The expression of P53 protein and infection of human papilloma virus in conjunctival and eyelid neoplasms

Human papilloma virus (HPV) infection and loss of P53 function have been identified as frequent events in various human tumors. The aim of this study was to evaluate P53 protein expression and to detect HPV in the tissue samples of 45 benign (papillomas) and 38 malignant conjunctival and eyelid lesions (27 basal cell carcinomas and 11 squamous cell carcinomas). We also looked for eventual relationships between P53 expression and clinicopathological features such as age, histological type of tumor, grading and staging. HPV infection was detected using the PCR-RFLP method. Specific primers were engaged and PCR products of HPV 16, 18, and 33, underwent enzymatic digestion at 37 degrees C. We revealed P53 protein expression in 30 out of 45 (66.6%) squamous cell papillomas. In the SCC and BCC groups, P53 was present in 31 out of 38 carcinomas and there was a statistically significant correlation between histological type of tumor and P53 protein expression. Malignant type HPV 16 and 18 were detected in three squamous cell papillomas, two BCCs and one SCC. However, we observed P53 protein expression in only two HPV-positive papillomas and one infiltrative type of BCC. P53 is probably involved in the development of conjunctival and eyelid tumors due to its high rate of presence in both benign and malignant neoplasms of these organs. HPV seems to occur rarely. In some cases its role in the pathogenesis of conjunctival and eyelid tumorigenesis should be considered as auxiliary.     

Detection of human papillomavirus type 6/11 DNA in conjunctival papillomas by in situ hybridization with radioactive probes

Twenty-three conjunctival papillomas and 28 conjunctival dysplasias were examined for human papillomavirus (HPV)-DNA sequences by in situ hybridization with nick-translated 35S-labeled HPV probes. Adjacent paraffin sections were hybridized with HPV type 2, 6, 16, and 18 probes at Tm - 17 degrees C. Fifteen tissues, all papillomas, displayed positive hybridization with the HPV-6 probe. Infection with HPV-6 (or the closely related HPV-11) appeared to be responsible for most of the conjunctival papillomas of children and young adults. The presence of genital tract HPV-6 in these lesions suggests that some of the infections were acquired during passage through an infected birth canal. The lack of hybridization in adult conjunctival dysplasias indicates either that HPVs are not associated with this condition or that the probes and the technique utilized were not adequate for demonstration of this association.     

Sinonasal Schneiderian papillomas: human papillomavirus typing by polymerase chain reaction.

A series of 35 formalin-fixed, paraffin-embedded Schneiderian papillomas (24 inverted, nine cylindrical cell type, and two fungiform) of the nasal cavity were evaluated for the presence of human papillomavirus (HPV) types 6b/11, 16, and 18 DNA sequences using both a highly sensitive and specific modification of the polymerase chain reaction technique and conventional in situ hybridization. The HPV gene sequences (E6-E7 portions) were not detected in any of the 24 inverted or nine cylindrical cell papillomas. One of the fungiform papillomas was positive for HPV 6b/11. We conclude: (a) the origin of most Schneiderian sinonasal papillomas is not associated with HPV infection of these common types, and (b) fungiform papilloma is a distinctive clinicopathologic subtype of Schneiderian papilloma that may be HPV-related.     

High prevalence of human papillomavirus infection and possible association with betel quid chewing and smoking in oral epidermoid carcinomas in Taiwan

Seventeen oral epidermoid carcinomas, three oral papillomas, and 17 normal gingival tissues were tested for the presence of human papillomavirus (HPV) types 6, 11, 16, and 18 sequences by Southern blot hybridization. Episomal HPV-16 sequences in various amounts were detected in 76.4% of the oral carcinomas and in all three cases of papilloma. However, only one of the 17 normal tissues was HPV positive with an unknown type. None of the samples contained HPV-6, -11, or -18 sequences. Examination of the habits of the patients showed that 59% of the patients were betel quid chewers and 82% were smokers. Thus, the concurrent incidence of HPV infection and betal quid chewing and/or smoking habits in oral carcinoma patients observed in Taiwan is consistent with the view that both viral and chemical factors may be involved in the process of carcinogenesis.     

HPV type 16 in conjunctival and junctional papilloma, dysplasia, and squamous cell carcinoma.

