四川地区宫颈癌HPV59型感染及其E6基因突变

宫颈癌是妇科常见的恶性肿瘤,特定型别的人乳头瘤病毒(HPV)感染是引发宫颈癌的主要因素[1].根据HPV致癌性的不同可分为高危型,低危型.Munoz N[2]等对来自9个国家的1918例宫颈癌组织进行各型HPV筛查,最终确认HPV-16、18、26、31、33、35、39、45、51、52、53、56、58、59、66、68、73及82型为高危型.李洁等[3]用核酸印迹技术对1986至1994年间来自我国14个省市自治区的1008宫颈癌组织进行HPV型别检测,发现HPV-16、18、31、35、51、58等型的存在,并发现各地区主要流行型别随地理位置有所差异.     

新疆维族妇女感染的HPV型别分布的研究

目的:探讨人乳头瘤状病毒(Human papillomavirus,HPV)在新疆南部维族妇女人群中的型别分布情况.方法:以年龄在30-59岁的新疆伽师县夏普吐勒乡维吾尔族妇女人群为基础进行筛查,签署知情同意书后,采集受试者宫颈脱落细胞,利用PCR和基因芯片技术检测HPVDNA并分型.结果:共2473名妇女入选.HPV总的感染率为9.1%,高危型中HPV-16的感染率最高为6.9%,其他高危型的感染率从高到低依次为HPV-59、HPV-56、HPV-18、HPV-33、HPV-58、HPV-51、HPV-31、HPV-45、HPV-52、HPV-68、HPV-35和HPV-39.低危型中HPV11感染率最高,其他低危型的感染率从高到低依次为HPV-42、HPV-43、HPV-6、HPV-53、HPV-66和HPV-73.HPV-44、HPV-83、HPV-MM4没检测到.多重感染率为34.2%.结论:新疆维吾尔族妇女人群中以HPV16感染为主,其次为HPV59、56、18、33等.HPV59可能是新疆维吾尔族妇女较易感染的类型.体现了新疆维吾尔族妇女感染HPV的特殊性.     

新疆维族妇女感染的HPV 型别分布的研究

目的:探讨人乳头瘤状病毒(Human papillomavirus,HPV)在新疆南部维族妇女人群中的型别分布情况。方法:以年龄在30-59岁的新疆伽师县夏普吐勒乡维吾尔族妇女人群为基础进行筛查,签署知情同意书后,采集受试者宫颈脱落细胞,利用PCR和基因芯片技术检测HPV DNA并分型。结果:共2473名妇女入选。HPV总的感染率为9.1%,高危型中HPV-16的感染率最高为6.9%,其他高危型的感染率从高到低依次为:HPV-59、HPV-56、HPV-18、HPV-33、HPV-58、HPV-51、HPV-31、HPV-45、HPV-52、HPV-68、HPV-35、HPV-39。低危型中HPV11感染率最高,其他低危型的感染率从高到低依次为HPV-42、HPV-43、HPV-6、HPV-53、HPV-66、HPV-73。HPV-44、-83、-MM4没检测到。多重感染率为34.2%。结论:新疆维吾尔族妇女人群中以HPV16感染为主,其次为HPV59、56、18、33等。HPV59可能是新疆维吾尔族妇女较易感染的类型。体现了新疆维吾尔族妇女感染HPV的特殊性。     

人乳头状瘤病毒感染与肺癌关系的研究进展

近年越来越多的研究发现肺癌组织中有人乳头状瘤病毒（human papillomavirus,HPV）DNA和癌蛋白的检出,其检出率具有地域差异性,但不同地方HPV检出亚型具有高度的一致性,主要是高危型HPV-16和HPV-18。HPV可能主要通过血液循环从宫颈癌病灶等原发部位转移到肺,从而可能诱发肺癌的发生。HPV诱发肺癌的机制可能与影响p53的失活及端粒酶的激活等有关。     

