In situ DNA hybridization analysis of oral papillomas, leukoplakias, and carcinomas for human papillomavirus.

Twenty-one papillomas, 23 ordinary benign keratoses, 13 smokeless tobacco keratoses, 10 verrucous hyperplasias, 10 verrucous carcinomas, 17 squamous cell carcinomas, 3 epithelial dysplasias, and 6 lichen planus lesions were evaluated for human papillomavirus (HPV) types 6/11, 16/18, and 31/33/35, with biotinylated double-stranded DNA probes by in situ hybridization. Sixty-two percent (13/21) of oral squamous papillomas were positive for HPV DNA. HPV DNA types 6 and 11 demonstrated the strongest reactivity. Of the 13 cases, 10 also showed some reactivity with HPV-16/18 and -31/33/35. None of the cases of keratoses, epithelial dysplasia, squamous cell carcinoma, verrucous hyperplasia, verrucous carcinoma, or lichen planus were positive for HPV DNA. This study confirms the consistent and frequent finding of HPV DNA in oral squamous cell papillomas and the inconsistency of being able to identify HPV DNA in keratotic, premalignant, or cancerous lesions of the oral mucous membranes.     

Oral squamous papillomas: detection of HPV DNA by in situ hybridization

Oral squamous papillomas were segregated from other papillary lesions on the basis of histopathologic features. Twenty representative papillomas were evaluated for the presence of papillomavirus genus-specific antigen with the use of an immunoperoxidase technique. These same tumors were analyzed for human papillomavirus (HPV) types 2, 4, 6, and 11 with biotinylated full-length double-stranded DNA probes by in situ hybridization. Only one case exhibited papillomavirus antigen reactivity. Alternatively, seven of twenty cases (35%) yielded positive results for HPV 6 or 11 DNA; one papilloma exhibited a dual infection with both HPV 2 and 6 when assayed under conditions of high-stringency hybridization. It is concluded that some oral squamous papillomas harbor HPV genotypes akin to those encountered in genital tract condylomas. Viral DNA can be detected in the absence of capsid antigen immunoreactivity, thereby obviating the use of antigen detection assays for determining the presence or absence of virus.     

The prevalence and distribution of human papillomavirus genotypes in oral epithelial hyperplasia: proposal of a concept

Background:  Evidence is accumulating for the aetiological role of human papillomavirus (HPV) in the pathogenesis of potentially malignant oral mucosal lesions and squamous cell carcinomas.
Methods:  Paraffin tissue sections from 49 patients with 'white patches' of the oral mucosa were investigated histologically, by broad-spectrum PCR followed by genotyping and chromogenic in situ hybridisation (CISH).
Results:  Histologically, 33 flat hyperplasias and 16 papillary hyperplasias were diagnosed. Twenty-two of 28 samples studied (78.6%) were positive for HPV DNA by PCR and six were negative. The following HPV types were detected in decreasing order of prevalence: HPV 35, HPV 6, HPV16, HPV 53, HPV 18, HPV 51 and HPV 55. Seventeen samples (60.7%) contained high-risk HPV DNA. Using CISH, ≥ 1 HPV signals were detected at least in a few epithelial cells in 95% of cases studied. All but one case were positive with the high-risk HPV probe and all HPV infections contained low viral load. Concordant positive results both by PCR and CISH were detected in 14 of 19 cases (73.7%) analysed.
Conclusions:  The high prevalence of HPV infection in hyperplastic 'white patches' of the oral mucosa supports the putative role of HPV at an early stage of oral carcinogenesis. These results further indicate that the majority of white oral mucosal lesions – flat, exophytic, wart-like or papillary proliferations – could be considered as the clinical manifestations of oral HPV infection. This finding has clinical relevance regarding therapy and patient management and may help in elucidating the role of HPV infection in oral carcinogenesis.     

