Viral DNA detection and RAS mutations in actinic keratosis and nonmelanoma skin cancers

Background Actinic keratosis (AK) is a well‐established precancerous skin lesion that has the potential to progress to squamous cell carcinoma (SCC). Basal cell carcinoma (BCC) is a locally aggressive slowly growing tumour that rarely metastasizes. A number of viruses have been proposed to play a role in the development of nonmelanoma skin cancers (NMSC), but the most plausible evidence to date suggests that cutaneous human papillomavirus (HPV) is the key instigating factor. Objectives To evaluate the prevalence of HPV, cytomegalovirus (CMV), herpes simplex virus (HSV) and Epstein–Barr virus (EBV) and investigate their relationship with the presence of RAS gene mutations in cutaneous lesions obtained from nonimmunosuppressed patients. Methods HPV, CMV, HSV and EBV detection was performed using polymerase chain reaction (PCR) in skin biopsies (26 AK, 12 SCC and 15 BCC samples) that were collected from immunocompetent patients. The RAS mutation incidence was also investigated in all cutaneous lesions by use of PCR/restriction fragment length polymorphism and direct DNA sequencing. Results Seventeen out of 53 (32%) skin lesions were found to be positive for HPV DNA. The highest incidences of HPV infection were five of 15 (33%) in BCC and four of 12 (33%) in SCC specimens. The HPV incidence was eight of 26 (31%) in AK and eight of 53 (15%) in normal skin tissue. Twelve out of 53 (23%) skin lesions were CMV‐positive. The highest incidence of CMV infection was six of 15 (40%), observed in BCC specimens. The CMV incidence was two of 26 (8%) in AK and four of 12 (33%) in SCC. No normal skin biopsy was found to be positive for CMV. All cutaneous samples were negative for HSV and EBV DNA, as assessed by our PCR‐based assays. Only three samples, one AK (4%), one BCC (6%) and one SCC (8%), were found to carry a G>T transversion at the second position of HRAS codon 12. Both HRAS mutant SCC and BCC biopsies were HPV‐ and CMV‐positive, as well. Conclusions HPV DNA is detected in NMSC, AK and normal skin biopsies. Our results also indicate that CMV is involved in NMSC at higher levels than in premalignant lesions, whereas the virus was not detected in normal skin biopsies. HSV and EBV do not appear to be involved in the pathogenesis of cutaneous lesions. Moreover, we suggest that the HRAS codon 12 mutation is not a very common event in AK or NMSC. Finally, both viral infection and HRAS activation appear to represent independent factors in the aetiology of NMSC, samples of which were obtained from immunocompetent patients.     

SOUTHERN BLOT ANALYSIS OF SKIN BIOPSIES FOR HUMAN PAPILLOMAVIRUS DNA: RENAL ALLOGRAFT RECIPIENTS IN SOUTH-EASTERN QUEENSLAND

The 104 skin biopsies from 34 patients who attended a Renal Transplant Unit in Brisbane over 12 months included 40 squamous cell carcinoma (SCC), 22 solar keratoses, 4 hyperkeratoses, 18 warts and 11 basal cell carcinoma (BCC). Human papillomavirus (HPV) DNA was identified by Southern blot hybridisation using, as individual probes, purified insert DNA from recombinant HPV 1, 2, 3 or 3/10, 4, 5 or 5/8, 7, 11, 16, 18 and 41 under relaxed conditions and characterised by restriction enzyme analysis and Southern blot hybridisation under more stringent conditions. Genomic HPV DNA was characterised in 7 skin biopsies from 4 renal allograft recipients (RARs): HPV 1A in a SCC (20 copies/cell) and a BCC (10 copies/cell) from the one patient, HPV 36 (20 copies/cell) in a SCC, HPV 1A (1000 copies/cell) in a wart and HPV 2B (200-800 copies/cell) in 3 warts from the one patient. Only HPV 1A in the SCC exhibited a significant degree of subtype variation. HPV DNA was identified in another 5 skin biopsies from another 4 RARs: HPV 3A in a wart and a hyperkeratosis, HPV3/10-related DNA in 2 solar keratoses and HPV5/8-related DNA in another (20-50 copies/cell). The incidence of HPV 5 (or 5-related HPVs) in RAR SCC was very low and that of HPV DNA in RAR warts was lower than that recorded elsewhere but this was not due to insensitivity of the assays. There was no evidence for a role for HPV in the aetiology of skin cancer in RARs in south-eastern Queensland but the possibility remains that as yet unidentified HPV types are involved.     