AIMS--To clarify the role of human papillomavirus (HPV) infection in the development of papilloma, dysplasia, squamous cell carcinoma, and basal cell epithelioma arising from the eyelids, including the tunica conjunctiva palpebrum (conjunctiva), its junction to epidemis of eyelid skin (junction), and eyelid skin. METHODS--Sixteen cases of papilloma, four of dysplasia, four of squamous cell carcinoma, and 12 of basal cell epithelioma were examined using formalin fixed and paraffin embedded samples. Detection of HPV-DNA was performed by PCR-RFLP and in situ hybridisation (ISH) methods. RESULTS--HPV-16 was detected in 12/16 papillomas (75%), 2/4 dysplasias (50%), and 1/4 squamous cell carcinomas (25%) but in none of the basal cell epitheliomas. No other HPV subtypes were found. ISH assay showed positive signals in only two cases of dysplasia and squamous cell carcinoma. The mean age of HPV-16 positive dysplasia and squamous cell carcinoma cases (81.7 years) was significantly higher than that of HPV-16 positive papilloma cases (p < 0.01). CONCLUSIONS--Based on the presence of HPV-16 in both benign and malignant lesions and the age distribution, it seems likely that HPV-16 alone may be incapable of causing development of conjunctival and junctional dysplasia and squamous cell carcinoma, and that any correlation between the papilloma-squamous cell carcinoma sequence and HPV infection may be due to rare events.     

Sinonasal papillomas and human papillomavirus: human papillomavirus 11 detected in fungiform Schneiderian papillomas by in situ hybridization and the polymerase chain reaction.

A series of 19 paraffin-embedded sinonasal papillomas (four squamous papillomas, three fungiform papillomas, nine inverted papillomas, and three cylindrical cell papillomas) were investigated for evidence of human papillomavirus (HPV) infection using immunohistochemistry (polyclonal antibody to HPV capsid antigen), in situ hybridization (DNA probes for HPV 6/11, 16/18, and 31/33/35), and the polymerase chain reaction (primers and probes for HPV 6, 11, 16, 18, and 33). All three fungiform papillomas were positive by all three techniques: immunohistochemistry, in situ hybridization for HPV 6/11, and the polymerase chain reaction for HPV 11. None of the other lesions contained detectable HPV using the specific probes included in this study. These results support the continued classification of fungiform papilloma as a distinctive variant of schneiderian papilloma characterized by a predominantly exophytic growth pattern and an association with HPV 11.     

Prevalence of human papillomavirus in sinonasal papillomas: a study using polymerase chain reaction and in situ hybridization.

Inverted and fungiform papillomas of the sinonasal cavity share a common origin from the Schneiderian membrane, but they differ widely in their rates of recurrence and progression to carcinoma. To determine the role of human papillomavirus in the etiology of these lesions, 15 inverted papillomas, five fungiform papillomas, and two squamous cell carcinomas associated with inverted papilloma were examined for the presence of HPV by in situ hybridization (ISH) and polymerase chain reaction (PCR). ISH was carried out on formalin-fixed, paraffin-embedded material using HPV types 6/11, 16/18, and 31/33/35 DNA probes. Tissue DNA was amplified by PCR with HPV L1 consensus primers, and the product was detected by gel electrophoresis, Southern blotting, and hybridization with type specific probes (HPV types 6/11, 16, 18). Three of 15 inverted papillomas and two of five fungiform papillomas were positive for HPV 6/11 by ISH, whereas PCR detected HPV 6/11 sequences in two of 15 inverted and three of five fungiform papillomas. Biopsies from two patients who had serial resections contained HPV 6/11 in the original lesions and all recurrences. No HPV was detected in the carcinomas by ISH, whereas PCR detected HPV 16 in one carcinoma. These findings confirm the presence of HPV DNA sequences in both inverted and fungiform sinonasal papillomas as well as in an associated squamous carcinoma. This would suggest a role for HPV in the pathogenesis of Schneiderian membrane lesions. Furthermore, our data indicate that ISH and PCR are equally sensitive in detecting HPV in sinonasal papillomas.     

Detection of human papillomavirus (HPV) DNA type 6/11 in a conjunctival papilloma by in situ hybridization with biotinylated probes.

