HLA—G在肾移植受者外周血外泌体中表达的初步研究  

HLA-G expression in peripheral blood of renal transplant recipients

作　　者：肖漓 黄海燕 毕丽丽 韩永 何秀云 许晓光 樊文梅 高钰 孔祥瑞 魏玉香 石炳毅 

机构地区：[1]解放军第三0九医院全军器官移植研究所移植研究所北京市器官移植与免疫调节重点实验室,北京100091

出　　处：《中华器官移植杂志》2015年第9期515-519,共5页Chinese Journal of Organ Transplantation

基　　金：国家自然科学基金（81571555）;国家高技术研究发展计划（2012AA021002）;首都卫生发展科研专项（2014-2-5071）

摘　　要：目的研究人豢白细胞抗原-G（HLA-G）在肾移植受者外周血外泌体中的表达情况。方法以2012年1月至2014年6月在单中心进行同种异体肾移植术且术后无排斥反应的12例肾移植受者（肾移植组）以及健康志愿者（对照组）5例为研究对象。每个对象收集5ml外周血并采用总外泌体提取试剂盒提取总外泌体。通过透射电子显微镜观察外泌体的大小及形态；流式细胞术检测血浆外泌体表面CD63、CD73、CD81和CD106的表达；蛋白印迹方法分析HLA-G在外泌体的表达情况。结果在肾移植组12例中9例以及对照组5例中2例的外周血血浆获得外泌体，可见大小不一、呈现类似圆形或椭圆形、直径为40-100nm的膜性微囊泡结构，透射电子显微镜下观察，符合外泌体形态和结构。将其标志分子CD63抗体包被磁珠实现捕获，肾移植组CD81阳性率为（78．57±12．35）％，CD106为（45．39±11．23）％，CD73为（63．37±15．62）％；对照组CD81阳性率为（35．37±8．42）％，CD106为（12．44±2．31）％，CD73为（10．23±2．57）％。肾移植组9例受者血浆外泌体均表达HLA-G;对照组5例仅中1例呈低表达。结论肾移植术后无排斥反应患者血浆外泌体高表达HLA-G，外泌体可能在移植后的免疫耐受中发挥着重要作用。Objective To-study the expression of human leucocyte antigen G （HLA-G） in peripheral blood exosomes of kidney transplantation recipients. Method Twelve patients received allogenic kidney transplantation in 309th Hospital of PLA from January 2012 to June 2014 （kidney transplantation recipient group） and 5 healthy volunteers （control group） were selected for the whole experiments. 5 mL peripheral blood was collected and total exosomes were extracted by exosome extraction kit （Life Technologies）. Transmission electron microscope （TEM） was used to observe the size and morphology of exosomes. Flow cytometry was used to detect CD63, CD73, CD81 and CD106 expression on the surface of plasma exosomes. Western blotting was used to analyze the expression of HLA-G in exosomes. Result Exosomes were obtained in 9 cases of kidney transplantation recipients （n= 12） and 2 cases of control group, and TEM results showed these membranous microvesicles resembled round or oval shape at different sizes and in diameter of 40-100 nm, which conformed to the morphology and architect of exosomes. The marker CD63 with antibody-coating immunomagnetic beads was used for capturing. CD81, CD106 and CD73 positive rate in kidney transplantation recipient group was （78. 57 ±12. 35） %, （45.39± 11.23） % and （63. 37 ± 15.62）% respectively, and that in control group was （35. 37 ± 8. 42）%, （12. 44 ± 2. 31）% and （10. 23±2. 57）% respectively. Plasma exosomes in all kidney transplantation recipients expressed HLA-G, while those in only 1 case of control group showed low expression of HLA-G. Conclusion HLA-G is expressed at high levels in the plasma exosomes of patients with non-rejection after renal transplantation and exosomes may play an important role in immune tolerance after transplantation.

关 键 词：HLA-G抗原 肾移植 

分 类 号：R699.2[医药卫生—泌尿科学]