高危型HPV E6/E7 mRNA检测对宫颈高级别病变的预警分流价值

目的:通过比较检测HPV E6/E7 mRNA和HPV DNA两种HPV检测方法对宫颈高级别病变的检出能力,探讨HPV E6/E7 mRNA检测在宫颈癌筛查中的预警分流价值。方法:采集2016年7月至2017年6月就诊于我院接受宫颈癌筛查的1 515例女性宫颈脱落细胞,利用转录介导扩增法检测高危型HPV E6/E7 mRNA（n=505）和PCR+膜杂交法检测HPV DNA（n=1 010）,以组织病理学诊断为金标准,比较两种检测方法对宫颈高级别病变检出率。结果:利用HPV E6/E7 mRNA检测组阴道镜转诊率32.69%（17/52）、HPV DNA检测组阴道镜转诊率10.62%（12/113）,差异有统计学意义（P＝0.001）;HPV E6/E7 mRNA检测组CIN2+的检出率42.42%;HPV DNA组 CIN2+的检出率28.16%,差异有统计学意义（P＝0.034）。结论:与HPV DNA检测方法相比,HPV E6/E7 mRNA检测对CIN2+病变预警分流价值较高。     

p16INK4A和HPV16 E7/HPV18 E6在ASCUS中的表达及临床意义

目的：探讨p16^INK4A和HPV16 E7／HPV18 E6在宫颈细胞学诊断为非典型性鳞状细胞不明确意义型（ASCUS）中的表达及筛查潜在宫颈病变的价值。方法：对150例ASCUS患者行阴道镜检查并取活检，同时对该150例患者的TCT标本进行免疫细胞化学染色检测p16^INK4A的表达和RT-PCR法检测其中HPV16型口蛋白和18型E6蛋白（HPV16 E7／HPV18 E6）mRNA的表达。结果：p16^INK4A和HPV16 E7／HPV18 E6 mRNA在ASCUS中表达的阳性率分别为37．33％和46．67％，随着病理级别的增加，p16^INK4A和HPV16 E7／HPV18 E6 mRNA表达的阳性率也随之增加；p16^INK4A和HPV16 E7／HPV18 E6 mRNA筛查ASCUS中宫颈病变的灵敏度、特异度、阳性预测值和阴性预测值分别为0．88、0．95、0．91、0．93和0．81、0．75、0．67、0．86，在p16^INK4A和HPV16 E7／HPV18 E6 mRNA阳性的样本中，宫颈病变发生率分别为91．07％和67．14％，均明显高于阴性样本中的发病率7．45％和13．75％（P〈0．001）；ASCUS中宫颈病变样本中p16^INK4A和HPV16 E7／HPV18 E6 mRNA呈高表达，且具有较高的一致性（K=0．6475）。结论：p16^INK4A和HPV16 E7／HPV18 E6 mRNA在ASCUS中病理诊断为宫颈病变的样本中均呈高表达，对筛查潜在的宫颈病变具有重要意义，其中p16^INK4A的筛查效能优于HPV16 E7／HPV18 E6 mRNA；p16^INK4A能间接反映HPV的转录活性，在ASCUS的分流中有重要意义，且可视性的p16^INK4A免疫染色比HPV检测更直观。     

HPV E6/E7 mRNA和HPV DNA检测技术在宫颈上皮内瘤变中的诊断价值分析

目的比较人乳头瘤病毒(HPV)E6/E7 mRNA和HPV DNA检测技术对宫颈上皮内瘤变(CIN)2级及以上(CIN2+)患者的诊断价值,并评价HPV E6/E7 mRNA检测结果在不同实验室间的一致性。方法采用HPV E6/E7 mRNA和HPV DNA检测技术对212例门诊体检的健康者和住院的宫颈病变患者的宫颈脱落细胞学标本进行检测。以病理诊断结果为金标准,评价两种检测技术诊断CIN2+的灵敏度和特异度。北京市迪安中心实验室和北京市怀柔妇幼保健院实验室均采用HPV E6/E7 m RNA检测技术检测同一批标本,评价实验室间检测的一致性。结果HPV E6/E7 m RNA检测的阳性率为38.7%,与HPV DNA的阳性率43.9%比较,差异无统计学意义(P﹥0.05)。HPV E6/E7 mRNA和HPV DNA的检测阳性率均随着病理分级的升高而增加(P<0.01)。HPV E6/E7mRNA检测CIN2+的灵敏度为92.96%,与HPV DNA的90.14%相比,差异无统计学意义(P﹥0.05),而HPV E6/E7mRNA检测CIN2+的特异度为88.65%,高于HPV DNA的79.43%,差异有统计学意义(P<0.05)。两个实验室采用HPV E6/E7 m RNA检测阳性一致的标本例数为78,阴性一致的标本例数为121,总一致率为93.87%,Kappa=0.872,一致性较好。结论与HPV DNA检测技术相比,HPV E6/E7 mRNA检测宫颈病变的特异度更具优势,实验室间重复性检测的一致率较高,有望成为中国宫颈癌HPV筛查的首选方法。     