口腔粘膜病损HPV的检测

应用ＰＣＲ技术检测口腔粘膜病损中ＨＰＶ的感染情况。病损包括白斑、扁平苔藓、慢性盘状红斑狼疮、日光性唇炎、鳞癌及鳞癌术后上皮高度增生共３４例。结果表明在口腔癌与白斑中斑可存在ＨＰＶ的感染，ＨＰＶ可能是口腔癌的致病因子之一。     

Human papillomavirus as a risk factor in oral carcinogenesis: a study using in situ hybridization with signal amplification

Introduction:  It is still controversial whether human papillomavirus (HPV) can be considered a risk factor in oral carcinogenesis. The aim of this study was to detect HPV DNA in 50 cases diagnosed as oral leukoplakias, with different degrees of epithelial dysplasia, and as oral squamous cell carcinomas, using in situ hybridization with signal amplification (CSA-ISH).
Methods:  HPV DNA was assessed in paraffin sections using CSA-ISH with a wide-spectrum biotinylated DNA probe. In HPV-positive cases, genotyping with specific probes to HPV types 6/11, 16/18 and 31/33 was performed.
Results:  The overall prevalence of HPV infection was 24%, markedly higher than that found in the control group. Results showed a discrete proportional relationship in the indices found in leukoplakia with no dysplasia, leukoplakia with dysplasia, and squamous cell carcinoma, but this was not statistically significant. When separating the group of leukoplakia by degrees of dysplasia, this relation of proportion was not observed. In genotyping, HPV types 16/18 were the most prevalent, and types 6/11 were only found in groups of mild or no dysplasia.
Conclusion:  The results suggest that HPV is not likely to play a role in the progression of malignant transformation in oral lesions. Nevertheless, the increased prevalence of HPV infection compared to normal oral mucosa and the fact that high-risk HPV types were the most frequently identified do not allow the exclusion of HPV as a risk factor in oral carcinogenesis.     

口腔粘膜组织中HPV16DNA和空泡细胞关系的研究

本研究采用斑点杂交技术，检测８３例口腔扁平苔藓、口腔白斑、口腔鳞癌及正常口腔粘膜组织中的ＨＰＶ１６ ＤＮＡ，并分析ＨＰＶ１６ ＤＮＡ与空泡细胞出现的关系。口腔粘膜组织ＨＰＶ１６ ＤＮＡ的检出率为１０．８％，空泡细胞的检出率为３４．９％。两者均阳性的检出率为７．２％，结果提示空泡细胞不是ＨＰＶ感染所特有的，不能作为诊断ＨＰＶ的特征性指标。     

Valuation of exfoliative cytology as prediction factor in oral mucosa lesions

The aim of this study was immunolabeling oncoproteins Ck14, p53, p21 and Bcl-2 in order to evaluate their expression in premalignant and malignant stomatological lesions in oral epithelial, and to compare this expression with exfoliative cytology alterations in the same patients. It was studied biopsies and cytologies of 13 subjects with oral lichen planus, with or without Human Papilloma Virus (HPV), leukoplakia and squamous cell carcinoma clinically diagnosed and confirmed by anatomopathological studies. The oral lichen planus lesion presented binuclei orange cells; and in leukoplakia lesions only orange stained was observed; meanwhile koilocytes, inflammatory cells, enlarge nuclear volume and pathogenic microorganisms were observed in the HPV infections and squamous cells carcinoma (SCC). The Ck14, p53, p21 and Bcl-2 proteins were found modified in the leukoplakia, oral lichen planus and cancer. Cytological alterations and positive immunolabeling or over-expression of Ck14 cytokeratine in the upper epithelial stratus should be indicator of malignant transformations as doing subsequence exams.     

Papillary lesions of the oral cavity: relationship to human papillomaviruses

Human papillomaviruses are a group of genetically related organisms that infect stratified squamous epithelium. Unlike many other viruses that infect oral epithelium and induce lysis of the cells they penetrate, HPVs induce proliferative changes in these cells that result in both benign and malignant tumors. The common skin wart (verruca vulgaris) is induced by HPV 2 and 4. Genital warts (condylomas) and the common solitary oral papilloma are associated with HPV 6 and 11. Either HPV 13 or 32 causes focal epithelial hyperplasia. All of these wart-like lesions are benign growths of the stratified squamous lining of the oral cavity and lips and can be treated by surgical excision or laser ablation. HPV 16 and other less frequently encountered genotypes are associated with uterine cervix cancer in 95 percent to 98 percent of cases, and the evidence for a causal role is robust. There are emerging data that implicate HPV in certain subsets of oral cancer, particularly those that arise in the oropharynx/tonsillar region. Some instances of the various histologic subtypes subsumed under proliferative verrucous leukoplakia and verrucous carcinoma also harbor HPV.     

