本文選題：尖銳濕疣 + 復發(fā)　； 參考：《天津醫(yī)科大學》2011年碩士論文

【摘要】：尖銳濕疣(condyloma acuminatum, CA)是由人乳頭瘤病毒(human papillomavirus, HPV)引起的,以發(fā)生在生殖器、肛周為主的性傳播疾病(sexually transmitted disease, STD)。CA不僅發(fā)病率高,而且治療后其復發(fā)率也很高。目前有多種針對CA的治療方法,但任何一種方法都不能避免其復發(fā)。不同型的HPV感染可以引起不同的臨床表現(xiàn),對HPV進行分型將有利于臨床疾病的診斷及預后的判定。 整合素(integrin)是細胞黏附分子家族中重要的成員之一。通過大量對整合素與病毒感染關系的研究發(fā)現(xiàn),多種病毒都能利用整合素作為病毒受體或共受體(co-receptor)進入宿主細胞。整合素α6主要在正常上皮組織基底層特異性表達,而HPV對宿主造成的損害,首先是以感染宿主上皮組織基底層為起始的,二者是否存在相關性?整合素α6在HPV感染過程中是否發(fā)揮作用?如果能在這個起始階段將整合素的表達進行封閉,是否可使病毒對宿主細胞的黏附受到抑制?對這些問題的進一步研究將有助于從根本上阻斷HPV病毒的感染和CA的復發(fā)。 目的①鑒定和評估PCR聯(lián)合限制性片段長度多態(tài)性(RFLP)及基因測序技術對CA及四種外生殖器和肛周皮膚病進行HPV DNA檢測和分型的可行性；②檢測CA患者皮損處整合素α6 mRNA的表達,初步探討其在CA的發(fā)病與復發(fā)中的意義；③探討整合素α6單克隆抗體(GoH3)對HPV 6/11病毒顆粒體外感染永生化人角質(zhì)形成細胞株(HaCaT細胞)的影響。 方法①應用PCR-RFLP-基因測序技術對發(fā)生于外生殖器和肛周部位的CA、鮑溫樣丘疹病(BP)、鱗狀細胞癌(SCC)、硬化萎縮性苔蘚(LSA)和乳房外Paget病(EMPD)的皮損組織進行HPV DNA檢測和分型；②采用RT-PCR方法測定初發(fā)和復發(fā)CA患者皮損處整合素α6 mRNA的表達；③應用熒光定量聚合酶鏈反應(FQ-PCR)方法檢測GoH3對臨床CA組織中提取的HPV 6/病毒顆粒感染HaCaT細胞后HPV DNA載量的影響。 結(jié)果①50例經(jīng)病理確診的CA及四種外生殖器和肛周皮膚病標本中,共有35例檢測出HPV DNA,其陽性率分別為CA 100.0%(26/26)、BP 47.1%(8/17)、SCC 50.0%(1/2)、LSA 0.0%(0/4)和EMPD 0.0%(0/1); HPV DNA陽性標本中,HPV 6型19例,HPV-11型3例,HPV 16型8例,HPV 6型+11型4例,HPV 62型1例；基因測序結(jié)果與PCR-RFLP判定的HPV型別相符；②整合素α6在初發(fā)和復發(fā)CA皮損中.nRNA的相對病毒載量分別0.79±0.16和1.07±0.29,均高于正常包皮組織的0.25+0.10,差異有統(tǒng)計學意義(P=0.001)；復發(fā)CA皮損中整合素α6的表達高于初發(fā),差異有統(tǒng)計學意義(P0.05)；③4組不同濃度的HPV 6/11病毒顆粒感染HaCaT細胞后測得的HPV病毒載量不同(P=0.001)。當病毒顆粒濃度為106copies/ml時,HaCaT細胞被感染率為100%,且所測得的HPV載量高于其它3組(P0.05)；當GoH3稀釋倍數(shù)1：100時,其對HPV感染HaCaT細胞的抑制率與其濃度成正比；但當稀釋倍數(shù)1：100時,再增加GoH3的濃度,其抑制率也未見明顯增長；8例臨床分離的HPV 6/11病毒顆粒在加入GoH3之前感染HaCaT細胞的平均病毒載量為(5.8±2.51)×104copies/ml,加入GoH3后感染HaCaT細胞的平均病毒載量為(3.02±1.21)×104copies/ml, GoH3的平均抑制率為(46.9±4.7)%。 結(jié)論①PCR-RFLP方法可用于HPV DNA檢測與分型；②整合素α6可能在CA發(fā)病和復發(fā)中發(fā)揮作用；③GoH3對HPV 6/11型感染HaCaT細胞具有一定的抑制作用。
[Abstract]:Condyloma acuminatum (CA) is caused by the human papillomavirus (human papillomavirus, HPV), which occurs in the genitals, and the sexually transmitted disease (sexually transmitted disease, STD) is not only high, but also has a high relapse rate after treatment. There are a variety of treatment methods for CA, but any one of them. The methods can not avoid the recurrence. Different types of HPV infection can cause different clinical manifestations. The classification of HPV will be beneficial to the diagnosis and prognosis of clinical diseases.