13933例妇科门诊患者人乳头瘤病毒感染状况分析

目的探讨河北医科大学第四医院妇科门诊患者人乳头瘤病毒(HPV)感染情况及型别分布情况,为预防HPV感染和宫颈癌防治提供依据。方法采用核酸分子快速导流杂交技术对13 933例妇科门诊患者的宫颈细胞标本进行21种HPV亚型的基因检测,不同年龄组HPV感染率比较采用卡方检验。结果 13933例受检者中,HPV感染5 979例,感染率为42.9%。共计检出21种亚型,9 023例次HPV感染,其中HPV高危型7 823例次占86.7%,感染前三位分别为HPV-16、HPV-52、HPV-58;HPV低危型1 200例次占13.3%,感染前三位分别为HPV-CP8304、HPV-11、HPV-6;17~25岁和≥56岁年龄组HPV感染率相对较高,与其他年龄组比较差异有统计学意义。结论妇科门诊患者HPV感染率较高,且以高危型HPV感染为主,其中最常见的感染亚型是HPV-16、HPV-52、HPV-58。不同年龄组HPV感染率差异有统计学意义,17~25岁和≥56岁年龄组HPV感染率相对较高。     

MALDI-TOF-MS技术检测宫颈癌中HPV感染状态

目的:本研究拟评MALDI-TOF-MS技术在HPV分型中的实际应用效果.方法:采用MALDI-TOF-MS技术检测河南安阳地区94例宫颈癌样本中HPV感染状态,并与普通PCR方法得到的结果相比较.结果:94例宫颈癌组织中HPV感染率为92.6%,共检测到10种常见的高危型别,以16为主,其次是58和18型.样本单一HPV型别感染为主,其中16型单一型别感染占6I.7%,见多种型别和组合的多重感染.与普通PCR检测结果比较发现两种方法样本HPV(不分型)检测一致率为95.7,样本HPV16型检测一致率为93.6%.结论:MALDI-TOF-MS技术能够精确、快速、高通量的对宫颈癌中HPV进行检测和分型.     

宫颈癌组织人乳头瘤病毒的荧光偏振基因分型

采用荧光偏振人乳头瘤病毒(human papillomavirus,HPV)分型新方法探讨了8种常见型别HPV在陕西宫颈癌患者中的流行情况。首先,用HPV GP5 ／GP6 通用引物PCR扩增65例早期宫颈癌(Ⅱa期内)和72例慢性宫颈炎病变组织DNA粗提物,继之将模板指导的末端延伸反应与荧光偏振检测技术结合(TDI-FP),用GP5 ／GP6 扩增区内的HPV6、11、16、18、31、33、35和58型特异性探针与PCR产物杂交后,荧光素标记的特异碱基(TAMRA-ddTTP或R110-ddGTP)在GP5 ／GP6 产物中相应的模板指导下,掺入延伸至相应探针末端,致使对应的TAMRA或R110 FP值升高,从而对扩增的HPV阳性产物进行HPV分型。65例宫颈癌患者中检出HPV57例,阳性率87．69％,72例慢性宫颈炎患者中检出HPV28例,阳性率38．89％,两组间HPV阳性率有显著性差异。宫颈癌与慢性宫颈炎患者中4种最常见的HPV型别分别是HPV 16(45．6％)、HPV 18(22．8％)、HPV 58(17．5％)、HPV 31(7．02％)和HPV 16(35．7％)、HPV 11(32．1％)、HPV 6(21．4％)、HPV 18(10．7％)。慢性宫颈炎患者中检出的HPV型别57．14％属高危型。HPV 16在两组中均最为多见。中国陕西宫颈疾病患者中HPV感染有其特点,世界范围内少见的HPV 58在陕西宫颈癌与慢性宫颈炎患者中均较为多见,在进行HPV新诊断方法及疫苗研制时应考虑到这种特点。     