Serological association of beta and gamma human papillomaviruses with squamous cell carcinoma of the skin

Background  Cutaneous human papillomaviruses (HPVs) may play a role in the development of squamous cell carcinomas (SCC) of the skin.
Objectives  Available serological studies on HPV and skin SCC have analysed only few HPV types from the phylogenetic genus beta. The potential association of cutaneous HPV types from the genera alpha, gamma, mu and nu with skin SCC has not been thoroughly analysed so far.
Methods  Using multiplex serology, a method that allows analysing sera for antibodies to up to 100 different antigens simultaneously, we re-analysed an SCC case–control study in immunocompetent individuals (43 cases, 77 controls) for antibodies to L1 capsid proteins of 29 cutaneous and two mucosal HPV types from five different genera.
Results  Significantly increased SCC risks were observed for the beta HPV types 15, 17 and 38, as well as for the gamma HPV type 50, with type-specific odds ratios (ORs) ranging from 2·6 to 3·4. Significant associations were also found in cases of seropositivity for any type of the beta 2 species (OR 3·3, 95% confidence interval [CI] 1·2–8·7) and for any type of the gamma genus (OR 3·1, 95% CI 1·1–8·6). With regression models that included all HPV types and forward stepwise selection, two gamma HPV types (HPV 95, OR 25, 95% CI 1·2–509; HPV 50, OR 3·6, 95% CI 1·4–9·4) were each significantly associated with skin SCC.
Conclusions  Our study confirms a possible role of cutaneous HPV in the development of skin SCC. Future studies should include skin HPV types from more than only the beta genus, especially gamma types.     

E6 and E7 proteins from different beta-papillomaviruses types do not interfere in UVB-induced apoptosis of HaCaT keratinocytes

Beta-papillomaviruses (beta-HPV) have been linked to the development of skin cancer in humans. Because both E6 and E7 proteins from beta-HPV have been involved in the potential carcinogenicity of these viruses, we investigated their role on UVB-induced apoptosis in HaCaT cell line. HaCaT cells have been transduced with both E6/E7 using a retroviral system and treated with PRIMA-1. Apoptosis was assessed by flow cytometry to measure mitochondrial membrane potential and DNA fragmentation. HaCat keratinocytes transduced with both E6 and E7 genes of seven beta-HPV types (HPV5, HPV8, HPV14, HPV24, HPV36, HPV38 and HPV49) did not demonstrate any inhibition of UVB-induced apoptosis, even after p53 reactivation through PRIMA-1. Our data suggest that the expression of E6 and E7 exert different modulatory effects on UVB-induced apoptosis according to beta-HPV types and to the cellular genetic context.     