Integrin (integrin) is one of the most important members of the cell adhesion molecule family. Through a large number of studies on the relationship between integrin and virus infection, it is found that a variety of viruses can use integrin as a viral receptor or a common receptor (co-receptor) into the host cell. Integrin alpha 6 is mainly expressed in the basal layer of normal epithelial tissue, and HPV The damage to the host, first of all, begins with the infection of the basal layer of the epithelial tissue of the host. Is there a correlation between the two? Does the integrin alpha 6 play a role in the HPV infection? If the expression of the integrin is closed at this beginning, can the adhesion of the virus to the host cell be inhibited? Further studies will help to fundamentally block the infection of HPV virus and the recurrence of CA.
Objective (1) to identify and evaluate the feasibility of PCR combined restriction fragment length polymorphism (RFLP) and gene sequencing technology for HPV DNA detection and typing of CA and four kinds of external and perianal skin diseases. (2) the expression of integrin alpha 6 mRNA in the skin lesions of CA patients was detected and its significance in the pathogenesis and recurrence of CA was preliminarily discussed. The effect of monoclonal antibody GoH3 (HPV) on HPV 6/11 virus particles infecting immortalized human keratinocyte line (HaCaT cells) in vitro.
Methods (1) PCR-RFLP- gene sequencing was used to detect CA, bautherid papular disease (BP), squamous cell carcinoma (SCC), BP, LSA and Paget disease (EMPD) in the genital and perianal areas, and to detect and classify the lesions of the lesions of the skin tissues of Paget disease (EMPD). (2) the determination of integrin alpha at the skin lesions of primary and recurrence CA patients by RT-PCR method. 6 mRNA expression; (3) using fluorescence quantitative polymerase chain reaction (FQ-PCR) to detect the effect of GoH3 on the HPV DNA load of HPV 6/ virus infected HaCaT cells extracted from the clinical CA tissue.
Results (1) in 50 cases of CA and four types of external genitals and perianal skin diseases, there were 35 cases of HPV DNA detection, the positive rates were CA 100% (26/26), BP 47.1% (8/17), SCC 50% (1/2), LSA 0% (0/4) and EMPD 0%. The results of gene sequencing in 1 cases were consistent with the HPV type determined by PCR-RFLP, and the relative viral load of integrin alpha 6 in the primary and recurrent CA lesions was 0.79 + 0.16 and 1.07 + 0.29, respectively, higher than that of normal foreskin (P=0.001), and the expression of integrin alpha 6 in the recurrent CA lesions was higher than that of the initial hair. The difference was statistically significant (P0.05); (3) the HPV virus load of the 4 groups of different concentrations of HPV 6/11 virus particles infected with HaCaT cells was different (P=0.001). When the concentration of the virus particles was 106copies/ml, the infection rate of HaCaT cells was 100%, and the measured HPV load was higher than that of the other 3 groups (P0.05); when GoH3 dilution multiple 1:100, it was infected with the infection The inhibitory rate of HaCaT cells was proportional to its concentration, but when the dilution multiple of 1:100, the inhibitory rate of GoH3 was not significantly increased, and the average viral load of the 8 clinically isolated HPV 6/11 virus particles infected with HaCaT cells before adding GoH3 was (5.8 + 2.51) x 104copies/ml, and the average infection of HaCaT cells after the addition of GoH3 was observed. The viral load was (3.02 + 1.21) x 104copies/ml, and the average inhibition rate of GoH3 was (46.9 + 4.7)%.
Conclusion (1) PCR-RFLP method can be used for HPV DNA detection and typing, and integrin alpha 6 may play a role in the pathogenesis and recurrence of CA; (3) GoH3 has a certain inhibitory effect on HaCaT cells of HPV 6/11 infection.
【學位授予單位】：天津醫(yī)科大學
【學位級別】：碩士
【學位授予年份】：2011
【分類號】：R759

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本文編號：2118398