叶酸水平及 HPV16感染对妇女发生宫颈癌变的协同作用研究

目的：探讨叶酸水平、HPV16感染及HPV16 E2和E6 mRNA水平与宫颈癌变的关系,并研究叶酸水平及HPV16感染在宫颈癌变中的协同作用。方法选择新发宫颈炎症（ CI）患者40例、宫颈上皮内瘤样变（ CIN）患者80例（ CINⅠ患者36例,CINⅡ／Ⅲ44例）、宫颈鳞状细胞癌（ SCC）患者48例作为研究对象,采用微生物法测定血清叶酸和红细胞叶酸含量,采用PCR检测宫颈癌组织HPV16的感染情况,采用荧光定量PCR检测宫颈癌组织和叶酸体外干预后Caski宫颈癌细胞中HPV16 E2及E6 mRNA水平,检测不同叶酸浓度对Caski和C33A细胞抑制情况。结果 CINⅠ组、CINⅡ／Ⅲ组和SCC 组HPV16感染率分别与CI 组宫颈组织HPV16感染率比较,差异具有统计学意义（ P＜0．05）。CINⅠ、CINⅡ／Ⅲ、SCC 各组HPV16 E2和E6 mRNA水平与CI组比较差异均具有统计学意义（P＜0．0001）。宫颈癌的严重程度与HPV16 E2和E6 mRNA水平均呈明显正相关性,与血清叶酸和红细胞叶酸含量均呈显著负相关性。叶酸浓度与Caski细胞和C33A细胞的相对抑制率及HPV16 E2和E6 mRNA水平呈明显负相关性。结论叶酸水平低、HPV16感染及HPV16 E2及E6 mRNA低表达均与宫颈癌变有关,补充叶酸明显抑制宫颈癌细胞的增殖,逆转HPV16 E2及E6高表达,提示叶酸水平及HPV16感染在宫颈癌变的过程中具有协同作用。     

HPV E6/E7 mRNA检测对宫颈癌筛查意义的初步评价

目的:利用超薄液基细胞学检测标本(LCT)分析高危人乳头瘤病毒(HPV)E6/E7mRNA检测在宫颈癌筛查中的意义。方法:选取231例液基细胞学检测标本,根据病理级别分为炎症/良性组(n=78)、ASCUS组(n=140)、LSIL组(n=8)和HSIL组(n=5)。利用bDNA技术检测细胞学标本中HPV E6/E7mRNA、DNA,结合病理诊断资料,进行统计学分析。结果:对不同细胞学级别中mRNA及DNA检测情况行配对χ2检验显示,炎症/正常细胞组DNA和mR-NA,阳性检出率差异无统计学意义,χ2=2.307,P>0.05;ASCUS组阳性检出率差异有统计学意义,χ2=9.082,P=0.020;LSIL组与HSIL组例数少,阳性检出率差异无统计学意义,P值分别为0.464和0.800。34例行宫颈活检的AS-CUS标本,2种检测指标在检出率差异无统计学意义,P值均>0.05;在进行活检的43例标本中,对不同病理级别HPVE6/E7mRNA及DNA检测情况行配对Fisher精确概率检验,2种检测指标检出率差异无统计学意义,P值均>0.05。分析预测效能,HPV E6/E7mRNA对LSIL/HSIL诊断敏感性为33.33%,95%CI为17.19～54.63;特异性为72.73%,95%CI为51.85～86.85;阳性预测值为53.85%,95%CI为29.14～76.79;阴性预测值为53.33%,95%CI为36.14～69.77。HPV E6/E7mRNA联合细胞学检测诊断敏感性为100%,95%CI为84.54～100.00;阴性预测值为100%,95%CI为43.85～100.00。结论:高危HPV E6/E7mRNA可以应用于宫颈癌筛查,在筛查中联合HPV E6/E7mRNA及细胞学检测有利于提高诊断准确性。     