Four cases of conjunctival papilloma in two different patients were examined by in situ hybridization for HPV DNA type 6/11, 16/18 and 31/33/51. Formalin-fixed and paraffin-embedded tissues were hybridized by biotinylated probes. One tumor and one of its recurrences showed nuclear positivity for HPV 6/11 in the superficial cells of the epithelium. The results suggest that HPV type 6/11 may be etiologic agent of conjunctival papillomas. The benign behavior of these neoplasms may be related to the etiologic role of this type of HPV.     

High frequency of human papillomavirus 6/11, 16, and 18 infections in precancerous lesions and squamous cell carcinoma of the conjunctiva in subtropical Tanzania

Dysplastic lesions and epithelial neoplasms of the conjunctiva account for approximately 2% of all malignant tumors in subtropical Tanzania. We examined the pathophysiologic role of human papillomavirus (HPV) in the development of conjunctival carcinoma in subtropical Tanzania, which has a high HPV prevalence. Tissue samples from 14 patients were obtained from the cancer registry archives at the medical center of the university in Dar es Salaam, Tanzania. A highly sensitive nonradioactive in situ hybridization technique (ImmunoMax) was applied to paraffin-embedded tissue samples to identify HPV DNA in conjunctival epithelial dysplasia and epithelial neoplasms. In each case, conventional morphologic evaluation revealed a transitional lesion extending from koilocytic dysplasia to severe dysplasia or invasive squamous cell carcinoma. Highly specific, morphologically easily distinguishable labeling of HPV-6/11, HPV-16, and HPV-18 was found in most cases. Coinfections were observed frequently. The signals showed varying intensities and different patterns of distribution. In general, higher signal intensity was found in dysplasia grades 1 and 2 and in well-differentiated areas of the invasive component of conjunctival carcinoma compared with less differentiated areas. This observation underlines the central role of HPV-16 and HPV-18 in the oncogenesis of conjunctival cancers in subtropical Tanzania.     

Human papillomavirus 6, 11, and 16 in laryngeal papillomas.

Twenty-seven cases of benign laryngeal papillomas, both single and multiple variants, were analysed for human papillomavirus (HPV) by DNA slot-blot hybridization chiefly to determine the pattern of infection in Hong Kong Chinese. DNA was extracted from paraffin blocks of formalin-fixed tissue and probed separately for HPV 6, 11, 16, and 18. Sixteen cases (59 per cent) showed the presence of at least one of these four HPV genomes. Thirteen cases (48 per cent) were positive for HPV 11 only. Three other cases (11 per cent) showed triple positivity for HPV 6, 11, and 16. None were positive for HPV 18. The predominance of HPV 11 infection contrasts with other series which have shown either an almost equal distribution of HPV 6 and 11 or a predominance of HPV 6. The finding of HPV 16 in three cases was unexpected. Using the polymerase chain reaction (PCR) with primers complementary to the upstream regulatory region of the HPV 16 viral DNA, the presence of HPV 16 genome was confirmed in all three cases. As the number of HPV 16-positive cases in this study is small, analysis of more cases using fresh biopsy material and a wider range of HPV type-specific PCR primers is warranted to determine the relative incidence of HPV subtypes in these benign laryngeal papillomas.     

Etiological role of human papillomavirus infection for inverted papilloma of the bladder

The status of human papillomavirus (HPV) infection in urothelial inverted papilloma was examined in the present study. Formalin-fixed and paraffin-embedded tissues from eight cases of inverted papilloma of the bladder were studied. The presence of HPV-DNA was examined by modified GP5/6+PCR using archival tissue sections by microdissection. HPV genotype was determined with a Hybri-Max HPV genotyping kit. Immunohistochemical analysis for p16-INK4a, mcm7, HPV-E4, and L1, and in situ hybridization for the HPV genome were performed. HPV was detected in seven of eight cases (87.5%) of inverted papilloma. Three cases were diagnosed as inverted papilloma with atypia, while the remaining five were typical cases. HPV-18 was detected in two cases, including one inverted papilloma with atypia, and HPV-16 was detected in four cases, including one inverted papilloma with atypia. Multiple HPV type infection was detected in one typical case and one atypical case. High-risk HPV was present in all HPV-positive cases. Cellular proteins, p16-INK4a and mcm7, which are surrogate markers for HPV-E7 expression, were detected in all HPV-positive cases, and their levels were higher in inverted papilloma with atypia than in typical cases. In contrast, HPV-E4 and L1, which are markers for HPV propagation, were observed in some parts of the typical inverted papilloma tissue. High-risk HPV infection may be one of the causes of urothelial inverted papilloma, and inverted papilloma with atypia may have malignant potential.     

