Host defenses against human papillomaviruses: lessons from epidermodysplasia verruciformis.

Epidermodysplasia verruciformis (EV) is a rare, autosomal recessive genodermatosis associated with a high risk of skin carcinoma (MIM 226400). EV is characterized by the abnormal susceptibility of otherwise healthy patients to infection by specific, weakly virulent human papillomaviruses (HPVs), including the potentially oncogenic HPV-5. Inactivating mutations in either of the related EVER1/TMC6 and EVER2/TMC8 genes cause most EV cases. New insights in EV pathogenesis have been gained from the following recent observations: (1) EV-specific HPVs (betapapillomaviruses) are defective for an important growth-promoting function encoded by an E5/E8 gene present in other HPVs, and inactivation of EVER proteins may compensate for the missing viral function; (2) the transmembrane viral E5/E8 and cellular EVER proteins interact both with the zinc transporter ZnT1, and are likely to modulate zinc homeostasis. EV may thus represent a primary deficiency in intrinsic, constitutive immunity to betapapillomaviruses, or constitute a primary deficiency in innate immunity (or both). Keratinocytes, the home cells of HPVs, are likely to play a central role in both cases. An important issue is to establish which cellular genes involved in intrinsic and innate antiviral responses play a part in the outcome of infections with other HPV types, such as genital oncogenic HPVs.

Acceptance of NCPAP in a sample of patients admitted for geriatric rehabilitation.

OBJECTIVE: Sleep apnea syndrome (SAS) is common in older people. Nasal continuous airway pressure (NCPAP) therapy is the treatment of choice for sleep apnea, but is not always accepted by patients. The rate of successful initiation of NCPAP is unknown in geriatric patients. METHODS: All patients admitted for geriatric rehabilitation were considered for sleep studies. Sleep apnea was assessed using an Edentrace (Nellcor, Hayward, CA) multi-channel recording system. SAS was defined as an apnea-hypopnea-index (AHI) of more than five events per hour plus excessive daytime sleepiness, or an AHI of more than fifteen events per hour regardless of reported sleepiness. Disability was assessed using the Barthel Index of Activities of Daily Living. RESULTS: Two hundred sixty nine of 322 consecutive patients (84%) had adequate sleep studies and gave informed consent. SAS was found in 169 subjects (68%). There was no gender difference in the prevalence of SAS. Six subjects (4%) accepted NCPAP therapy. Individuals who accepted NCPAP were younger and less disabled (p<0.03). Multiple logistic regression analysis revealed disability as the only significant factor predicting NCPAP acceptance. CONCLUSION: NCPAP should not be withheld in the elderly. However, initiation of treatment for SAS remains to be a great challenge in those patients. Geriatric assessment procedures may help better manage older subjects with sleep apnea syndrome.

Cyclical nocturnal oxygen desaturation and impact on activities of daily living in elderly patients.

It is well known, that sleep disordered breathing (SDB) is associated with functional impairment in stroke patients. In elderly non-stroke subjects this relation is unclear. We evaluated 539 elderly patients (median age was 81 years, range 41 to 100 years). We measured activities of daily living (ADL) with the Barthel-Index (BI, 0-100) on admission and discharge, excessive daytime sleepiness (EDS), and the nocturnal oxygen desaturation index (ODI). BI on admission was 54+/-33 in men and 55+/-31 in women (n.s.). More than 50% of the subjects had an ODI above 10/h. The mean BI on discharge was 65+/-31 in men and 67+/-29 in women (n.s.). With increasing SDB severity (quartiles of ODI), BI on admission decreased from 58+/-33 to 46+/-31 (P<0.001) and BI on discharge decreased from 70+/-31 to 59+/-29 (P<0.001). The frequency of EDS increased with increasing severity of SDB from 14.150% (first quartile of ODI) to 40.3% (forth quartile of ODI) (P<0.001). In 330 subjects without EDS, BI on admission did not differ regarding ODI. BI on discharge decreased from 78+/-22 (first quartile of ODI) to 66+/-25 (forth quartile of ODI) (P<0.04). SDB and EDS have a negative and independent impact on activities of daily living in elderly non-stroke subjects. Regarding the high frequency of SDB in the elderly and the effect size on ADL further interventional studies are warranted.