宫颈癌患者人乳头瘤病毒(HPV)主要型别及其感染研究

本文探讨了江西省和广东省宫颈癌患者人乳头瘤病毒(Human papillomavirus,HPV)感染及其型别分布,分析了高危型HPV对各种宫颈病变的感染情况,为宫颈癌的早期发现和临床诊治提供科学依据。首先采用细胞学、HPV DNA检测(第二代杂交捕获法,HC2)、电子阴道镜和宫颈化学着色方法筛查宫颈癌患者,经病理镜检确诊,然后用GP PCR-SBT法对宫颈癌患者进行HPV基因分型。江西省溪口镇、古市镇及修水县城宫颈癌癌前病变发生率为5．7‰。HC2方法发现宫颈癌患者13种高危型HPV DNA阳性率为89．9％,宫颈上皮内瘤样病变的为84．8％,对照组为24．5％。采用GP PCR-SBT方法进行基因分型发现,江西省宫颈癌患者存在HPV16、58、31、33、18、66、6、11、56和81十种型别,其中HPV81型在国内外鲜有报道。据此提出生殖道高危型HPV感染是妇女宫颈癌发病的重要因素。并发现江西省宫颈癌高发区妇女高危型HPV感染率为24．5％。建立了HPV基因分型的方法,对HPV致宫颈病变的分子机制进行了分析。     

人乳头瘤病毒58型的系统地理学分析

人乳头瘤病毒58型（HPV58）是具有高度致癌危险性的HPV型别之一,在亚洲和非洲等地呈现出特殊的流行状况．由于全球HPV58数据分布零散,加之一些高发地区（如中国内地）数据缺乏,迄今为止尚未有对世界范围内HPV58地理分布的全面分析．本研究对中国内地妇女宫颈癌组织标本进行了HPV58检测,共获得14条HPV58-E6,L1基因序列．对GenBank收录的自1985年起分离的HPV58序列进行了系统地理学分析．结果表明,在上海、江苏和四川等地检出的HPV58-E6,L1序列均与以往来自中国香港、西安和日本等HPV58亚洲株同源．HPV58毒株可能由非洲西部起源,而中国内地及东南亚地区则在接受“根部”来源的变异株之后,成为毒株播散源“中转站”和新的策源地．HPV58型可能与HPV16,HPV18相同,也是宫颈癌发生和散播的主要HPV型别．     

南京部分地区女性宫颈HPV感染状况与分析

目的:探讨南京部分地区女性人乳头瘤病毒(HPV)感染的型别和年龄特征,为宫颈癌的防治提供参考资料.方法:采用人乳头状瘤病毒的基因扩增技术(HPV PCR)对2010年9月至2011年7月3382例就诊于南京妇幼保健院宫颈疾病科的女性进行HPV基因亚型检测,21种型别包括15种高危型和6种低危型,分析比较上述样品的感染程度、感染型别、感染年龄等.结果:HPV总阳性率为33.47％(1132/3382),21种型别均被检出,该地区女性阳性率最高的型别是HPV16,其他常见为58、52、CP8304、53、18、33等,多重感染率为9.58％.该地区女性HPV阳性率与各年龄组无显著差异(P＞0.1).结论:该地区女性HPV阳性率为33.47％,常见型别为HPV16、58、52、CP8304、53、18、33、68、31、39、11、6和66型;人群感染存在在＜20岁和＞70岁两个高峰.     

北京地区宫颈癌HPV16上游调控序列、E6、E7癌基因序列初步分析

为检测HPV16上游调控序列(Upstream regulatory region, URR)、E6、E7癌基因变异在北京地区宫颈癌患者癌组织中的分布特征, 探讨该地区宫颈癌发生同HPV16变异株间的相关性, 文章以提取的31例HPV16检测阳性宫颈癌组织DNA为模板, 设计针对性引物扩增URR、E6、E7 3个目的片段, PCR产物直接测序并通过GenBank对比分析变异和分支鉴定情况。在所分析的宫颈癌组织中, URR是突变频率最高的片段, 其次为E7, 最保守的序列为E6。共发现热突变位点8个, 分别为URR序列上G7521A(100%)、C7435G(96.77%)、C24T(45.16%)、A7729C(45.16%)、G7839A(45.16%); E6序列上T178G(41.94%); E7序列上A647G(45.16%)、T846C(45.16%)。HPV16分支分布频率最广的是As型(54.84%), 其次为E型(45.16%)。研究结果提示, HPV16URR序列上G7521A、A7729C、G7839A, E6序列上T178G、T350G, E7序列上A647G、G658A等位点的变异可能与病毒致癌潜能及宫颈癌的发生相关。北京地区宫颈癌患者中As和E型可能是两种最主要的HPV16分支, 这有可能会为HPV疫苗的研制和感染治疗提供有价值的信息。As型和E型病毒在不同年龄组和不同肿瘤分期组的患者中分布频率有差异, 这可能会为揭示宫颈癌年轻化趋势提供新的线索。     