OC11Hyperimmunoglobulin E syndrome presenting with oral premalignant papillomas

Hyperimmunoglobulin E syndrome (HIES) is a rare primary immunodeficiency characterized by the clinical triad of recurrent staphylococcal abscesses, recurrent cyst-forming pneumonia and an elevated serum IgE level. A 10-year-old female with HIES presenting with widespread oral premalignant papillomas, which are an uncommon co-existing manifestation, and labial herpes simplex virus infection is reported. The patient experienced recurrent furuncles, pneumonia, tuberculosis, otitis media and sinusitis attacks since she was 6 months-old. She had been diagnosed with HIES at the age of 6 when the papillomatous lesions first occurred in oral cavity. Family history was not remarkable. Intraoral examination revealed diffuse multiple papillomas covering the tongue, lips, buccal, labial and palatinal mucosa and gingiva extending to the crowns of teeth. The lips were enlarged to the double of normal size due to numerous papilloma formations and the outline could hardly be traced. These lesions caused nutritional, aesthetic, psycological problems besides loss of appetite. Histopathological examination of the lesions revealed papillary proliferations of squamous epithelium which was confirmed to be positive for human papilloma virus DNA by molecular based test (Inno lipa HPV genotyping test) which is based on reverse hybridization principle and PCR. Typing of HPV was found to be as HPV 6, 11, 16, 18, 31, 33, 35, 39, 40, 42–45, 51–53, 56, 58, 59, 66, 68, 70 and 74, forming a high/intermediate risk group for malignancy. For the management of widespread papillomas, diode laser was applied. After therapy, significant clinical improvement, reduction in oral discomfort and improvement in appetite was observed. Since infection with HPV is a significant independent factor for oral squamous cell carcinoma, the patient remains under regular review.     

Human papilloma virus in erosive oral lichen planus

Several types of human papilloma viruses (HPV) have been associated with benign and malignant squamous cell tumours of mucosal epithelium. To identify HPV in erosive oral lichen planus (OLPe), considered as a premalignant lesion, tissues from 20 patients were examined by Southern blot hybridization with 32P-labeled HPV DNA probes. Type 11 was found in 6 of the lesions while HPV types 6, 16 and 18 were not detected in any of the tissues examined. Using a type-specific polymerase chain reaction (PCR) assay for HPV-6, 11, 16 and 18, HPV-11 was detected in 8 of the samples (all of those positive by Southern blot), and, in addition, HPV-6 was found in 5 samples and HPV-16 in 3 samples. Overall, by the more sensitive PCR assay, 65% of samples were positive for HPV DNA. The finding of HPV DNA in many of the samples using two different techniques indicates a high prevalence of HPV in the OLPe afflicted oral mucosa. However, the role of HPV in the pathogenesis of OLPe has yet to be determined.     

Detection of human papillomavirus-genomic DNA in oral epithelial dysplasias, oral smokeless tobacco-associated leukoplakias, and epithelial malignancies

Human papillomavirus (HPV) is an infectious agent that is increasingly associated with mucosal cancers, in particular cancer of the cervix. The present investigation was undertaken in an attempt to determine whether HPV could be easily detected in biopsies of oral tissues, specifically oral squamous cell carcinomas, oral epithelial dysplasias, smokeless tobacco keratoses, verrucous hyperplasia, and verrucous carcinoma. In situ DNA hybridization methods were used to isolate specific HPV genomes. Among 100 instances of benign leukoplakia, only 4% of non-tobacco-related and 10% of smokeless tobacco-related lesions harbored viral sequences. We were able to detect viral sequences in dysplastic lesions 3% of the time. Alternatively, 17% and 20% of the verrucous hyperplasias and verrucous carcinomas were positive for viral nucleic acids. Six percent of the squamous cell carcinomas harbored HPV. On the basis of these findings, it is concluded that HPV of known genotype can be identified in oral premalignant and malignant neoplasms.     

