本文選題：人巨細(xì)胞病毒 + 包膜糖蛋白B(gB)��； 參考：《浙江大學(xué)》2012年碩士論文

【摘要】：背景 人巨細(xì)胞病毒(Human Cytomegalovirus,HCMV)感染和潛伏激活是骨髓移植受體術(shù)后引起移植失敗及發(fā)病率和死亡率最主要的病原體之一。HCMV是胞內(nèi)病毒,HCMV在骨髓移植受體體內(nèi)的感染和激活主要由機(jī)體的特異性T細(xì)胞對(duì)其免疫應(yīng)答程度所決定；HCMV包膜糖蛋白B(gB)是HCMV編碼的蛋白中最具有抗原性的一個(gè),檢測(cè)骨髓移植受體對(duì)HCMV gB特異性CTLs表達(dá)水平對(duì)防治移植術(shù)后HCMV感染有重要價(jià)值。 方法和目的 選取2009.2-2010.10在我院行骨髓移植術(shù)受體16例,HLA-A*0201受體組9例,HLA-A*1101受體組7例,共收集224份全血標(biāo)本。采用免疫組化法檢測(cè)外周血HCMV pp65抗原血癥,采用酶聯(lián)免疫吸附實(shí)驗(yàn)檢測(cè)術(shù)后8周和12周時(shí)外周血IFN-γ、IL-6分泌水平；采用五聚體結(jié)合流式細(xì)胞術(shù)測(cè)定HCMV gB特異性CTLs在術(shù)后8周和12周時(shí)體內(nèi)的表達(dá)水平,根據(jù)各個(gè)特異性五聚體在骨髓移植受體體內(nèi)的表達(dá)水平,進(jìn)一步篩選出gB優(yōu)勢(shì)抗原表位肽；同時(shí)分析HCMVgB特異性CTLs在骨髓移植受體體內(nèi)的表達(dá)水平與pp65抗原血癥及細(xì)胞因子水平的相關(guān)性,從而探析HCMV gB特異性CTLs在骨髓移植受體內(nèi)的免疫應(yīng)答狀況。 結(jié)果 1.HLA-A*0201受體組IFN-γ、IL-6分泌水平分別是12.43±7.60pg/ml、282.41±149.28pg/ml;HLA-A*1101受體組IFN-γ、IL-6分泌水平分別是16.36±6.55pg/ml、247.59±102.63pg/ml。IFN-γ、IL-6分泌水平在HLA-A*0201受體組和HLA-A*1101受體組間差異均無(wú)統(tǒng)計(jì)學(xué)意義(P0.05)； 2.HLA-A*0201受體組7條HLA-A*0201～gB抗原肽特異性CTLs在骨髓移植受體內(nèi)表達(dá)水平差異無(wú)統(tǒng)計(jì)學(xué)意義(P0.05);HLA-A*1101受體組10條HLA-A*1101～gB抗原肽中,HCMV gB527-535(AILSAIYNK)特異性CTLs在骨髓移植受體內(nèi)表達(dá)水平為1.26±0.86%,較其他HLA-A*1101～gB特異性CTLs表達(dá)水平差異有統(tǒng)計(jì)學(xué)意義(P0.05)； 3.外周血pp65抗原血癥與HLA-A*1101～gB特異性CTLs表達(dá)水平呈負(fù)相關(guān)(P=0.04),與HLA-A*0201～gB特異性CTLs表達(dá)水平無(wú)明顯相關(guān)性(P=0.209),與16例骨髓移植受體HTA～gB特異性CTLs表達(dá)水平呈負(fù)相關(guān)(P=0.003)；16例骨髓移植受體HTA～gB特異性CTLs表達(dá)水平與外周血IFN-γ分泌水平呈負(fù)相關(guān)(P=0.022),與IL-6分泌水平呈正相關(guān)(P=0.005)。 結(jié)論 1.建立MHC限制性的HCMV包膜糖蛋白B(gB)特異性CTLs測(cè)定技術(shù)； 2.在本研究選擇的23條gB抗原肽中,成功篩選出HLA-A*1101限制性HCMC gB527-535(AILSAIYNK)優(yōu)勢(shì)肽； 3.HCMV gB特異性CTLs可能在骨髓移植受體內(nèi)對(duì)HCMVpp65抗原血癥起抑制作用；IL-6可能促進(jìn)HCMVgB特異性CTLs的表達(dá)。
[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 鹵7.60 PG / ml 282.41 鹵149.28 PG / ml respectively. The level of IL-6 secretion was 16.36 鹵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 鹵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- 緯 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.
【學(xué)位授予單位】：浙江大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2012
【分類號(hào)】：R392

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