人乳头瘤病毒6b型早期蛋白E6和E7基因的克隆及表达

目的:从尖锐湿疣(CA)标本中克隆出人乳头瘤病毒6b型(HPV-6b)早期蛋白E6、E7基因,并进行序列分析比对及原核表达,为HPV感染的检测和基因工程疫苗研究奠定基础。方法:用PCR法,从CA标本中扩增HPV-6的E6、E7基因,构建pUCHPV-6E6、pUCHPV-6E7两个重组体,酶切鉴定及测序分析比对。经双酶切与表达载体pGEX-5X-1定向连接,构建pGEX-5X-l重组表达质粒,转入BL21大肠杆菌,IPTG诱导GST融合蛋白表达及Westernblot鉴定。结果:克隆出HPV-6b早期蛋白E6、E7基因,成功构建pUCmHPV-6bE6、pUCmHPV-6bE7重组质粒,克隆获得HPV-6bE6、E7基因与GenBank标准株序列完全相同,经双酶切定向克隆,成功构建重组表达质粒pGEX-5X-lHPV-6bE6和pGEX-5X-lHPV-6bE7,转入BL21大肠杆菌,高效表达GST融合蛋白。结论:本研究克隆出的HPV-6bE6、E7基因与标准株相同,HPV-6bE6、E7GST融合蛋白获得高效表达,为HPV-6bE6、E7基因表达产物的纯化、体外活性以及E6、E7为靶位的基因疫苗研究奠定基础。     

Identification of human papillomavirus in keratoacanthomas

BACKGROUND: Keratoacanthomas are benign, clinically distinct skin tumors that may infiltrate and show cellular atypia. A viral etiology has been suggested, and the aim was to search for human papillomavirus (HPV) in keratoacanthomas. METHODS: From 21 immunosuppressed organ transplant recipients and 11 non-immunosuppressed patients, 72 fresh biopsies with diagnosis of keratoacanthomas were analyzed. For detection of cutaneous and genital HPV DNA, single-tube nested "hanging droplet" polymerase chain reaction (PCR) and another PCR (GP5+ and 6+) were used, respectively. RESULTS: Among 21 immunosuppressed patients, 71% (15/21) harbored HPV DNA at least in one sample. Of the keratoacanthoma lesions, 55% (33/60) were HPV DNA positive. Fourteen samples from eight immunosuppressed patients contained HPV types 5, 9, 10, 14, 19, 20, 21, 38, 49, 80, putative HPV types as HPVvs20-4, HPVvs75, and HPVvs92 and FA16.1, FA23.2, FA37, FA75, and FA81. Among 11 non-immunosuppressed patients, 36% (4/11) harbored HPV DNA at least in one sample, and 33% (4/12) of their keratoacanthomas were HPV DNA positive. In total, HPV DNA was detected in 51% (37/72) of the keratoacanthomas. CONCLUSIONS: By the use of PCR, cutaneous HPV DNA was detected in 51% (37/72) of the keratoacanthomas. No predominating HPV type or genital HPV type was identified. The role of HPV in keratoacanthomas remains thus elusive.     

北京地区人乳头瘤病毒16E6E7基因变异和序列分析

目的 了解北京地区人乳头瘤病毒(HPV)16型E6E7结构特点，为研制HPV16疫苗奠定基础。方法 从HPV感染组织中提取DNA，PCR鉴别其型别，从单独HPV16感染的标本中分离HPV16 E6E7基因，克隆入载体pGEM-3ZF，进行双向测序，与德同标准株进行比较。结果 构建了北京地区HPV16E6E7的重组质粒。测序结果表明该基因全长为776bp，与已发表的德同标准株长度相等，3处核苷酸发生变异，同源性99．7％。分别位于第60、96和565nt(相当于HPV16全长序列中第142、178和647nt)，2处位于F6区，1处位于E7，其对应氨基酸分别为第20、32和188个氨基酸，分别由Pro(CCA)、Asp(GAT)和Asn(AAT）变异为Pro(CCG)、Glu(GAG)和Ser(AGT)。结论 北京地区HPV16E6E7基因结构与德国标准株存在差异。     

A broad spectrum of human papillomavirus types is present in the skin of Australian patients with non-melanoma skin cancers and solar keratosis