A general primer pair for amplification and detection of genital human papillomavirus types.

A general primer pair localized in the E7 and E1 regions was identified and used for the detection of genital human papillomaviruses (HPVs). The genital HPV types 6b, 11, 16, 18, 31 and 33 were amplified and detected by the polymerase chain reaction (PCR) performed at a high stringency annealing temperature (60 degrees C). HPV-2, -3, -7, -13 and -30 were amplified only at lower temperatures. Twelve biopsies from women with invasive cancer in the cervix were analysed with the general primer pair. The amplification product specific for the general primer pair was detected in 11 of the 12 biopsies. The eleven HPV DNA positive specimens were shown to contain HPV-6b, HPV-16 and/or HPV-18 by Southern blot hybridization of the PCR products. The general primers were also used for analysis of 57 cervical scrapes from women with normal cytology, condyloma or CIN. By ethidium bromide staining after agarose gel electrophoresis we could detect 21 positives. Slot-blot analysis of the amplification products from all 57 scrapes confirmed the specificity of the 21 positives and revealed 5 additional positives. Among the 57 scrapes, 15/21 CIN scrapes, 10/21 condyloma scrapes and 1/15 normal scrapes contained HPV DNA. Eight different HPV types were detected. The general primer pair from the E7/E1 region is thus a powerful tool for the detection of HPV in clinical samples. The amplimer obtained offers a possibility for further typing by slot-blot hybridization using HPV-type specific probes.     

The characteristics of human papillomavirus DNA in head and neck cancers and papillomas

AIM: To determine the prevalence, type, physical state, and viral load of human papillomavirus (HPV) DNA in cases of head and neck cancer and recurrent respiratory papillomatosis (RRP). METHODS: The prevalence and type of HPV DNA was determined in 27 fresh frozen tissue specimens from patients with head and neck cancers and 16 specimens from 10 patients with RRP by MY09/MY11 and GP5+/GP6+ nested polymerase chain reaction (PCR) and subsequent restriction enzyme cleavage. The physical state of HPV DNA was analysed by E1, E2, and E1E2 specific PCRs and Southern blot hybridisation (SBH). RESULTS: HPV DNA was detected in 13 of 27 cancers and 10 of 10 papillomas. Both low risk HPV-6 and HPV-11 and high risk HPV-16 were present in cancers in low copy numbers, whereas papillomas exclusively harboured low risk HPV-6 and HPV-11. E1E2 PCRs failed to determine the physical state of HPV in cancers except one case where HPV-6 DNA was integrated. In contrast to cancers, all papillomas showed the episomal state of HPV DNA and a relatively higher viral load. CONCLUSIONS: Based on the prevalence, type, physical state, and copy number of HPV DNA, cancers and papillomas tend to show a different HPV DNA profile. The 100% positivity rate of low risk HPV types confirms the role of HPV-6 and HPV-11 in the aetiology of RRP.     

Epidemiology and risk factors for human papillomavirus infection in a diverse sample of low-income young women

BackgroundTwo HPV vaccines prevent infection with HPV-16 and HPV-18, high-risk (cancer-associated) HPV types which together cause approximately 70% of cervical cancers; one vaccine also prevents HPV-6 and HPV-11, which together cause approximately 90% of anogenital warts. Defining type-specific HPV epidemiology in sexually experienced women will help estimate the potential clinical benefits of vaccinating this population.ObjectivesTo examine HPV epidemiology in a diverse sample of sexually experienced women, and to determine factors associated with high-risk HPV and vaccine-type HPV (HPV-6, HPV-11, HPV-16 and HPV-18).Study designCross-sectional study of 13–26-year-old women (N = 409) who completed a questionnaire and provided a cervicovaginal swab. Swabs were genotyped for HPV using PCR amplification. Logistic regression models were used to determine whether participant characteristics, knowledge, and behaviors were associated with high-risk and vaccine-type HPV.ResultsMost women (68.4%) were positive for ≥1 HPV type, 59.5% were positive for ≥1 high-risk type, 33.1% were positive for ≥1 vaccine-type HPV, and 3.5% were positive for both HPV-16 and HPV-18: none was positive for all four vaccine types. In adjusted logistic regression models, Black race (OR 2.03, 95% CI 1.21–3.41) and lifetime number of male sexual partners (OR 4.79, 95% CI 2.04–11.23 for ≥10 partner vs. ≤1 partner) were independently associated with high-risk HPV infection.ConclusionsHPV prevalence was very high in this sample of sexually active young women, but <5% were positive for both HPV-16 and HPV-18, suggesting that vaccination could be beneficial for many individual women who are sexually experienced.     

