联合 OX40Ig 和 CTLA4Ig 基因转染对移植物排斥反应的影响及其机制。

Effect of combined OX40Ig and CTLA4Ig gene local transfer on allograft rejection and the underlying mechanisms.

机构信息

Department of Plastic Surgery, Xijing Hospital, Fourth Military Medical University, Xi'an, PR China.

出版信息

J Surg Res. 2012 Dec;178(2):949-58. doi: 10.1016/j.jss.2012.05.034. Epub 2012 May 31.

DOI:10.1016/j.jss.2012.05.034

PMID:22694937

Abstract

BACKGROUND

OX40Ig and CTLA4Ig fusion proteins have been suggested to induce immune tolerance and prevent rejection in allografts. The present study aims to investigate and compare the effects of ex vivo combined OX40Ig and CTLA4Ig lentivirus-mediated gene transfer on the long-term survival of the graft, as well as potential underlying mechanisms.

METHODS

We ex vivo transferred Brown Norway rats' superficial groin free flap with lentivirus vectors expressing OX40Ig or CTLA4Ig, or OX40Ig and CTLA4Ig combined, and transplanted the free flaps to Lewis rats. Short-course rapamycin was administered after transfection and transplantation. RT-PCR and Western blot were employed to evaluate expression of OX40Ig and CTLA4Ig. We assessed the survival time of the grafts and the degree of acute graft rejection after indicated treatment. Mixed lymphocyte reaction, flow cytometry, and ELISA were also used to evaluate systemic immune reactions.

RESULTS

Ex vivo transfer of OX40Ig or CTLA4Ig lentivirus vectors led to local expression of corresponding mRNA and proteins in the donor flap without affecting other organs of the recipient. The graft survival time was significantly expanded and rejection was markedly attenuated after transfection. Mixed lymphocyte reaction, flow cytometry (CD4(+) and CD8(+) T lymphocyte proportions), and serum ELISA analysis (IL-2, IFN-γ, IL-4, and IL-10) also showed decreased immune response following transfection. Combined OX40Ig and CTLA4Ig transfer exerted superior effect on improving graft survival and preventing graft rejection, inhibiting the immune response and decreasing the production of proinflammatory cytokines, compared with singular transfer of either OX40Ig or CTLA4Ig.

CONCLUSION

Combined ex vivo transfer of OX40Ig and CTLA4Ig lentivirus vectors provided superior benefits on long-term survival and restoration of the graft through inhibiting immune response and decreasing the production of proinflammatory cytokines.

摘要

背景

OX40Ig 和 CTLA4Ig 融合蛋白被认为可以诱导同种异体移植物中的免疫耐受和预防排斥反应。本研究旨在探讨和比较外源性 OX40Ig 和 CTLA4Ig 慢病毒介导的基因转移对移植物长期存活的影响，以及潜在的机制。

方法

我们用表达 OX40Ig 或 CTLA4Ig 或 OX40Ig 和 CTLA4Ig 联合的慢病毒载体对外源性转染布朗-挪威大鼠腹股沟浅游离皮瓣，并将游离皮瓣移植到 Lewis 大鼠。转染和移植后给予短期雷帕霉素治疗。采用 RT-PCR 和 Western blot 检测 OX40Ig 和 CTLA4Ig 的表达。评估移植后移植物的存活时间和急性排斥反应的程度。混合淋巴细胞反应、流式细胞术和 ELISA 也用于评估全身免疫反应。

结果

OX40Ig 或 CTLA4Ig 慢病毒载体的外源性转染导致供体皮瓣局部表达相应的 mRNA 和蛋白，而不影响受体的其他器官。转染后移植物存活时间明显延长，排斥反应明显减轻。混合淋巴细胞反应、流式细胞术（CD4(+)和 CD8(+)T 淋巴细胞比例）和血清 ELISA 分析（IL-2、IFN-γ、IL-4 和 IL-10）也显示转染后免疫反应降低。与单独转染 OX40Ig 或 CTLA4Ig 相比，联合转染 OX40Ig 和 CTLA4Ig 对改善移植物存活和预防移植物排斥反应、抑制免疫反应和减少促炎细胞因子的产生具有更好的效果。