Identification of an immunologically distinct papillomavirus from lesions of epidermodysplasia verruciformis.

Virions isolated from warts of 2 siblings with epidermodysplasia verruciformis (EV), a rare disease characterized by the lifelong growth of warty skin tumors containing papova-like virions, were compared to isolates of human papillomavirus (HPV) from 3 pools of plantar and common hand warts. The length of relaxed, circular (form II) molecules of EV virion DNA approximated the length of HPV DNA but was 3.3% longer. Antisera prepared in rabbits against the 3 HPV pools coated and aggregated HPV in immune electron microscopy (IEM) tests but did not react with EV virions. These antisera reacted at high titers in complement fixation (CF) tests with HPV and reacted only weakly in CF tests with EV virions. Rabbit antisera to EV virions coated and aggregated EV virions but reacted only weakly or not at all with HPV virions in IEM tests. These sera reacted in CF with EV virions only. The data indicated that virions from the EV patients represent an immunologically distinct papillomavirus.

Early regression of cervical lesions in HIV-seropositive women receiving highly active antiretroviral therapy.

OBJECTIVE: Advanced HIV disease is associated with a high prevalence of cervical squamous intra-epithelial lesions (SIL) and of infection with oncogenic human papillomavirus (HPV) genotypes. Triple-combination antiretroviral therapy results in decreased plasma HIV viral load, increased CD4 cell counts and partial restoration of immune functions in patients with severe HIV disease. This study investigated the outcome of SIL in HIV-seropositive women undergoing triple combination antiretroviral treatment. METHODS: Forty-nine women who started triple-combination antiretroviral therapy, including a protease inhibitor, were examined prior to and after a median 5-month treatment. We collected cytological, colposcopic and histologic data and assessed the presence of HPV DNA in cervical smears by PCR and Southern blot hybridization (SBH). RESULTS: The prevalence of SIL decreased from 69 to 53% during follow-up (P < 0.0001). Among 13 women who initially presented with high-grade SIL, conversion to lower grade was observed in two women and a full regression to normality was observed in one. Cytology also returned to normality in nine out of 21 women who initially presented with low-grade SIL. The high prevalence of HPV infection as detected by SBH and PCR was similar at the first and second examinations and the same high-risk viral genotypes were identified at both examinations in all infected patients but one. There was a higher increase in absolute CD4 cells in the subgroup of patients whose lesions regressed (99 versus 50 x 10(6)/l, P=0.03). CONCLUSION: Our observations demonstrate that active antiretroviral therapy may result in a reduced prevalence of cervical squamous intra-epithelial lesions despite the absence of clearance of HPV infection.

Comparative prevalence, incidence and short-term prognosis of cervical squamous intraepithelial lesions amongst HIV-positive and HIV-negative women.

OBJECTIVE: To investigate the impact of HIV infection on the prevalence, incidence and short-term prognosis of squamous intraepithelial lesions (SIL), in a prospective study with 1-year follow-up. METHODS: Between 1993 and 1995, 271 HIV-positive and 171 HIV-negative women at high risk of HIV infection were recruited, 365 (82.6%) of whom completed the 1-year follow-up. The women underwent a Papanicolaou smear test at inclusion and at 6 and 12 months. Human papillomavirus (HPV) was detected at inclusion by Southern blot and PCR. RESULTS: The SIL prevalence ranged from 7.5% for HIV-negative to 31.3% for HIV-positive women with CD4 cell counts < 500 x 10(6)/l (P < 0.001). Other factors associated independently and significantly with SIL prevalence were HPV-16, 18, 33 and related types, HPV-31, -35, -39 and related types, lifetime number of partners, younger age, past history of SIL and lack of past cervical screening. The SIL incidence ranged from 4.9% in HIV-negative women to 27% in HIV-positive women with CD4 cells < 500 x 10(6)/l (P < 0.001). Progression from low- to high-grade SIL during follow-up was detected in 38.1% of HIV-positive women with CD4 cells < or = 500 x 10(6)/l but in no HIV-negative nor HIV-positive women with CD4 cells > 500 x 10(6)/l. HPV-16, 18, 33 and related types were also associated with higher incidence of SIL and progression from low- to high-grade SIL. CONCLUSION: HIV-induced immunodeficiency is associated with high prevalence, incidence and persistence/progression of SIL. A pejorative influence of HIV infection without marked immunodeficiency is less clear. HIV-positive women with SIL may thus benefit from early treatment when a useful immune response is still present.