BACKGROUND: Human papillomavirus (HPV) may play a role in the pathogenesis of non-melanoma skin cancer (NMSC) in epidermodysplasia verruciformis (EV) patients, but in the general population no specific HPV types have been associated with these lesions. Objectives To examine the spectrum of HPV types present in the skin and tumours of Australian patients with NMSC or solar keratosis (SK). METHODS: Biopsies from tumours, and cotton swab samples of perilesional skin and buttock skin from each of 59 Australian patients with basal cell carcinoma (BCC), squamous cell carcinoma (SCC) or SK were tested for HPV DNA by polymerase chain reaction (PCR) using HPV consensus (FAP) primers and by type-specific primers for HPV 38 and candidate HPV 92. The identification of HPV type from consensus PCR was performed by sequencing and comparison with GenBank. RESULTS: In total, 49 of 59 (83%) patients harboured HPV DNA, which was detected in 28 of 64 (44%) biopsies, 48 of 64 (75%; P < 0.001) perilesional swabs and 36 of 59 (61%; P = 0.04) buttock swabs. Forty-five different HPV types/putative types were detected: 15 were previously characterized HPV types, 17 were earlier described putative types and 13 were new putative types. In addition, six subtypes and four variants of HPV sequences were identified. HPV types within the B1 group (EV HPV types) were found in 26 of 64 (40%) lesions, 44 of 64 (69%) perilesional swabs and 35 of 59 (59%) buttock swabs. HPV 38 was detected in 23 of 59 (39%) patients, and was found in seven of 16 (43%) SKs, but was less common in SCCs [three of 23 (13%); P = 0.037] and BCCs [four of 25 (16%); P = 0.056]. Candidate HPV 92 was found in seven of 59 (12%) patients. CONCLUSIONS: A broad spectrum of HPV types, the majority from the B1 group, was found in skin of Australian patients with skin tumours. HPV 38 was found significantly more often in SK than in SCC. However, the role of cutaneous HPV infection in the pathogenesis of NMSC remains elusive.     

Frequency and spectrum of HPV types detected in cutaneous squamous-cell carcinomas depend on the HPV detection system: a comparison of four PCR assays

BACKGROUND: The association of human papillomavirus (HPV) with cutaneous squamous-cell carcinomas (SCCs) has been described recently, but the frequency and spectrum of HPV types identified differed substantially in distinct studies. OBJECTIVE: Comparison of different PCR assays with respect to sensitivity and range of HPV types detected. METHOD: Cutaneous SCC were analyzed for HPV DNA using both consensus PCR assays with degenerate primers and PCR assays with nondegenerate primers derived from HPV types 5 and 8. RESULTS: HPV DNA was found in 50% of SCC specimens using degenerate primers. The rate of HPV-DNA-positive specimens increased to 69% when PCR assays with nondegenerate primers were applied in addition. The spectrum of HPV types detected with each of the PCR assays differed considerably. CONCLUSIONS: The frequency and spectrum of HPV types detected in cutaneous SCC strongly depends on the HPV detection system used and urges the need for standardization of HPV detection and typing in skin lesions in order to characterize HPV types predominating in distinct tumors.     

Epidermodysplasia verruciformis and cutaneous human papillomavirus DNA, but not genital human papillomavirus DNAs, are frequently detected in vulvar and vaginal melanoma

Vulvovaginal melanomas are rare and their etiology is unknown. Genital mucosal human papillomavirus (HPV) 16 has been identified in both cutaneous and mucosal melanoma, suggesting that it might play a role in the pathogenesis or progression of melanoma. In this study, we investigated the prevalence of HPV DNA by using a broad spectrum of degenerate and type-specific primers for genital-mucosal, epidermodysplasia verruciformis-associated (EV), and cutaneous HPV types in 6 vulvar and 3 vaginal melanomas. The patients were mostly postmenopausal women (8/9), had a mean age of 67 years (range, 44-85 years), and had mucosal lentiginous (7) or nodular (2) melanomas. In the adjacent skin/mucosa, mucosal melanosis was found in 5, lichen sclerosus or a lichenoid mucositis in 4, and blue nevi in 2 women. With nested polymerase chain reaction techniques followed by direct sequencing, HPV DNA was identified in 6 of 9 (67%) melanomas; these were either cutaneous (HPV 3) (4/9) or epidermodysplasia verruciformis-associated types (HPV 38, Z95969, AJ00151) (4/9). Epidermodysplasia verruciformis-associated HPV (type 15) was found solely in 1/10 (10%) normal vulvar controls. Genital-mucosal HPV types were not detected either by degenerate nested polymerase chain reaction or type-specific probes for HPV 16. We propose that the above findings are not coincidental but may represent a molecular record of HPV involvement in pathogenesis or progression of melanoma, which is consistent with the strong but poorly defined association of cutaneous HPV types with nonmelanoma skin cancers. The theory that HPV may act as a cofactor in melanoma development deserves further clinical and experimental investigations.     