口腔鳞状细胞乳头状瘤组织中HPV DNA的原位杂交研究

目的 探讨人乳头瘤病毒（ｈｕｍａｎ ｐａｐｉｌｌｏｍａｖｉｒｕｓ，ＨＰＶ）感染与口腔鳞状细胞乳头状瘤（ｓｑｕａｍｏｕｓ ｃｅｌｌ ｐａｐｉｌｌｏｍａ，ＳＣＰ）的发生之间的关系。方法 应用地高辛标记的ＨＰＶ６／１１和ＨＰＶ１６／１８核酸探针分别在３０例口腔ＳＣＰ组织上进行原位杂交，检测口腔ＳＣＰ组织中ＨＰＶ ＤＮＡ的特征。结果 ＨＰＶ６／１１ ＤＮＡ阳性１６例（５３％），ＨＰＶ１６／１８ＤＮＡ未检出，ＨＰＶ６／１１ＤＮＡ阳性细胞多数分布在鳞状上皮的表层、中层和基底层。结论 原位杂交方法可以检测口腔ＳＣＰ组织中ＨＰＶ ＤＮＡ的存在并能准确组织定位，进一步支持ＨＰＶ６／１感染与口腔ＳＣＰ的发生密切相关。     

Morphological and immunohistochemical evidence suggesting human papillomavirus (HPV) involvement in oral squamous cell carcinogenesis

Human papillomavirus (HPV) is an agent responsible for squamous cell tumors (verrucae, condylomata and papillomas) at various sites of the body, the oral cavity included. Due to the recently pointed-out association between these HPV lesions and the squamous cell dysplasias and malignancies in the uterine cervix, in the bronchus and in the larynx, the present work was carried out to assess, whether morphological signs (cytopathic effects of HPV) or HPV antigens could be found in lesions of oral squamous cell carcinomas. Biopsies from 40 cases of oral squamous cell carcinomas were surveyed by light microscopy with special emphasis on the presence of the histopathological features suggesting an HPV lesion, i.e. whether flat, inverted or papillomatous condylomas are present concomitantly with the malignancy. All specimens were also subjected to immunoperoxidase staining with anti-HPV serum to disclose the possible HPV antigens in the lesions. Morphological signs of the flat-type HPV lesion were found in 4 cases (10%), those of an inverted type in 3 cases (7.5%), and those of a papillomatous type in 9 cases (22.5%). Epithelial cells (mostly koilocytes) showing HPV-positive nuclei were disclosed in 5 papillomatous lesions, in 2 inverted lesions and in 1 flat lesion. The results suggest that HPV might be the agent involved in the development of at least certain special types of oral squamous cell carcinomas; albeit further confirmatory evidence with other techniques (DNA hybridization) is still needed.     

Human papillomavirus infection in oral potentially malignant disorders and cancer

Human papillomavirus (HPV) infects keratinocytes in the mucosa or skin, and persistent infection with HPV may lead to premalignant lesions and invasive cancer, especially cervical cancer. It has also been hypothesized that HPV infection is an etiological factor of oral squamous cell carcinoma and oral precancerous disorders such as lichen planus, leukoplakia, and erythroplakia. A high percentage of HPV in oral lesions supports the possible viral contribution, but an association of HPV infection with these lesions remains to be established. The current paper will update the latest progress of HPV infection in several oral potentially malignant disorders and oral squamous cell carcinoma and discuss the impact of HPV infection on the progression of oral potentially malignant disorders.     