Clinical observations, virologic studies, and treatment trials in patients with epidermodysplasia verruciformis, a disease induced by specific human papillomaviruses.

We have studied 11 patients with the papillomavirus-induced disease epidermodysplasia verruciformis (EV). Clinical diagnostic features are widespread, long-lasting, pityriasis versicolor-like macules and flat, wart-like papules, both usually occurring in early childhood, with the subsequent development in the third decade of multiple skin cancers of the Bowenoid in situ and squamous cell types, primarily in sun-exposed skin. Virologic studies using the methods of immunofluorescence microscopy, restriction endonuclease analysis, and DNA blot hybridization have shown benign lesions to be associated with one or several types of the human papillomaviruses (HPVs) specifically associated with EV (at least 15 types recognized on the basis of sequence homology studies of molecularly cloned genomes). Skin cancers in these patients were associated with the genomes of either HPV-5, HPV-8 or HPV-14, suggesting that these three viruses are potentially oncogenic. A genetic factor appears to play a role in the pathogenesis of EV, since 5 of our patients were children of consanguineous parents and 2 had siblings also suffering with EV, suggesting a recessive inheritance pattern. Treatment of 4 EV patients with an oral retinoid resulted in partial temporary improvement of benign lesions, and the treatment of 2 patients with intralesional interferon injections into multiple Bowenoid cancers in situ has resulted in the disappearance of these lesions. Finally, EV serves as a model for studying the interplay of oncogenic viruses, genetic and immunologic factors, and sunlight in the production of skin cancer in humans.

The search for a culture system for papillomavirus.

Papillomaviruses induce tumors of keratinocytes. Vegetative viral DNA replication and virion assembly are seen in those cells which are in the process of keratinizing or are keratinized. To date, no cell culture system has been developed that permits expression of the complete viral life cycle. Keratinocytes infected in culture may harbor the virus as a stable, replicating episome, but they do not support vegetative viral growth, nor do they become immortalized or transformed. The major obstacle in using keratinocyte cultures may be related to a dual need for transformation and full differentiation. Some animal papillomaviruses have been shown to be capable of transforming cultured murine fibroblasts. The fibroblast model is useful for identifying the viral-transforming gene(s) and their products.

Premalignant lesions and cancers of the skin in the general population: evaluation of the role of human papillomaviruses.

To evaluate the role of human papillomaviruses (HPV) in the development of premalignant lesions and cancers of the skin in the general population, 314 biopsies obtained from 227 patients with benign neoplasms, premalignant lesions, and cancers of the skin and from 25 patients with squamous cell carcinoma of the lip were analyzed by Southern blot hybridization. DNA probes specific for various cutaneous and genital HPV types were used in hybridizations conducted under nonstringent or stringent conditions. HPV DNA sequences were only detected in eight specimens obtained from six patients: HPV 34 in one case of periungual Bowen's disease, HPV 36 and an as yet uncharacterized HPV in two cases of actinic keratosis, HPV 20 in one case of basal cell carcinoma, an as yet unrecognized HPV in one case of squamous cell carcinoma, and HPV 16 in one case of squamous cell carcinoma of the lip. None of the specimens of cutaneous horn and keratoacanthoma contained detectable HPV DNA. In contrast, HPV DNA sequences, mostly HPV 16, were detected in 13 of 23 cases of anogenital Bowen's disease and invasive Bowen's carcinoma. HPV DNA sequences were not detected in 90 cutaneous samples further analyzed by the polymerase chain-reaction technique, using amplification primers that contain conserved sequences among the genomes of HPV. These results strongly suggest that the known HPV types play only a minor role, if any, in skin carcinogenesis in the general population.