人乳头瘤病毒11型早期蛋白E6和E7基因的克隆及序列分析

目的：从尖锐湿疣(CA)标本中克隆出人乳头瘤病毒11型(HPV11)早期蛋白E6、E7基因，并进行序列测定及编码氨基酸序列分析，为HPV感染的检测和基因工程疫苗研究奠定基础。方法：用PCR法，从CA标本中扩增出HPV11E6、E7基因，与载体pGEX-6P-1连接成重组质粒pGEX-6P-1／E6、pGEX-6P-1／E7，酶切鉴定及双脱氧法测序观察其变异状况。结果：克隆出HPV 11早期蛋白E6、E7基因，成功构建重组质粒pGEX-6P-1／E6、pGEX-6P-1／E7。本研究克隆出的HPV11 E7基因与GenBank标准株序列完全相同，E6基因有两个位点的变化。结论：本研究克隆出的HPV11 E6、E7基因与标准株基本相同，这将为进一步研究E6、E7基因的表达、免疫活性及流行病学奠定基础。     

Activation of Src-family tyrosine kinases in hyperproliferative epidermal disorders

Background:  Src-family tyrosine kinases (SFKs) are important regulators of keratinocyte growth and differentiation. In a broad range of cell types, persistent activation of SFKs correlates with increased cell proliferation. In this study, we determined if SFK activity is increased in cutaneous neoplasia and psoriasis, common hyperproliferative epidermal disorders.
Methods:  Formalin-fixed tissue sections of unremarkable epidermis, psoriasis, actinic keratoses (AKs), squamous cell carcinoma in situ (SCIS) and squamous cell carcinoma (SCC) were subjected to immunohistochemical staining for activated SFKs.
Results:  All psoriasis specimens displayed significantly greater staining for activated SFKs than sections of unremarkable skin. In the psoriasis biopsies, the degree of epidermal hyperplasia was proportional to the level of activated SFK staining. All AKs, SCISs and SCCs exhibited more prominent staining than sections of unremarkable epidermis. No discernable difference in activated SFK staining was seen between AKs, SCIS and SCC specimens.
Conclusions:  This study shows increased staining of activated SFKs in human biopsy specimens of psoriasis and cutaneous neoplasia. These data provide direct evidence for increased activation of SFKs in the pathogenesis of hyperproliferative epidermal disorders.     

The role of human papillomavirus in the development of pyogenic granulomas

Background Pyogenic granulomas (lobular capillary hemangiomas) and condyloma acuminata share similar locations and risk factors. Human papillomavirus (HPV) types 6 and 11 are commonly associated with condyloma acuminata, but their association with pyogenic granulomas has not been evaluated. The purpose of this study was to determine whether pyogenic granulomas contain evidence of infection with condyloma producing HPVs. Methods Polymerase chain reaction assays for the E6 and E7 gene sequences of HPV types 6 and 11 and another assay for the E7 region of HPV types 16, 31, 33, 35, 42, and 58 were used to evaluate decxyribonucleic acid (DNA) extracted from archival pyogenic granuloma biopsies taken from cutaneous and oral epithelium. Results Neither cutaneous nor oral pyogenic granulomas contain amplifiable E6 or E7 sequences from any of these viruses. Conclusions Pyogenic granulomas are not caused by HPV 6, 11, 16, 31, 33, 35, 42, or 58. This study does not exclude the possibility that other viruses may be responsible for these tumors.     