人乳头瘤病毒和人巨细胞病毒与口腔癌的关系

目的：检测口腔白斑和口腔鳞状细胞癌中的人乳头瘤病毒（HPV16）DNA和人巨细胞病毒（HCMV）DNA,探讨HPV16和HCMV与口腔癌之间的关系。方法：应用多聚酶链反应（PCR）技术扩增HPV16DNA和HCMVDNA,采用2%琼脂糖凝胶电泳分析,检测口腔白斑和口腔鳞状细胞癌中的HPV16DNA和HCMVDNA。结果：6例口腔白斑中,HPV16DNA的阳性率为16.7%,HCMVDNA为16.7%,16例口腔鳞癌组织HPV16DNA的阳性率为31.3%,HCMVDNA为25%,例正常组织均未检测出。结论：4HCMV与HPV16在口腔癌的发生中具有协同致癌作用。     

Human papillomavirus (HPV) DNA sequences in oral precancerous lesions and squamous cell carcinoma demonstrated by in situ hybridization

A series of routinely processed, paraffin-embedded biopsies from 73 surgically treated oral precancerous lesions (OPL) (22 cases), and oral squamous cell carcinomas (SCC) (51 cases), was first screened using an in situ DNA hybridization technique with a human papillomavirus (HPV) DNA probe cocktail containing the 35S-labelled DNA of HPV types 6, 11, 13, 16, 18 and 30. The specific HPV types in lesions shown to contain HPV DNA in this procedure were further analysed by using in situ hybridization and the 6 HPV DNA probes separately. A total of 12/73 (16.4%) of the lesions proved to contain HPV DNA; 6/51 (11.8%) carcinomas and 6/21 (28.6%) dysplasias. The most frequent sites of HPV DNA-positive lesions were palate (4/7; 57%), followed by the floor of the mouth (2/8; 25%), the tongue and gingiva (11.8%). HPV 13 or HPV 30 were not found in any of the lesions studied. HPV 11 DNA was demonstrated in 2 mild dysplasia lesions, but not in carcinomas. One additional mild dysplasia proved to contain HPV 6 DNA. HPV 16 DNA was present in 5 biopsies; 3 carcinomas and 2 dysplasias. In one of the HPV 16-positive carcinomas, HPV 18 DNA was simultaneously present. HPV 18 alone was found in 3 additional carcinomas and in one moderate dysplasia lesion. The results confirm the recently reported evidence on HPV involvement in OPL and oral cancer. The implications of these findings are discussed in terms of the possible HPV etiology of oral SCC.(ABSTRACT TRUNCATED AT 250 WORDS)     

Condyloma acuminatum and human papilloma virus infection in the oral mucosa of children

PURPOSE: The purpose of this study was to investigate the clinicopathological features of oral condylomas in children and condylomatous lesions of their mothers. Moreover, the authors sought to determine the mode of transmission of this disease and to find the genotype of human papilloma virus (HPV) in the children's oral condyloma. METHODS: Nine instances of oral condyloma acuminatum in children and lesions in their mothers were reviewed. Their HPV genotypes were evaluated by in situ hybridization (ISH). RESULTS: This study revealed that the lesions appeared during 3 years of age and the most common location was the hard and soft palate. Seven of the 9 mothers had experienced vulva and/or oral cavity condylomata during pregnancy. Social evaluation confirmed sexual abuse in 1 girl, and probable sexual abuse in another girl. The results of ISH demonstrated HPV 16/18 DNA being positive in 5 of the 9 cases, and HPV 6 and HPV 11, HPV 6 and HPV 16/18, HPV 6, and HPV 11 DNA being positive, respectively, in 1 case. HPV DNA types in mother-child pairs were not concordant. CONCLUSIONS: Oral condyloma acuminatum in children is probably induced by HPV 16/18. The mode of transmission by sexual abuse is the most likely route. Prenatal transmission of HPV to children is rare. This study provides further confirmation of possible different genotype and transmission in oral CA of children and adults.     