Cytopathic effect in human papillomavirus type 1-induced inclusion warts: in vitro analysis of the contribution of two forms of the viral E4 protein.

Myrmecia warts induced by human papillomavirus type 1 (HPV1) are characterized by abundant eosinophilic inclusions associated with HPV1 E4 gene products. The major HPV1 E4 proteins are a 17-kilodalton (kDa) E1-E4 fusion protein and a 16-kDa species lacking the five E1 amino acids and a few E4 residues. To study the contribution of E4 proteins to the formation of myrmecia inclusions, we used a previously designed transient expression system in the rabbit VX2-R keratinocyte line. We find that the E1-E4 and an E4 protein without the E1 residues (E4-3200) form eosinophilic inclusions. Ultrastructural and immunoelectron microscopic studies show that the electron-dense, keratohyalin-like myrmecia inclusions are recognized by anti-E4 antibodies. They are associated with tonofilament bundles at their periphery in the cytoplasm or free of filaments in the nucleus. The E1-E4 inclusions formed in vitro are also homogeneously electron dense, and are usually associated with tonofilaments at their periphery in the cytoplasm and free of filaments in the nucleus. The E4-3200 inclusions are exclusively cytoplasmic and heterogeneously electron dense, with a fibrillar structure made of entangled 10-nm filaments. The expression of either protein in VX2-R cells does not result in the collapse of the cytokeratin network, as shown by immunofluorescence double-labeling experiments. This is in contrast to data reported for the HPV16 E1-E4 protein. Our findings indicate that the E1-E4 protein by itself accounts for the formation of myrmecia inclusions, and suggest that the five N-terminal E1 amino acids play a major role in the interaction of E4 proteins with intermediate filaments.

Analysis of the nucleotide sequence variation of the antigen-binding domain of DR alpha and DQ alpha molecules as related to the evolution of papillomavirus-induced warts in rabbits.

We previously found that regression of skin warts induced by the Shope cottontail rabbit papillomavirus in New Zealand White rabbits, as well as malignant conversion of persistent warts, are linked to a restriction fragment length polymorphism of the major histocompatibility complex class II DR alpha and DQ alpha genes. To find out whether this immunogenetic control could be connected with the antigen binding and presentation function of the alpha 1 domain of class II molecules, we have sequenced the exon 2 of the four DR alpha EcoRI and six of the seven DQ alpha PvuII restriction fragment length polymorphism alleles identified, and deduced the encoded amino acid sequences. We found no amino acid polymorphism among DR alpha alleles, indicating that the alpha 1 domain of the DR alpha chain does not condition wart regression or cancer development. In contrast, 27 of the 82 amino acids of the DQ alpha 1 domain were found variable, defining five amino acid sequence alleles. The restriction fragment length polymorphism allele linked to regression and another allele not linked to regression share the same alpha 1 domain, indicating that wart regression is rather conditioned by a closely linked gene. The most divergent DQ alpha 1 allele, however, was that associated with a higher risk of cancer. Alignment of rabbit and human DQ alpha exon 2 alleles disclosed that amino acid charge variations occur at positions assumed to be important for peptide binding in humans. By modulating the affinity for tumor-specific antigenic peptides, such transitions could affect immune surveillance and, thus, condition the risk for progression to carcinoma of papillomavirus-associated lesions.

A susceptibility locus for epidermodysplasia verruciformis, an abnormal predisposition to infection with the oncogenic human papillomavirus type 5, maps to chromosome 17qter in a region containing a psoriasis locus.