子宫颈石蜡包埋样本检测HPV E6/E7mRNA用于子宫颈癌筛查的价值

目的 探讨检测石蜡包埋样本HPV E6/E7 mRNA在子宫颈癌筛查及预后判断中的应用价值.方法 应用支链DNA技术对石蜡包埋样本行HPV E6/E7 mRNA定量检测,以病理学诊断结果为标准分组并对拷贝值进行分析.比较各组间HPV E6/E7 mRNA阳性率及拷贝均数.结果 子宫颈癌组的HPVE 6/E7 mRNA阳性率高于子宫颈上皮内瘤变（CIN）1、CIN2、CIN3组、慢性子宫颈炎组[分别为94.29％（66/70）、30.30％（20/66）、50.00％（19/38）、59.46％（44/74）、5.66 ％（3/50）,均P＜ 0.004 5],CIN组的阳性率均高于慢性子宫颈炎组（P＜0.004 5）.CIN2组和CIN3组的阳性率均高于CIN1组（P＜0.0045）,CIN2组和CIN3组间的阳性率差异无统计学意义（P＞0.004 5）.子宫颈癌组的拷贝值高于慢性子宫颈炎组、CIN1、CIN2、CIN3组（分别为16 508.730±8 741.402、929.337±460.880、3 239.681±1 447.399、5 286.224±2 933.445、4.941±2.263,均P＜0.05）,CIN组的拷贝值均高于慢性子宫颈炎组（P＜0.05）,CIN3组的拷贝数值高于CIN1组（P＜0.05）,CIN1组和CIN2组、CIN2组和CIN3组间的拷贝值差异均无统计学意义（P＞0.05）.结论 HPV E6/E7 mRNA阳性率和拷贝值的变化趋势与子宫颈病变的程度有关.子宫颈石蜡包埋组织检测HPV E6/E7 mRNA在预见疾病风险上有较高的临床应用价值,可作为子宫颈癌筛查和指导随访的有效措施.     

宫颈癌及其癌前病变组织中HPV16 E6、HPV16 E7蛋白的表达及其意义

目的 探讨宫颈癌及其癌前病变组织中HPV16E6、E7蛋白的表达及其意义。方法 采用免疫组化SP法对15例正常宫颈、25例宫颈上皮内瘤变（C1N）以及61例浸润性宫颈癌组织进行了HPVl6E6、HPVl6E7蛋白表达的检测。结果在正常宫颈、CINI～Ⅱ、CINⅢ及浸润性宫颈癌中，HPVl6E6蛋白的阳性表达率分别为0（0／15）、7．14％（1／14）、36．36％（4／11）、59．02％（36／61）；CINⅢ和浸润性宫颈癌中HPVl6E6蛋白阳性表达率明显高于正常宫颈组织和CINI～Ⅱ（P〈0．05）；在高、中、低分化宫颈癌中，HPVl6E6蛋白阳性表达率分别为45．45％（5／11）、77．78％（14／18）、53．13％（17／32）；HPVl6E6蛋白在不同分化程度的宫颈癌组织中的阳性表达率有显著性差异（P〈0．05），但HPVl6E6蛋白表达与官颈癌组织分化程度无明显相关性（ys=0．123），HPVl6E6蛋白阳性表达率与宫颈癌临床分期和淋巴结转移无关（P〉0．05）。HPVl6E7蛋白在正常宫颈上皮、CINI-Ⅱ、CIN Ⅲ及浸润性富颈癌组织中的阳性表达率分别为20．00％（3／15）、42．86％（6／14）、63．64％（7／11）、57．38％（35／61），HPVl6E7蛋白在不同宫颈组织中的阳性表达率无明显差异（P=0．05）；HPVl6E7蛋白的表达与富颈癌临床分期、淋巴结转移和组织分化均无关（P〉0．05）。结论 HPVl6E6蛋白的检测有可能作为宫颈癌前病变转归的指标。     