Conservation of E6 and E7 regions of human papillomavirus types 16 and 18 present in cervical cancers

The E6 and E7 regions of human papillomavirus (HPV) type 16 were present in the DNA samples from cervical cancer cell lines, SKG-IIIa and SKG-IIIb, and those from cervical cancer tissues of three different patients. T601 cells, an NIH3T3 transformant obtained by transfection of DNA from a surgical specimen of a cervical cancer, also contained the E6 and E7 regions. The E6 region of HPV type 16 was expressed as mRNA in SKG-IIIa, SKG-IIIb and T601 cells. The E6 and E7 regions of HPV type 18 were present in the DNA samples from cervical cancer cell lines, SKG-I and SKG-II, and those from cervical cancer tissues of two different patients. SKG-I and SKG-II cells expressed the E6 region of HPV type 18 as mRNAs. These results strongly suggest that the E6 and E7 regions or the sequence surrounding these regions are important for maintaining malignant phenotype of cervical cancer cells.     

宫颈癌患者高危型人乳头瘤病毒感染 与免疫功能和癌基因表达的相关性

摘要：目的 探讨宫颈癌患者高危型人乳头瘤病毒（HPV）感染的检测及其与免疫功能和癌基因表达的相关性。方法 选择2017年9月至2018年12月在我院接受手术治疗的118例原发性宫颈癌患者为宫颈癌组,根据高危型HPV感染情况进一步分为高危型HPV组（89例）和非高危型HPV组（29例）。选择同期在我院行子宫全切术的57例子宫肌瘤患者为子宫肌瘤组。对比各组患者外周血T淋巴细胞亚群分布情况,病灶组织中原癌基因（E6/E7、c-Met、SALL4、PGRN）和抑癌基因（p53、pRb、PTEN、LKB1）表达量的差异。结果 宫颈癌组患者病灶组织中高危型HPV感染率显著高于子宫肌瘤组（75.42% vs 22.81%,2=43.764,P＜0.05）。高危型HPV组患者外周血中CD4+ T淋巴细胞比例、CD4+T/CD8+ T比值低于非高危型HPV组,CD8+ T淋巴细胞比例高于非高危型HPV组（均P＜0.05）。高危型HPV组患者病灶组织中E6/E7、c-Met、SALL4、PGRN mRNA的表达量高于非高危型HPV组,而p53、pRb、PTEN、LKB1 mRNA的表达量低于非高危型HPV组（均P＜0.05）。结论 高危型HPV感染可导致宫颈癌患者细胞免疫功能下降及肿瘤恶性程度增加,可能是导致病情恶化、治疗效果不佳的危险因素之一。     

Prevalence and risk factors of Human Papillomavirus (HPV) infection in southern Chinese women - a population-based study

Background

Persistent high-risk type Human papillomavirus (HPV) infection is recognized as a necessary cause of cervical cancer. This study aimed to compare the HPV prevalence and risk factors between women residing in Hong Kong (HK) and Guangzhou (GZ) region of China.

Methodology/Principal Findings

A total of 1,570 and 1,369 women were recruited from HK and GZ, respectively. The cytology samples were collected and tested for HPV infection. The overall and type-specific HPV prevalence and the potential risk factors for acquisition of HPV infection were studied. Women with normal cytology in the GZ cohort had significantly higher HPV prevalence (10%) than those in the HK cohort (6.2%, p<0.001). The patterns of the age-specific HPV prevalence were also different between the two cohorts. In the HK cohort, women at the age of 20–29 years old had the highest prevalence and a second peak was observed in the age of ≥60 years old. In the GZ cohort, the highest HPV prevalence was also observed in 20–29 years old but declined as the age increased and a second peak was not seen. HPV16 and HPV52 were the most common high-risk types found in the HK and GZ cohorts, respectively. Age was the most consistently observed independent risk factor for HPV infection in the HK, while the number of sexual partners had association in the GZ cohort.

