸润 ;对照组的排斥植片有大量炎性细胞和T淋巴细胞浸润 (包括CD 4 ,CD 8及CD 11细胞 )。术后 2周发生免疫排斥的角膜植片中检测到白细胞介素 10 (IL 10 ) ,肿瘤坏死因子α(TNF α) ,γ干扰素 (IFN γ) ,B7及C

CTLA4-Ig prevents corneal allograft rejection in mice

OBJECTIVE: Selective inhibition of T cell activation using the B7 specific fusion protein CTLA4-Ig has been shown to block CD28/B7 signal pathway. In this study, the capacity of soluble CTLA4-Ig alone in the suppressing corneal allograft rejection is further tested in mice, and its mechanism of preventing the cornea from immune rejection in local eye is analyzed. METHODS: The mouse models of corneal allografts were established. In one group (25 mice), the corneal donors from C57BL/6 mice were incubated in corneal storage medium containing 10 microg/ml of CTLA4-Ig for 24 hours, and then transplanted orthotopically into the recipients cornea of BALB/c mice. In another group (25 mice), as a control, no treatment was conducted on the donor corneas before surgery. The condition of the allografts was monitored using slit lamp microscopy every 3 days and the cellar architecture of selected graft was examined by histological and immunohistochemical techniques weekly. Local corneal cytokines expressing in the rejected grafts were examined. Some mice, which were treated with CTLA4-Ig and their grafts survived beyond 6 weeks, were selected as recipients of skin grafts from C57BL/6 donors. Delayed type hypersensitivity (DTH) was measured after the skin grafts rejection. RESULTS: All mice in the control group were rejected within 14 days after transplantation, while grafts incubated in CTLA4-Ig for 24 hours survived well beyond 100 days. The histopathology of surviving allografts had a normal cellular architecture, whereas the rejected allografts were heavily infiltrated with inflammatory cells and T lymphocytes (including CD4+, CD8+, and CD11+ cells). The cytokines including IL-10, TNF-alpha, IFN-gamma, B7-1 and CD40 were detected in the rejected corneas. Skin grafts from the C57BL/6 donor into the BALB/c mice after corneal transplantation were rejected with the expected median survival time of 11 days. DTH ear swelling challenge was induced after skin rejection, and a significant difference was found comparing with negative control mice. CONCLUSION: The results indicate that CTLA4-Ig could competitively combine with B7, block CD28/B7 T-cell signals, and inhibit immune responses.