hTERC和高危HPV及细胞学联合检测在宫颈癌筛查中的价值

目的：探讨联合应用hTERC、高危HPV检测及液基细胞学检查TCT在宫颈癌筛查中的意义。方法：选择2010年5月至2012年1月因宫颈异常在佛山市妇幼保健院宫颈门诊行TCT检查的患者100例为研究对象,同时行HC2检测高危HPV感染,FISH技术检测hTERC基因扩增及阴道镜下病理活组织检查,以病理学结果为金标准,将三种检测方法的结果与其进行比较。结果：非CIN组、CIN1、高级别CIN及浸润癌中TCT检查与病理学结果符合率分别为88．9％（16／18）、58．33％（14／24）、68．29％（28／41）、76．47％（13／17）,高危HPV阳性率分别为27．78％（5／18）、91．67％（22／24）、97．56％（40／41）、100％（17／17）,hTERC阳性率分别为11．1％（2／18）、29．17％（7／24）、87．80％（36／41）、100％（17／17）。随着宫颈病变病理级别的递增,高危HPV的表达及hTERC基因扩增出现递次增高。非CIN组高危HPV感染率低于CIN组,差异有统计学意义（P〈0．01）,CIN1、CIN2及CIN3三组间比较差异无统计学意义（P〉0．05）,CIN组与宫颈浸润癌组差异无统计学意义（P〉0．05）。hTERC基因扩增率高级别CIN组及浸润癌组明显高于低级别CIN1组（P〈0．01）。结论：在宫颈癌筛查中,细胞学检查具有较高的特异性（92．5％）,但灵敏度较差（53．4％）,高危HPV检测灵敏度高（96．7％）,但特异性差（44．6％）,hTERC基因扩增筛查高级别CIN病变特异性高（98．6％）,灵敏度稍低（71．7％）,在宫颈组织的表达率与宫颈病变程度密切相关,三种方法联合检测可以大大提高宫颈癌前病变的阳性检出率,有利于较特异地识别出具有较高的转化为宫颈癌潜能的高级别CIN病变。     

头颈部鳞癌患者HPV DNA、E6/E7 mRNA的分布及其在预后中的作用

摘 要：［目的］ 检测头颈部鳞癌组织标本人乳头瘤病毒(HPV)DNA、E6/E7 mRNA感染情况,并分析HPV感染对预后的影响。［方法］ 收集2016—2018年头颈部鳞癌手术标本121例,采用MassARRAY?誖HPV基因分型技术和RT-qPCR方法分别进行HPV DNA和HPV E6/E7 mRNA检测,比较不同肿瘤部位检出率差异。采用多因素Cox比例风险回归模型分析HPV E6/E7 mRNA对头颈部鳞癌预后的影响,计算HR值及其95%可信区间。［结果］ 121例头颈部鳞癌患者HPV阳性率为66.94%（81/121）,其中口腔癌阳性率为70.37%（19/27）,喉癌为65.79%（50/76）,口咽癌为66.67%（12/18）,HPV DNA阳性率在不同肿瘤部位无统计学差异（P>0.05）。HPV16阳性率为4.13%（5/121）,HPV18阳性率为6.61%（8/121）。头颈部鳞癌HPV E6/E7 mRNA阳性率为28.93%（35/121）,HPV E6/E7 mRNA阳性率在不同肿瘤部位中无统计学差异（P>0.05）。多因素分析结果显示,与HPV E6/E7 mRNA阴性组相比,HPV E6/E7 mRNA阳性组预后较好（HR=0.13,95%CI：0.04~0.46）。［结论］ 头颈部鳞癌患者HPV DNA阳性率高,但HPV E6/E7 mRNA阳性率明显降低。HPV感染为头颈部鳞癌独立的预后因素。     