Conclusions/Significance

Our study provides the current status and the epidemiological characteristics of HPV prevalence in Southern Chinese women. The results strongly suggested that population education and the effective cervical cancer screening would be vital in the prevention of cervical cancer.     

Human papillomavirus type 16 E7 peptide-directed CD8+ T cells from patients with cervical cancer are cross-reactive with the coronavirus NS2 protein

Human papillomavirus type 16 (HPV16) E6 and E7 oncoproteins are required for cellular transformation and represent candidate targets for HPV-specific and major histocompatibility complex class I-restricted CD8(+)-T-cell responses in patients with cervical cancer. Recent evidence suggests that cross-reactivity represents the inherent nature of the T-cell repertoire. We identified HLA-A2 binding HPV16 E7 variant peptides from human, bacterial, or viral origin which are able to drive CD8(+)-T-cell responses directed against wild-type HPV16 E7 amino acid 11 to 19/20 (E7(11-19/20)) epitope YMLDLQPET(T) in vitro. CD8(+) T cells reacting to the HLA-A2-presented peptide from HPV16 E7(11-19(20)) recognized also the HLA-A2 binding peptide TMLDIQPED (amino acids 52 to 60) from the human coronavirus OC43 NS2 gene product. Establishment of coronavirus NS2-specific, HLA-A2-restricted CD8(+)-T-cell clones and ex vivo analysis of HPV16 E7 specific T cells obtained by HLA-A2 tetramer-guided sorting from PBL or tumor-infiltrating lymphocytes obtained from patients with cervical cancer showed that cross-reactivity with HPV16 E7(11-19(20)) and coronavirus NS2(52-60) represents a common feature of this antiviral immune response defined by cytokine production. Zero of 10 patients with carcinoma in situ neoplasia and 3 of 18 patients with cervical cancer showed > or =0.1% HPV16 E7-reactive T cells in CD8(+) peripheral blood lymphocytes. In vivo priming with HPV16 was confirmed in patients with cervical cancer or preinvasive HPV16-positive lesions using HLA-A2 tetramer complexes loaded with the E6-derived epitope KLPQLCTEL. In contrast, we could not detect E6-reactive T cells in healthy individuals. These data imply that the measurement of the HPV16 E7(11-19(20)) CD8(+)-T-cell response may reflect cross-reactivity with a common pathogen and that variant peptides may be employed to drive an effective cellular immune response against HPV.     

Antigen processing defects in cervical carcinomas limit the presentation of a CTL epitope from human papillomavirus 16 E6.

Human papillomavirus (HPV) infection, particularly type 16, is causally associated with the development of cervical cancer. The E6 and E7 proteins of HPV are constitutively expressed in cervical carcinoma cells making them attractive targets for CTL-based immunotherapy. However, few studies have addressed whether cervical carcinomas can process and present HPV E6/E7-derived Ags for recognition by CTL. We generated HLA-A*0201-restricted CTL clones against HPV16 E6(29-38) that recognized HPV16 E6 Ags transfected into B lymphoblastoid cells. These CTL were unable to recognize HLA-A*0201(+) HPV16 E6(+) cervical carcinoma cell lines even when the level of endogenous HPV16 E6 in these cells was increased by transfection. This defect in presentation of HPV16 E6(29-38) correlated with low level expression of HLA class I, proteasome subunits low molecular mass protein 2 and 7, and the transporter proteins TAP1 and TAP2 in the cervical carcinoma cell lines. The expression of all of these proteins could be up-regulated by IFN-gamma, but this was insufficient for CTL recognition unless the level of HPV16 E6 Ag was also increased by transfection. CTL recognition of the HPV16 E6(29-38) epitope in 721.174 B cells was dependent on TAP expression but independent of immunoproteasome expression. Collectively, these findings suggest that presentation of the HPV16 E6(29-38) epitope in cervical carcinoma cell lines is limited both by the level of TAP expression and by the low level or availability of the source HPV E6 oncoprotein. These observations place constraints on the use of this, and potentially other, HPV-derived CTL epitopes for the immunotherapy of cervical cancer.     