用聚合酶链反应检测食管癌组织中人乳头瘤病毒DNA

应用聚合酶链反应（ＰＣＲ）技术对汕头市区６８例食管癌的石蜡包埋标本进行人乳头瘤病毒（ＨＰＶ）ＤＮＡ序列检测，结果显示，ＨＰＶＤＮＡ总阳性率为６６．１８％（４５／６８），检出型别主要为ＨＰＶ６、１１、１６，检出率分别为２７．９４％、３６．７６％和２７．９４％，经统计学处理三型间无显著性差异；ＨＰＶ－１８及未定型别各占８．８２％。值得注意的是ＨＰＶ感染中多重感染占阳性病例的５３．３３％（２４／４５）。初步结果表明，汕头市食管癌高发区有较高的ＨＰＶ感染率，此与食管癌的发生，可能有密切关系。     

广州东部妇女宫颈人乳头瘤病毒16型感染及基因分析

目的 研究广州东部妇女中人乳头瘤病毒16型(HPV-16)宫颈感染分布,分析其早基因E6/E7的多态性,分析L1和E6基因定量与病程的关系.方法 通过导流杂交基因芯片技术检测宫颈脱落细胞的HPV-16感染;通过特异性扩增获取病毒早基因E6/E7序列,克隆测序并进行多态性分析;荧光定量PCR技术对E6基因和L1基因进行定量分析.结果 806例宫颈脱落细胞样本中HPV-16感染阳性36例(4.5%),其中18例(50.0%)宫颈细胞发生高度以上病变;7例(4例低度或以下病变,3例高度以上病变或浸润癌)阳性标本得到E6/g7序列有15个位点分别出现变异;高度病变组(A组,11例)与低度或以下病变组(B组,14例)的L1基因和E6基因定量数据对数值均有显著差异(P<0.05),但L1/E6比值差异无统计学意义(P=0.19).结论 本地区在17～62岁妇女中HPV-16感染阳性发生率约4.5%,50.0%发生高度以上宫颈病变,本研究显示病毒基因拷贝数与宫颈病变程度可能有关,L1/E6比值未能提示病毒整合的发生.     

A prospective study of antibody responses to defined epitopes of human papillomavirus (HPV) type 16 in relationship to genital and anorectal presence of HPV DNA.

The aim of this study was to investigate whether antibody responses against synthetic peptides derived from genital human papillomavirus (HPV) proteins are associated with laboratory-proven genital and anorectal HPV infection. In this study, 158 heterosexual patients (110 women and 48 men) were followed prospectively. At each visit we collected serum samples as well as specimens from several sites in the anogenital area for detection of HPV type 6/11 (HPV-6/11), -16, -18, and -33 DNAs by PCR. Immunoglobulin A (IgA) and IgG responses against disrupted bovine papilloma virions and eight different synthetic peptides derived from HPV-6/11, -16, and -18 were determined for serum samples from the first and the last visits. The subjects attended the Municipal Sexually Transmitted Disease Clinic in Amsterdam, The Netherlands, two to seven times (mean, four times) at approximately 4-month intervals. Women were monitored over a period of 155 person-years, and men were monitored over 65 person-years. The magnitudes of the IgA responses against HPV-16 late protein epitopes L1:13, L1:31, and L2:49 were significantly higher in the sera from the last visit among the currently HPV DNA-positive participants than in HPV DNA-negative persons (P = 0.02). When the persons positive for any HPV type at any time during the follow-up period were compared with those who were negative at all times during the follow-up period, we also found a significant elevation of IgA responses against L1:31 and L2:49 (P = 0.04 and 0.01, respectively).(ABSTRACT TRUNCATED AT 250 WORDS)     