宫颈细胞内p16/Ki67表达与宫颈癌及癌前病变的关系及其用于HPV阳性人群分流的诊断价值

摘 要：［目的］ 探讨p16/Ki67双染技术对宫颈癌及癌前病变的检出能力,比较其用于HPV阳性人群分流的诊断价值。［方法］ 选取2016年9月至2017年5月山西长治三家三甲医院21~70岁参加宫颈癌筛查的妇女及门诊患者共计497例,所有入组研究对象均进行HPV E6/E7mRNA检测、液基细胞学检测（LBC）与p16/Ki67双染。［结果］ p16/Ki67双染的阳性率随细胞学诊断及病理诊断的严重程度的增加而升高(P<0.05）;p16/Ki67对62例宫颈上皮内瘤变2级及以上（CIN2+）研究对象检出的灵敏度、特异性分别为69.4%、83.6%,HPV E6/E7mRNA检测分别为82.3%、74.6%,LBC检测分别为80.6%、65.2%。对49例CIN3+研究对象检出的灵敏度、特异性分别为69.4%、81.9%,HPV E6/E7mRNA检测分别为85.7%、73.3%。在CIN2+/CIN3+病例中p16/Ki67的灵敏度与LBC均衡可比,但其特异性明显高于 LBC 和HPV E6/E7mRNA检测。在HPV E6/E7mRNA检测阳性人群中p16/Ki67双染对CIN2+检出的灵敏度和特异性分别为82.4%和56.9%,LBC检测为90.2%和29.4%;p16/Ki67双染对CIN3+检出的灵敏度和特异性分别为81.0%和53.2%,LBC检测为88.1%和27.0%。p16/Ki67双染中对CIN2+及CIN3+检出的灵敏度与LBC检测相比均无明显差异（P>0.05）,但其特异性明显高于LBC检测（P<0.001）。［结论］ p16/Ki67双染技术用于HPV阳性人群的分流具有优于LBC的特异性和与LBC检测相似灵敏度的特性。在经济条件允许或者缺乏病理医生的地区,可以考虑使用p16/Ki67代替细胞学用于宫颈癌的初筛或HPV阳性人群的分流。     

Diagnostic Performance of HPV E6/E7 mRNA and HPV DNA Assays for the Detection and Screening of Oncogenic Human Papillomavirus Infection among Woman with Cervical Lesions in China

Background: Human papillomavirus (HPV) is the most common sexually transmitted infection worldwide and it is responsible for most cases of cervical uterine cancer. Although HPV infections of the cervix do not always progress to cancer, 90% of cervical cancer cases have been found to be associated with high risk HPV (HRHPV) infection. HPV DNA testing is widely used, along with Papanicolaou (Pap) testing, to screen for cervical abnormalities. However, there are no data on the prevalence of genotype-specific HPV infections assessed by measuring HPV E6/E7 mRNA in women representative of the Chinese population across a broad age range. Materials and Methods: In the present study, we compared the results with the CervicGen HPV RT-qDx assay, which detects 16 HR-HPV genotypes (Alpha-9: HPV 16, 31, 33, 35, 52, and 58; Alpha-7: HPV 18, 39, 45, 51, 59, and 68; and Alpha-5, 6: HPV 53, 56, 66, and 69), and the REBA HPV-ID assay, which detects 32 HPV genotypes based on the reverse blot hybridization assay (REBA) for the detection of oncogenic HPV infection according to cytological diagnosis. We also investigated the prevalence and genotype distribution of HPV infection with a total of 324 liquid-based cytology samples collected in western Shandong province, East China. Results: The overall HPV prevalences determined by HPV DNA and HPV E6/E7 mRNA assays in this study were 79.9% (259/324) and 55.6% (180/324), respectively. Although the positivity of HPV E6/E7 mRNA expression was significantly lower than HPV DNA positivity, the HPV E6/E7 mRNA assay showed greater specificity than the HPV DNA assay (88.6% vs. 48.1%) in normal cytology samples. The prevalence of Alpha-9 (HPV 16, 31, 33, 35, 52, and 58) HPV infection among these women accounted for up to 80.3% and 76.1% of the high-grade lesions detected in the HPV mRNA and DNA tests, respectively. The HR-HPV genotype distribution, based on HPV DNA and E6/E7 mRNA expression by age group in patients with cytologically confirmed lesions, was highest in women aged 40 to 49 years (35.9% for cytologically confirmed cases, Pearson correlation r value=0.993, p<0.001) for high-grade lesions. Among the oncogenic HR-HPV genotypes for all age groups, there was little difference in the distribution of HPV genotypes between the HPV DNA (HPV -16, 53, 18, 58, and 33) and HPV E6/E7 mRNA (HPV -16, 53, 33, 58, and 18) assays. HPV 16 was the most common HPV genotype among women with highgrade lesions. Conclusions: Our results suggest that the HPV E6/E7 mRNA assay can be a sensitive and specific tool for the screening and investigation of cervical cancer. Furthermore, it may provide useful information regarding the necessity for early cervical cancer screenings and the development of additional effective HPV vaccines, such as one for HPV 53 and 58. Additionally, gaining knowledge of HPV distribution may also inform us about ecological changes in HPV after the vaccination.     