Oral mucosal changes in women with genital HPV infection

Sixty different types of human papillomavirus (HPV) are currently recognized. Of these, HPV types 1, 2, 4, 6, 7, 11, 13, 16, 18, 30, 32 and 57 have been identified in oral squamous cell lesions. The prevalence and incidence of clinical HPV infections of oral mucosa are incompletely established, and the figures on subclinical and latent infections are completely lacking. Similarly, no data exist on transmission of oral HPV infections. A long-term prospective follow-up study was started to assess the oral mucosal changes related to HPV infection in women with genital HPV infections. The aim was to elucidate whether genital HPV infections predispose the oral mucosa to this virus. This study reports the clinical, histologic and cytologic findings of oral mucosa as related to the genital status of 334 women prospectively followed-up in Kuopio University Central Hospital since 1981. At the time of examination, 5% presented with hand warts. Clinical wart in oral cavity was found in three patients (0.9%) only. Oral mucosa was clinically normal in 207 of 334 (62%) patients examined. Biopsies (n = 255) were taken from the buccal mucosa beneath the linea alba and above the sulcus in the region of the first molar, or from a lesion whenever present. Cytologic scrapings were taken from both sides of the buccal mucosa. Koilocytes were found in 0.9% of the cytologic scrapings, and in 9.4% of the biopsies. Altogether, four squamous cell papillomas and five flat condylomas were found in the biopsies. Morphologic changes suggestive for HPV were present in 25 biopsies. Hyperkeratosis proved to be a frequent finding (34%) in these biopsies.(ABSTRACT TRUNCATED AT 250 WORDS)     

Human papillomavirus (HPV) DNA sequences in oral precancerous lesions and squamous cell carcinoma demonstrated by in situ hybridization

A series of routinely processed, paraffin-embedded biopsies from 73 surgically treated oral precancerous lesions (OPL) (22 cases), and oral squamous cell carcinomas (SCC) (51 cases), was first screened using an in situ DNA hybridization technique with a human papillomavirus (HPV) DNA probe cocktail containing the 35S-labelled DNA of HPV types 6, 11, 13, 16, 18 and 30. The specific HPV types in lesions shown to contain HPV DNA in this procedure were further analysed by using in situ hybridization and the 6 HPV DNA probes separately. A total of 12/73 (16.4%) of the lesions proved to contain HPV DNA; 6/51 (11.8%) carcinomas and 6/21 (28.6%) dysplasias. The most frequent sites of HPV DNA-positive lesions were palate (4/7; 57%), followed by the floor of the mouth (2/8; 25%), the tongue and gingiva (11.8%). HPV 13 or HPV 30 were not found in any of the lesions studied. HPV 11 DNA was demonstrated in 2 mild dysplasia lesions, but not in carcinomas. One additional mild dysplasia proved to contain HPV 6 DNA. HPV 16 DNA was present in 5 biopsies; 3 carcinomas and 2 dysplasias. In one of the HPV 16-positive carcinomas, HPV 18 DNA was simultaneously present. HPV 18 alone was found in 3 additional carcinomas and in one moderate dysplasia lesion. The results confirm the recently reported evidence on HPV involvement in OPL and oral cancer. The implications of these findings are discussed in terms of the possible HPV etiology of oral SCC. The use of the in situ DNA hybridization as a powerful tool (enabling the localization of specific HPV DNA sequences and the proper classification of the lesion at the same site) in the study of routinely processed oral biopsies is strongly advocated.     

Association between proliferative verrucous leukoplakia and infection with human papillomavirus type 16

Proliferative verrucous leukoplakia (PVL) is a recently described clinical entity characterized by multifocal oral lesions that frequently progress to oral cancer despite abstinence from tobacco use by most patients. To determine if this condition is associated with human papillomavirus (HPV), a polymerase chain reaction (PCR) for HPV DNA was performed on 9 lesions from 7 patients with PVL, histologically diagnosed with focal keratosis (1), papilloma (1), epithelial dysplasia (5) and squamous cell cancer (2). Eight (89%) were HPV positive, 7 for HPV 16. For comparison, we studied 55 non-PVL-associated oral specimens, including 24 oral squamous cell cancers. Of the cancers, 8 (33%) were HPV positive, 4 for HPV 16. These data suggest that HPV 16 infection may play an important role in the pathogenesis of PVL-associated oral dysplasia and possibly cancer, but is found in only a small proportion of the more common, non-PVL associated-oral lesions.