Evidence that dysregulated DNA mismatch repair characterizes human nonmelanoma skin cancer

BACKGROUND: In addition to an established role in the repair of postreplicative DNA errors, DNA mismatch repair (MMR) proteins also contribute to cellular responses to exogenous DNA damage. Previously, we have shown that Msh2-null mice display increased sensitivity to ultraviolet (UV) B-induced tumorigenesis, but squamous cell carcinomas (SCC) generated are microsatellite stable, suggesting a role for MMR other than postreplicative repair in UV-induced cutaneous tumour formation. OBJECTIVES: We questioned whether there was evidence of MMR dysfunction in human SCC, thus validating the mouse models of MMR-dependent UVB-induced skin cancer. METHODS: Using tissue microarrays we examined both nuclear and cytoplasmic levels of MMR proteins MSH2, MSH6, MSH3, MLH1 and PMS2 in more than 200 cases of cutaneous SCC and basal cell carcinoma (BCC). RESULTS: We found that subsets of these 10 MMR protein measures were increased in nonmelanoma skin cancer (NMSC) compared with normal epidermal samples; this was particularly true of SCC. In fact, based on post hoc tests and MMR protein distribution patterns, BCC was distinct from SCC. With the exception of nuclear MSH2, the BCC had lower levels of identified MMR protein measures than SCC. We believe this to be important because not only is SCC more aggressive than BCC, but evidence suggests that these two NMSC subtypes arise through different molecular pathways. CONCLUSIONS: In combination with previously established roles for MMR proteins in response to UVB-induced DNA damage, our data point towards an expanded perspective of the importance of MMR proteins in the suppression of UVB-induced tumorigenesis and, potentially, tumour behaviour.     

Human papillomavirus-DNA loads in actinic keratoses exceed those in non-melanoma skin cancers

Recent studies suggest a role of cutaneous human papillomaviruses (HPV) in non-melanoma skin cancer (NMSC) development. In this study viral DNA loads of six frequent HPV types were determined by quantitative, type-specific real-time-PCR (Q-PCR) in actinic keratoses (AK, n=26), NMSC (n=31), perilesional tissue (n=22), and metastases of squamous cell carcinomas (SCC) (n=8) which were previously shown to be positive for HPV5, 8, 15, 20, 24, or 36. HPV-DNA loads in AK, (partially microdissected) NMSC, and perilesional skin ranged between one HPV-DNA copy per 0.02 and 14,200 cell equivalents (median: 1 HPV-DNA copy per 344 cell equivalents; n=48). In 32 of the 79 HPV-positive skin biopsies and in seven of the eight metastases viral loads were even below the detection limit of Q-PCR. Low viral loads in NMSC were confirmed by in situ-hybridization showing only a few HPV-DNA-positive nuclei per section. Viral loads in SCC, basal cell carcinomas, and perilesional tissue were similar. But, viral loads found in AK were significantly higher than in SCC (p=0.035). Our data suggest that persistence of HPV is not necessary for the maintenance of the malignant phenotype of individual NMSC cells. Although a passenger state cannot be excluded, the data are compatible with a carcinogenic role of HPV in early steps of tumor development.     