Human Papillomavirus Genotype Distribution and E6/ E7 Oncogene Expression in Turkish Women with Cervical Cytological Findings

Background: Infection with certain human papillomavirus (HPV) genotypes is the most important riskfactor related with cervical cancer. The objective of the present study was to investigate the prevalence of HPVinfection, the distribution of HPV genotypes and HPV E6/E7 oncogene mRNA expression in Turkish women withdifferent cervical cytological findings in Mersin province, Southern Turkey. Materials and Methods: A total of476 cytological samples belonging to women with normal and abnormal cervical Pap smears were enrolled in thestudy. For the detection and genotyping assay, a PCR/direct cycle sequencing approach was used. E6/E7 mRNAexpression of HPV-16, 18, 31, 33, and 45 was determined by type-specific real-time NASBA assay (NucliSENSEasyQ®HPV v1.1). Results: Of the 476 samples, 106 (22.3%) were found to be positive for HPV DNA by PCR.The presence of HPV was significantly more common (p<0.001) in HSIL (6/8, 75%) when compared with LSIL(6/14, 42.9%), ASC-US (22/74, 29.7%) and normal cytology (72/380, 18.9%). The most prevalent genotypes were,in descending order of frequency, HPV genotype 66 (22.6%), 16 (20.8%), 6 (14.2%), 31 (11.3%), 53 (5.7%), and83 (4.7%). HPV E6/E7 oncogene mRNA positivity (12/476, 2.5%) was lower than DNA positivity (38/476, 7.9%).Conclusions: Our data present a wide distribution of HPV genotypes in the analyzed population. HPV genotypes66, 16, 6, 31, 53 and 83 were the predominant types and most of them were potential carcinogenic types. Becauseof the differences between HPV E6/E7 mRNA and DNA positivity, further studies are required to test the roleof mRNA testing in the triage of women with abnormal cervical cytology or follow up of HPV DNA positive andcytology negative. These epidemiological data will be important to determine the future impact of vaccinationon HPV infected women in our region.     

Eight-type human papillomavirus E6/E7 oncoprotein detection as a novel and promising triage strategy for managing HPV-positive women

The management of HPV-positive women becomes particularly crucial in cervical cancer screening. Here we assessed whether detection of E6 or E7 oncoproteins targeting eight most prevalent HPV types could serve as a promising triage option. Women (N = 1,416) aged 50–60 from Shanxi, China underwent screening with HPV testing and liquid-based cytology (LBC), with any positive results referring to colposcopy and biopsy if necessary. Women with HPV-positive results received further tests using DNA-based genotyping, E6 or E7 oncoprotein detection targeting HPV16/18 (for short: E6 (16/18) Test) or HPV16/18/31/33/35/45/52/58 (for short: E6/E7 (8 types) Test), respectively. Among HPV-positive women, E6/E7 (8 types) oncoproteins had lower positivity (17.37%) compared to DNA-based genotyping for same eight types (58.30%) and LBC with ASC-US threshold (50.97%); HPV16 was the genotype showing the highest frequency (8.49%) for oncoprotein detection followed by HPV52 (3.47%), 58 (2.32%), 33 (1.54%), 18 (1.16%), 45 (0.77%), 35 (0.39%) and 31 (0%). For detection of cervical intraepithelial neoplasia Grade 3 or higher (CIN3+), E6/E7 (8 types) Test had similar sensitivity (100.00%) and superior specificity (85.94%) as well as positive predictive value (PPV, 22.22%) compared to both LBC and DNA-based genotyping (8 types); For detection of CIN2+, E6/E7 (8 types) Test was less sensitive (67.74%) but still more specific (89.47%) and risk predictive with PPV of 46.67%. Notably, E6/E7 (8 types) Test remarkably decreased the number of colposcopies needed to detect one CIN2+ and CIN3+ (2.14 and 4.50). E6/E7 oncoprotein detection showed a good “trade-off” between sensitivity and specificity with more efficient colposcopy referrals, which is of great importance to maximize the benefits of HPV-based screening program, especially applicable for the areas with high HPV prevalence and low-resources.     

A cross-sectional study of a prototype carcinogenic human papillomavirus E6/E7 messenger RNA assay for detection of cervical precancer and cancer.