Squamous papillary neoplasia of the adult upper aerodigestive tract

Selected papillary squamous tumors of the upper aerodigestive tract (UADT) mucosa in adult patients do not have well-defined histologic criteria and the clinical behavior is poorly understood. To better characterize this spectrum of neoplasms, UADT papillary neoplasms were evaluated by routine histology, determination of cellular DNA content using Feulgen-stained tissue sections, and the typing of human papillomavirus (HPV) by in situ hybridization. Solitary papillomas were studied in two patients; there was no recurrence in either case, both had normal DNA content, and one was typed as HPV-6 while the other was typed as HPV-11. Seven adult patients with recurrent papillomatosis and at least one biopsy with dysplasia/atypia were identified (mean age at diagnosis, 13.3 years; mean age at last contact, 42.7 years). Six of seven patients had abnormal DNA cellular content in foci of epithelial atypia. In all biopsies evaluated, the papillomas of the seven patients were consistently typed as either HPV-6 or HPV-11. Six patients with malignant papillary neoplasms also had abnormal DNA cellular content, but none revealed evidence of HPV type 6, 11, 16, or 18 by in situ hybridization of tissue sections. In many of the recurrent papillomas, the degree of epithelial atypia encountered was pronounced and was commonly misdiagnosed as carcinoma in situ or papillary carcinoma. The aneuploid DNA content of these foci of atypia reflected the abnormal cellular appearance and partially explained the overdiagnosis of malignancy. However, none of the seven patients were treated for malignant disease and none progressed to invasive carcinoma, with an average follow-up period of almost 30 years. We conclude that histologic and cytologic atypia in HPV-containing papillomatosis may be appreciable. The aneuploid DNA content may represent premalignant conditions and the patient may be at an increased risk for the subsequent development of squamous cancer. However, none of the seven patients with recurrent papillomatosis developed any evidence of malignancy. In addition, none of the patients with papillary carcinomas had previous recurrent papillomatosis.     

Papilloma virus infection in pretumor and tumor masses of the larynx

There has been recent evidence suggesting that papilloma virus infection is of no small importance in the pathogenesis of tumors of the head and neck. This study has involved a screening of the mucosa having laryngeal pretumors and tumors for papilloma virus infection by polymerase chain reaction and immunohistochemistry. Specimens were studied in 130 patients. All the patients with laryngeal papillomatosis were found to have human papilloma virus (HPV) types 6 and 11. HPV-18 was detected in 6 (13%) of 48 cases of laryngeal cancer and in 4 (9%) of 46 cases of laryngeal mucosal precancer. Mixed HPV infection of 2-3 different types was revealed in 6 (20%) of 30 HPV-positive patients. Such a wide survey of laryngeal pretumors and tumors in the Russian population for virus infection has been conducted for the first time.     

High prevalence of high-risk oncogenic human papillomaviruses harboring atypical distribution in women of childbearing age living in Libreville, Gabon

The extent of human papillomavirus (HPV) genital shedding and type-specific diversity were evaluated in 354 consecutive women of childbearing age living in Libreville, Gabon. Detection of HPV DNA was performed by PCR using the MY09/MY11 primer set on DNA extracted from endocervical swabs. All PCR positive specimens were subjected to direct sequencing and HPV genotypes were identified on the basis of >95% sequence homology in the L1 region. Reverse line blot hybridization assay was used when a genotype could not be resolved by sequencing alone. HPV DNA was detected in 163 (46%) women, all clinically asymptomatic for HPV-related lesions. The highest prevalence of genital HPV detection (45%) was in the age group from 22 to 29 years. A total of 90 women (55%) harbored high-risk (HR) genotypes, with the most common being HPV-53 (19; 12%), HPV-58 (17; 11%), and HPV-16 (16; 10%). Low-risk genotypes were found in 36 (22%) women with HPV-54 and HPV-70 being the most frequently detected (17; 11% and 10; 6%, respectively). Finally 37 women (23%) tested positive for genotypes of unknown oncogenic risk, the most common in this category being HPV-83 (20; 12%). Multiple infections were detected in 35 (21%) women. By multivariate analysis, HPV genital shedding was significantly associated with young age (OR: 0.34; P < 0.007). The multivalent vaccine currently available against cervical carcinomas, is only active against HPV-16 and HPV-18, and will thus have a low impact in this setting.