骨髓移植受体术后HCMV gB糖蛋白特异性CTLs免疫应答研究

发布时间：2018-05-12 14:33

  本文选题：人巨细胞病毒 + 包膜糖蛋白B(gB)　； 参考：《浙江大学》2012年硕士论文

【摘要】：背景 人巨细胞病毒(Human Cytomegalovirus,HCMV)感染和潜伏激活是骨髓移植受体术后引起移植失败及发病率和死亡率最主要的病原体之一。HCMV是胞内病毒,HCMV在骨髓移植受体体内的感染和激活主要由机体的特异性T细胞对其免疫应答程度所决定；HCMV包膜糖蛋白B(gB)是HCMV编码的蛋白中最具有抗原性的一个,检测骨髓移植受体对HCMV gB特异性CTLs表达水平对防治移植术后HCMV感染有重要价值。 方法和目的 选取2009.2-2010.10在我院行骨髓移植术受体16例,HLA-A*0201受体组9例,HLA-A*1101受体组7例,共收集224份全血标本。采用免疫组化法检测外周血HCMV pp65抗原血症,采用酶联免疫吸附实验检测术后8周和12周时外周血IFN-γ、IL-6分泌水平；采用五聚体结合流式细胞术测定HCMV gB特异性CTLs在术后8周和12周时体内的表达水平,根据各个特异性五聚体在骨髓移植受体体内的表达水平,进一步筛选出gB优势抗原表位肽；同时分析HCMVgB特异性CTLs在骨髓移植受体体内的表达水平与pp65抗原血症及细胞因子水平的相关性,从而探析HCMV gB特异性CTLs在骨髓移植受体内的免疫应答状况。 结果 1.HLA-A*0201受体组IFN-γ、IL-6分泌水平分别是12.43±7.60pg/ml、282.41±149.28pg/ml;HLA-A*1101受体组IFN-γ、IL-6分泌水平分别是16.36±6.55pg/ml、247.59±102.63pg/ml。IFN-γ、IL-6分泌水平在HLA-A*0201受体组和HLA-A*1101受体组间差异均无统计学意义(P0.05)； 2.HLA-A*0201受体组7条HLA-A*0201～gB抗原肽特异性CTLs在骨髓移植受体内表达水平差异无统计学意义(P0.05);HLA-A*1101受体组10条HLA-A*1101～gB抗原肽中,HCMV gB527-535(AILSAIYNK)特异性CTLs在骨髓移植受体内表达水平为1.26±0.86%,较其他HLA-A*1101～gB特异性CTLs表达水平差异有统计学意义(P0.05)； 3.外周血pp65抗原血症与HLA-A*1101～gB特异性CTLs表达水平呈负相关(P=0.04),与HLA-A*0201～gB特异性CTLs表达水平无明显相关性(P=0.209),与16例骨髓移植受体HTA～gB特异性CTLs表达水平呈负相关(P=0.003)；16例骨髓移植受体HTA～gB特异性CTLs表达水平与外周血IFN-γ分泌水平呈负相关(P=0.022),与IL-6分泌水平呈正相关(P=0.005)。 结论 1.建立MHC限制性的HCMV包膜糖蛋白B(gB)特异性CTLs测定技术； 2.在本研究选择的23条gB抗原肽中,成功筛选出HLA-A*1101限制性HCMC gB527-535(AILSAIYNK)优势肽； 3.HCMV gB特异性CTLs可能在骨髓移植受体内对HCMVpp65抗原血症起抑制作用；IL-6可能促进HCMVgB特异性CTLs的表达。
[Abstract]:Background Human cytomegalovirus (HCMV) infection and latent activation is one of the most important pathogens causing transplant failure, morbidity and mortality after bone marrow transplantation. HCMV is the infection and activation of HCMV in bone marrow transplant recipients. The most antigenicity of the protein encoded by HCMV is determined by the immune response of specific T cells to it. Detecting the expression of HCMV GB specific CTLs in bone marrow transplant recipients has important value in preventing and treating HCMV infection after transplantation. Methods and purposes A total of 224 whole blood samples were collected from 16 cases of bone marrow transplant recipients in our hospital from September 2 to October 2010.9 cases of HLA-An0201 receptor group 9 cases of HLA-Af1101 receptor group were collected. The HCMV pp65 antigenemia in peripheral blood was detected by immunohistochemical method, and the levels of IL-6 in peripheral blood at 8 and 12 weeks after operation were detected by enzyme linked immunosorbent assay (Elisa). The expression of HCMV GB specific CTLs in vivo was determined by pentamer combined with flow cytometry at 8 and 12 weeks after operation. According to the expression level of each specific tetramer in bone marrow transplant recipients, the dominant epitope peptide of GB antigen was further screened. At the same time, the relationship between the expression of HCMVgB specific CTLs and pp65 antigenemia and cytokine levels in bone marrow transplant recipients was analyzed, and the immune response of HCMV GB specific CTLs in bone marrow transplant recipients was analyzed. Result The level of IL-6 secretion in 1.HLA-A*0201 receptor group was 12.43 卤7.60 PG / ml 282.41 卤149.28 PG / ml respectively. The level of IL-6 secretion was 16.36 卤6.55 PG / ml, respectively. There was no significant difference between HLA-A*0201 receptor group and HLA-A*1101 receptor group in the level of IL-6 secretion. There was no significant difference in the expression level of 7 HLA-A*0201~gB antigenic peptide specific CTLs in bone marrow transplant recipients in 2.HLA-A*0201 receptor group. The expression level of CTLs specific CTLs in bone marrow transplant recipients was 1.26 卤0.86, which was higher than that in other HLA-A*1101~gB receptor groups. The difference of heterosexual CTLs expression was statistically significant (P 0.05). 3. There was a negative correlation between peripheral blood pp65 antigenemia and HLA-A*1101~gB specific CTLs expression, no significant correlation with HLA-A*0201~gB specific CTLs expression, and a negative correlation with 16 bone marrow transplant receptor HTA~gB specific CTLs expression levels. The expression of HTA~gB specific CTLs was negatively correlated with the level of IFN- 纬 secretion in peripheral blood, and positively correlated with the level of IL-6 secretion. Conclusion 1. To establish a specific CTLs technique for the determination of HCMV envelope glycoprotein (HCMV glycoprotein) by MHC. 2. Out of 23 GB peptides selected in this study, HLA-A*1101 restricted HCMC (gB527-535AILSAIYNK) peptide was successfully screened. 3.HCMV GB specific CTLs may inhibit HCMVpp65 antigenemia in bone marrow transplant recipients. IL-6 may promote the expression of HCMVgB specific CTLs.
【学位授予单位】：浙江大学
【学位级别】：硕士
【学位授予年份】：2012
【分类号】：R392

【参考文献】

相关期刊论文 前2条

1 黑爱莲;李伟;刘娜;邓志辉;程良红;何军;靳伟民;杜丹;周晓阳;肖尧;戴大鹏;单小燕;张志欣;蔡剑平;;中华骨髓库造血干细胞捐献志愿者HLA-A、-B、-C、-DRB1、-DQB1高分辨等位基因频率分析[J];中国输血杂志;2009年04期

2 马伟杭,范骏,钱景,黄河,蔡挺,陈智;骨髓移植受者巨细胞病毒感染的研究[J];浙江预防医学;2001年09期

，

本文编号：1878983