PURPOSE: To evaluate carcinogenic human papillomavirus (HPV) mRNA for E6 and E7 mRNA detection on clinical specimens to identify women with cervical precancer and cancer. EXPERIMENTAL DESIGN: We evaluated a prototype assay that collectively detects oncogenes E6/E7 mRNA for 14 carcinogenic HPV genotypes on a sample of liquid cytology specimens (n=531), masked to clinical data and to the presence of HPV genotypes detected by PGMY09/11 L1 consensus primer PCR assay. RESULTS: We found an increasing likelihood of testing positive for carcinogenic HPV E6/E7 mRNA with increasing severity of cytology (P(Trend) < 0.0001) and histology (P(Trend) < 0.0001), with 94% of cervical intraepithelial neoplasia grade 3 (CIN3) histology cases (46 of 49) and all five cancer cases testing positive for carcinogenic HPV E6/E7 mRNA. Overall, fewer specimens tested positive for carcinogenic HPV E6/E7 mRNA than for carcinogenic HPV DNA (P<0.0001, McNemar's chi(2) test), especially in women with 相似文献   

高危型HPV E6/E7mRNA检测在宫颈机会性筛查中应用价值研究

目的 人乳头瘤病毒(human papillomavirus,HPV) DNA能提高检测宫颈上皮内瘤变2级+(cervical intraepithelial neoplasia 2+,CIN2+)的灵敏度,但也增加了一过性HPV感染的检出率.本研究应用高危型HPV(high risk human papillomavirus,hrHPV) E6/E7 mRNA对比二代杂交捕获(hybrid CaptureⅡ,HC2)、HPV分型,描述机会性筛查人群中HPV感染及宫颈病变分布特征,探讨其用于宫颈筛查的可行性.方法 选取2013-01-01-2015-12-31在青岛大学附属青岛市立医院(6 138例)和青岛市城阳区人民医院(1 653例)妇科门诊行宫颈液基细胞学筛查的女性共计7 791例,年龄21～65岁.所有筛查女性行液基细胞学检查的同时行HPV检测,根据HPV检测方法的不同分为HC2组、HPV分型组和E6/E7组共3组.计算各组细胞学结果异常率、HPV阳性率和宫颈病变检出率,比较E6/E7所描述的流行病学特征与HPV-DNA方法的不同.结果 E6/E7组各级细胞学结果分别为,未见上皮内瘤变及恶性病变(negative for intraepithelial lesion or malignancy,NILM) 87.51％,意义不明的非典型鳞状细胞(atypical squamous cells of undetermined significance,ASCUS)5.47％,低级别鳞状上皮内瘤变(low-grade squamous intraepithelial lesion,LSIL)3.51％,不除外高级别鳞状上皮内瘤变的非典型细胞(atypical squamous cells cannot exclude HSIL,ASC-H)0.52％,高级别鳞状上皮内瘤变(high-grade squamous intraepithelial lesion,HSIL)2.99％.HPV阳性中异常细胞学结果的比例明显高于HPV阴性,而NILM的比例低于HPV阴性,P＜0.001.随着细胞学异常程度的加重,HPV阳性率逐渐增高,差异有统计学意义,x2值分别为611.089、512.036和767.260,P值均＜0.001.E6/E7组中NILM的HPV阳性率为8.02％,合计阳性率为14.73％,均较HC2及分型组低,x2=30.174,P=0.000;x2 =22.991,P＜0.001. E6/E7组CIN2+/CIN3+检出率分别为5.92％和1.20％,3组间比较差异无统计学意义,x2值分别为1.499和0.711,P值分别为0.473和0.701.E6/E7阳性率在正常至浸润性宫颈癌(invasive cervical cancer,ICC)之间随病变程度加重而升高,P＜0.05.在正常病理结果中E6/E7阳性率为61.36％,低于HC2和HPV分型,x2=15.767,P＜0.001;其余病理结果中各组间HPV阳性率差异无统计学意义,P＞0.05.结论 hrHPV E6/E7 mRNA较之HC2和HPV分型能在不降低宫颈病变检出率的前提下减少一过性HPV感染的检出率,提示其在宫颈筛查中具有潜在应用价值.     

HPV18 E6E7 反义荧光真核表达载体的构建及其在宫颈癌HeLa 细胞中的表达

 目的 构建HPV18型E6E7反义荧光真核表达载体,并观察其对宫颈癌HeLa细胞中HPV18 E6和E7基因表达的影响,探索反义技术用于治疗临床HPV感染及宫颈癌的可能性。方法 以HPV18型全基因质粒为模板,PCR法扩增HPV18型E6E7区716bp片段,利用基因重组技术将目的片段反向插入荧光真核表达载体pEGFP-C1,EcoR I酶切并测序鉴定;采用脂转法将重组质粒pEGFP-HPV18 E6E7as（EGFP-18AS）转染宫颈癌HeLa细胞株,通过RT-PCR及western blot检测细胞中E6、E7 mRNA和蛋白的表达。结果 成功构建HPV18E6E7反义荧光真核表达载体EGFP-18AS,经脂质体转染HeLa细胞,48h后在荧光倒置显微镜下可见明显的绿色荧光,且细胞中E6、E7 mRNA及蛋白表达水平均明显下调。结论 反义荧光真核表达载体可以有效的抑制HPV18E6、E7癌基因的表达,为治疗HPV感染和宫颈癌提供了一种新的方法。     