E6 and E7 fusion immunoglobulin from human papilloma virus 16 induces dendritic cell maturation and antigen specific activation of T helper 1 response

Human papilloma virus (HPV) 16 causes cervical cancer. Induction of oncogenesis by HPV 16 is primarily dependent on the function of E6 and E7 proteins, which inactivate the function of p53 and pRB, respectively. Thus, blocking the activity of the E6 and E7 proteins from HPV 16 is critical to inhibiting oncogenesis during infection. We have expressed and purified soluble HPV 16 E6 and E7 fusion immunoglobulin (Ig), which were combined with the constant region of an Ig heavy chain, in a mammalian system. To assess whether soluble E6 and E7 fusion Igs induce effective cellular immune responses, immature dendritic cells (DCs) were treated with these fusion proteins. Soluble E6 and E7 fusion Igs effectively induced maturation of DCs. Furthermore, immunization with soluble E6 and E7 fusion Igs in mice resulted in antigen-specific activation of T helper 1 (Th1) cells. This is the first comprehensive study to show the molecular basis of how soluble HPV 16 E6 or E7 fusion Igs induces Th1 responses through the maturation of DCs. In addition, we show that DC therapy using soluble HPV E6 and E7 fusion Igs may be a valuable tool for controlling the progress of cervical cancer.     

Human papillomavirus (HPV) 16 E6 sensitizes cells to atractyloside-induced apoptosis: Role of p53, ICE-like proteases and the mitochondrial permeability transition

Infection of cervical epithelial cells with certain high risk HPV genotypes is thought to play an etiologic role in the development of cervical cancer. In particular, HPV type 16 and 18 early protein 6 (E6) is thought to contribute to epithelial transformation by binding to the tumor suppressor protein p53, targeting it for rapid proteolysis, resulting in loss of its cell cycle arrest and apoptosis-inducing activities. Recent data indicate that factors responsible for triggering apoptosis reside in the cytoplasm of cells, and not in the nucleus. In particular, the findings that mitochondria are required in certain cell-free models for induction of apoptosis and that bcl-2 is localized to mitochondria have focused attention on the role of the mitochondrial membrane permeability transition (MPT) in apoptosis. Here we present data to indicate that HPV 16 E6 expression sensitizes cells to MPT-induced apoptosis. We also report that HPV 16 E6 sensitization of cells to MPT-induced apoptosis occurs only in the presence of wildtype (wt) p53 expression. The extent of apoptosis induced by atractyloside (an inducer of the MPT) in normal, temperature-sensitive (ts) p53, and HPV-16 E6 transfected J2-3T3 cells, and the HPV expressing cervical carcinoma cell lines SiHa, Hela and CaSki was determined. C33A cells, which express mutant p53 but not HPV, were also exposed to atractyloside in the presence or absence of HPV 16 E6 expression. Dose-dependent apoptosis induced by atractyloside in normal J2-3T3 cells and cervical carcinoma cells was measured by loss of cell viability, nuclear fragmentation and DNA laddering. The sensitivity of cells to atractyloside-induced apoptosis was found to be: HPV 16 E6-J2-3T3 > CaSki > normal-J2-3T3 cells ≈ ts p53-J2-3T3 ≈ vector-J2-3T3 cells > Hela > SiHa > C33A ≈ C33A 16 E6. Cyclosporin A (CsA), an inhibitor of the MPT, and ICE-I, a protease inhibitor, provided protection against atractyloside-induced apoptosis. These findings indicate that: 1) high risk HPV 16 E6 protein is capable of sensitizing cells to apoptosis; 2) HPV 16 E6 sensitization of cells to atractyloside-induced apoptosis occurs in a p53-dependent fashion; 3) the target of HPV 16 E6 sensitization of cells to atractyloside-induced apoptosis is the mitochondria; and 4) HPV 16 E6 sensitization of cells to atroctycoside-induced apoptosis involves an ICE-like protease-sensitive mechanism, regulating the onset of the MPT. These findings constitute the first evidence that mitochondria play a role in HPV 16 E6 modulation of apoptosis. J. Cell. Biochem. 66:245-255. © 1997 Wiley-Liss, Inc.