Fluorescence diagnosis in actinic keratosis and squamous cell carcinoma

Background: As different tissue types have distinct capabilities to accumulate protoporphyrin IX, fluorescence diagnosis with aminolevulinic acid‐induced porphyrins (FDAP) could be used to discriminate between different types of tissue. Previous results demonstrated higher fluorescence ratios in squamous cell carcinoma (SCC) compared with actinic keratoses (AKs). Objectives: The lesional : non‐lesional fluorescence ratio of AKs was compared with the ratio of SCC. Other factors influencing macroscopic fluorescence were also assessed, including stratum corneum thickness, which has been demonstrated to account for heterogeneous fluorescence in psoriasis and in AKs. Methods: After 1 week of keratolytic pretreatment, FDAP was performed in 13 patients with 36 lesions suspected for AK or SCC. Biopsies were taken for histopathological diagnosis and measurement of stratum corneum thickness. Results: No significant differences were found in the fluorescence ratio (lesional : non‐lesional skin) between AKs and SCCs, although macroscopic fluorescence was significantly higher in Bowen's disease and micro‐invasive SCCs. Conclusions: There could be a potential applicability of FDAP to differentiate premalignant lesions with a tendency to progress into SCC and squamous cutaneous lesions already progressing into early invasive cancer from other squamous cutaneous (pre)malignancies. The amount of hyperkeratosis, invasiveness and degree of differentiation seem to be responsible for variations in fluorescence intensity.     

宫颈癌标本中HPV16的E6、E7和LCR的变异研究

目的:探究分析E6、E7和LCR(long control region,LCR)在宫颈癌标本中HPV16中的变异情况。方法:随机选取我院病理实验室于2015年1月至2016年1月期间留存的HPV16阳性宫颈癌标本100例为研究对象,分别采用PCR技术进行E6、E7和LCR片段的扩增处理,并采用DNA序列进行PCR扩增产物的序列测定,分析E6、E7和LCR的变异表现。结果:E6基因中最常见变异为T350G(67.27%),E7基因中最常见变异为T789C(72.67%),LCR最常见变异为G7521A(90.90%),LCR区中出现G7799A、A7636C、C13T、C7678T新变异,E7区高度保守,YY1转录因子结合点是LCR变异的主要集中点。结论:宫颈癌标本中HPV16存在E6、E7和LCR变异情况,分析高危型HPV变异有助于宫颈癌HPV的早期诊断,可将其应用于宫颈癌防治的疫苗设计中,具有广泛的临床应用前景。     

The role of p53 codon 72 and human papilloma virus status of cutaneous squamous cell carcinoma in the Swedish population

The arginine variant of the p53 codon 72 polymorphism as well as anogenital and epidermodysplasia verruciformis (EV) types of human papilloma virus (HPV) are suggested to confer increased risk for developing cutaneous squamous cell carcinoma (SCC). In this pilot study, we analysed the p53 codon 72 genotype distribution in 106 microdissected samples from normal and tumour tissues of 53 cases of cutaneous SCC and 96 controls from Sweden. Both normal and tumour samples from cases of SCC were screened for anogenital and EV HPV. The p53Arg allele was not associated with the development of cutaneous SCC. Anogenital HPV (44%) was more prevalent than EV HPV (12%). Data also indicate that anogenital HPV is more common in tumour samples, but HPV infection was not identified as a significant risk factor for developing SCC. The presence of anogenital HPV, but not EV HPV might be a risk factor for development of cutaneous SCC.     

人乳头瘤病毒18型E6E7反义RNA表达重组体的构建

目的　为了探讨人乳头瘤病毒 (HPV)的致病机理和寻找由HPV所致疾病的治疗途径 ,研究了HPV1 8型E6E7反义RNA表达重组体的构建。方法　以HPV1 8型全基因质粒为模板 ,聚合酶链反应扩增出HPV 1 8型E6E7区 81 6bp片段 ,以逆转录病毒pLNSX为载体 ,构建HPV 1 8型E6E7逆转录病毒重组体 ,并以限制性内切酶酶切和Southern杂交分别鉴定插入方向和特异性检测。结果和结论　筛选出可表达HPV1 8型E6E7逆转录病毒重组体。该反义RNA表达重组体的构建为进一步研究E6E7基因的作用 ,以及探索是否能通过反义技术调控E6E7基因的表达打下基础。