High-risk human papillomavirus E6/E7 mRNA and L1 DNA as markers of residual/recurrent cervical intraepithelial neoplasia

The aim of this study was to assess the use of human papillomavirus (HPV) E6/E7 mRNA testing in the follow-up of women treated for cervical intraepithelial neoplasia (CIN) by conization and to compare the prognostic value of HPV E6/E7 mRNA to HPV L1 DNA and cytology. One hundred and forty-three women underwent cytological/histological testing, HPV DNA genotyping by Linear Array, and HPV E6/E7 mRNA testing by APTIMA HPV assay during follow-up after surgical treatment for histologically verified CIN. High-grade residual/recurrent disease (CIN2+/HSIL+) was identified in 7 (4.9%) women, and low-grade disease (CIN1/LSIL) in 25 (17.5%). At the inclusion visit 33 (23%) women were HPV DNA-positive; 13 (9.0%) were HPV E6/E7 mRNA-positive. HPV E6/E7 mRNA did not identify three women with high-grade disease. Presence of high-risk HPV DNA at the inclusion visit predicted 100% (95% CI 64.6-100) of high-grade residual/recurrent disease, with a specificity of 80.9% (95% CI 73.5-86.6); cytology had a sensitivity of 85.7%, and a specificity of 87.5%. HPV E6/E7 mRNA testing was a poor predictor of treatment failure, with a sensitivity of 57.1% (95% CI 25.0-84.2), but high specificity (93.4%; 95% CI 87.9-96.5). Detection of high-risk HPV DNA after treatment by conization identified 100% of women with residual/recurrent high-grade disease, whereas HPV E6/E7 mRNA testing was a poor predictor of treatment failure. This study suggests that a negative HPV mRNA result cannot exclude the risk of malignant progression, and that HPV E6/E7 mRNA testing by APTIMA HPV assay is not useful in the follow-up of women treated for CIN.     

早期宫颈癌组织中ATG-12表达与HPV16-E6/E7-DNA病毒载量及预后的关系

目的探讨早期宫颈癌组织中自噬相关蛋白-12(ATG-12)与人乳头瘤病毒(HPV)16-E6/E7-DNA病毒载量及预后的关系。方法选取2015年1月至2017年12月期间在安徽皖北煤电集团总医院保存的宫颈活检或手术切除宫颈组织203例,包括30例慢性宫颈炎、18例宫颈上皮内瘤变(CIN)Ⅰ、23例CINⅡ、26例CINⅢ和106例早期(ⅠA2~ⅡB期)宫颈鳞状上皮癌(CSCC)。采用巢式多重聚合酶链式反应(NM-PCR)和免疫组化染色SP法检测组织中HPV 16-E6/E7-DNA病毒载量和ATG-12蛋白表达。分析ATG-12蛋白表达与早期CSCC临床病理特征的关系。随访CSCC患者无瘤生存时间(DFS)。采用Cox风险比例回归模型分析影响CSCC预后的因素。结果CSCC组织ATG-12蛋白阳性表达率为26.42%(28/106),低于慢性宫颈炎和CIN组织(P<0.05)。ATG-12蛋白表达与HPV16-E6/E7-DNA病毒载量呈负相关(r=-0.306,P=0.001;r=-0.436,P=0.001)。ATG-12蛋白表达与FIGO分期、间质浸润、淋巴脉管间隙浸润、淋巴结转移有关(P<0.05)。随访11~64个月,ATG-12蛋白阳性表达患者5年无瘤生存率为78.57%(22/28),ATG-12蛋白表达阴性患者5年无瘤生存率为60.26%(47/78)。多因素Cox风险回归模型结果显示,HPV16-E6/E7-DNA病毒载量和ATG-12蛋白表达是影响患者DFS的独立预后因素(P<0.05)。结论早期宫颈鳞状上皮癌组织中ATG-12阳性表达率低,与HPV16-E6/E7-DNA病毒载量呈负相关,有望成为评估HPV阳性早期宫颈癌患者预后的重要指标。