Physical state and expression of HPV DNA in benign and dysplastic cervical tissue: different levels of viral integration are correlated with lesion grade
Physical state and expression of HPV DNA in benign and dysplastic cervical tissue: different levels of viral integration are correlated with lesion grade
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DOI：
10.1016/j.ygyno.2003.11.035
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发表时间：
2004-03-01
影响因子：
4.7
通讯作者：
Czerwenk, KF
中科院分区：
文献类型：
--
作者：
Hudelist, G;Manavi, M;Czerwenk, KF
Objective.人乳头瘤病毒（HPV）感染是宫颈上皮恶变的重要原因。已经鉴定了几种高危（HR-）HPV亚型，其导致CIN并随后导致浸润性癌。这种现象的原因尚不清楚，但它似乎与HPV DNA的物理状态有关。Digene HC 11检测用于确定HR和/或低风险（LR-）HPV感染的275名妇女的宫颈拭子参加我们的诊所进行常规细胞学筛查和/或阴道镜检查，因为异常巴氏涂片包括低度鳞状上皮内病变（LGSIL）和高度SIL（HGSIL）。应用PCR-限制性内切酶方法对10例宫颈良性病变和68例CIN Ⅰ-Ⅲ患者的宫颈活检标本进行了FIR（16，18，31，33，52 b，58）和LR（6，11）亚型的分析。通过双向（2D）凝胶电泳分析HPV DNA的物理状态（游离型、混合型和整合型）。RT-PCR检测E6/E7基因mRNA的表达。此外，在9例选定病例中测定了相对病毒载量。HPV DNA的物理状态和转录活性与组织病理学结果相关。宫颈拭子中LR-HPV感染27例（9.8%），HR-HPV感染121例（44%），与SIL的程度明显相关。78例妇女宫颈活检中进一步的HPV分型显示，HPV 6和II仅限于良性细胞病变，CIN I和II，而HPV 16和18主要见于CIN III/CIS（P= 0.01）。HPV 31、33、52 b和58未观察到明显的分布模式。HPV E6和E7转录本的表达与HPV DNA的不同物理状态一致相关。分析这些HPV亚型的物理状态，HPV 6和II只能作为游离型检测到，与SIL分级无关。在正常上皮和CIN I和II中，HPV 16和18仅以游离型存在。在CIN III/CIS中，30例HPV 16中的15例（50%）和17例HPV 18中的16例（94%）仅整合到宿主基因组中。与HPV 16/18一样，HPV 31、33、52 b和58在正常上皮和CIN I、II中也以游离型存在，但在CIN III/CIS中整合率为80%（4/5）。一些CIN III/CIS中HPV 16 DNA整合缺失表明，早期发育异常病变进展并不总是需要整合。相反，18型HPV和其他HPV的整合似乎对宫颈发育不良的转化效果至关重要。所应用的方法代表了评估HPV物理状态的敏感工具，并可用于预测疾病的进展。(C)2004年爱思唯尔公司All rights reserved.
Objective. Human papillomavirus (HPV) infection is the most important event in the malignant transformation of human cervical epithelium. Several high-risk (HR-)HPV subtypes have been identified, which lead to CIN and subsequently to invasive carcinoma. The reason for this phenomenon is still unknown, but it seems to be related to the physical state of HPV DNA.Methods. Digene HC 11 test was used to identify HR- and/or low-risk (LR-)HPV infections in cervical swabs of 275 women attending our clinic for routine cytological screening and/or colposcopy because of an abnormal Pap smear comprising low-grade squamous intraepithelial lesions (LGSIL) and high-grade SIL (HGSIL). Specific FIR (16, 18, 31, 33, 52b, 58) and LR (6, 11) subtypes were characterized in cervical biopsies of 10 women with benign cellular changes and of 68 women with CIN I-III by the PCR-restriction enzyme method. The physical state of HPV DNA (episomal, mixed and integrated form) was analyzed by bi-dimensional (2D)-gel electrophoresis. In addition, mRNA expression of E6/E7 genes was analyzed by RT-PCR. Furthermore, the relative virus load was determined in nine selected cases. The physical state and transcriptional activity of HPV DNA were then correlated to histopathological results.Results. LR-HPV infection [27 cases (9.8%)] and HR-HPV infection [121 cases (44%)] of cervical swabs were clearly correlated to the degree of SIL. Further HPV typing in cervical biopsies of 78 women showed that HPV6 and II were restricted to benign cellular changes, CIN I and II, whereas HPV16 and 18 were observed predominantly in CIN III/CIS (P= 0.01). No clear distribution pattern was observed for HPV31, 33, 52b and 58. Expression of HPV E6 and E7 transcripts was uniformly correlated with the different physical state of HPV DNA. Analyzing the physical state of these HPV subtypes, HPV6 and II could only be detected as an episomal form, independent of SIL grade. In normal epithelium and in CIN I and II, HPV 16 and 18 were exclusively found in the episomal form. In CIN III/CIS, 15 of 30 cases of HPV 16 (50%) and 16 of 17 cases of HPV18 (94%) were exclusively integrated into the host genome. Like HPV16/18, HPV31, 33, 52b and 58 were also present in the episomal form in normal epithelium and in CIN I and II, but were integrated in 80% of the CIN III/CIS (4/5) cases.Conclusion. Absent integration of HPV 16 DNA in some CIN III/CIS suggests that integration is not always required for progression early dysplastic lesions. In contrast, integration of HPV type 18 and others appears to be of major importance for the transforming efficacy of cervical dysplasia. The applied method represents a sensitive instrument to assess the physical state of HPV and is useful to predict the progression of disease. (C) 2004 Elsevier Inc. All rights reserved.