Epidermodysplasia verruciformis (EV) is a rare genodermatosis characterized by an abnormal susceptibility to infection with a specific group of related human papillomavirus (HPV) genotypes, including the oncogenic HPV5 associated with the skin carcinomas developing in about half of EV patients. EV is usually considered as an autosomal recessive condition. Taking EV as a model to identify a locus underlying the susceptibility to HPV infections, we performed a genome-wide search for linkage with 255 microsatellite genetic markers in three consanguineous EV families comprising six patients, using the homozygosity mapping approach. Homozygosity restricted to affected individuals was observed for a marker of chromosome 17q (D17S784) in two families and a marker about 17 centiMorgan (cM) distal (D17S1807) in the third family. Ten additional microsatellite markers spanning 29 cM in this region were analyzed. Two-point lod score values greater than 3 were obtained for four markers and multipoint linkage analysis yielded a maximum lod score of 10.17 between markers D17S939 and D17S802. Recombination events observed in two families allowed a candidate region for the EV susceptibility locus to be mapped to the 1 cM region defined by these two markers. The EV locus (named EV1) is included in the 17qter region recently found to contain a dominant locus for the susceptibility to familial psoriasis. It has been shown that patients suffering from psoriasis are likely to constitute the reservoir of HPV5. It is thus tempting to speculate that distinct defects affecting the same gene may be involved in the two skin conditions.

Antibodies to human papillomavirus type 5 are generated in epidermal repair processes.

We reported previously that patients with psoriasis harbored at a very high frequency DNA sequences of the oncogenic human papillomavirus type 5 (HPV5) associated with epidermodysplasia verruciformis. Moreover anti-HPV5 antibodies were detected in 25% of the cases. Our aim was to find out whether keratinocyte hyperproliferation and/or autoimmunity could be responsible for HPV5 expression in psoriasis. We found that epidermal repair in patients with extensive second degree burns (n = 19) is frequently associated with the generation of anti-HPV5 antibodies. In patients with autoimmune bullous diseases (n = 118), a condition in which keratinocyte proliferation is involved in repair mechanisms, the prevalence of anti-HPV5 antibodies (15%-25%) was similar to that reported in psoriasis and significantly higher than that (5%) observed in individuals with no known history of human papillomavirus infection (n = 119). A high detection rate (57.9%) of HPV5 DNA was observed in patients with bullous diseases. Anti-HPV5 antibodies were found in patients with autoimmune connective tissue disorders with cutaneous involvement (n = 40) as frequently as in patients with bullous diseases. HPV5 DNA was detected in one of the 10 patients studied. In contrast, the prevalence of anti-HPV5 antibodies in patients with autoimmune neurological disorders (n = 47) and in patients with common warts (n = 28) or invasive carcinomas of the skin (n = 40) was as low as in the general population. It is worth stressing that a similar prevalence of antibodies against HPV1 was found in all groups studied. Our data strongly suggest that extensive keratinocyte proliferation is a major factor for the generation of anti-HPV5 antibodies and that autoimmunity may contribute to this phenomenon. It remains to be determined whether HPV5 and other human papillomavirus genotypes associated with epidermodysplasia verruciformis contribute to the hyperproliferation of keratinocytes occurring in epidermal repair and in psoriasis.

Identification of papillomaviruses in butchers' warts.

We have studied the papillomaviruses found in the hand warts of 60 butchers, most of them from 2 distant slaughterhouses. Warts differing in morphology and location were studied separately. The viruses were identified by molecular hybridization, restriction enzyme analysis and immunofluorescence. Four known human papillomaviruses (HPV-1, HPV-2, HPV-3, HPV-4) were detected and one hitherto unknown papillomavirus was identified in 9 butchers. The DNA of the latter virus did not anneal with any of the RNAs complementary to either HPV-1 to HPV-5 or bovine papillomavirus type 1 (BPV-1) DNAs, and showed a Hind II + III restriction enzyme cleavage pattern distinct from those of known HPVs and BPVs. This virus showed distinct antigenic properties, as shown by immunofluorescence, using HPV-1, -2, -3, -5, and BPV-1 antisera. It may represent a new type of human papillomavirus (HPV-7) or a yet unidentified animal papillomavirus. In addition, 6 butchers were found to be infected with a papillomavirus, distinct from the known skin HPVs and from BPV-1, which could not be characterized by restriction enzyme analysis. Eleven butchers were found to be infected by 2 viruses. A characteristic histological pattern was found to be associated with the different papillomaviruses.

Psoriasis: A possible reservoir for human papillomavirus type 5, the virus associated with skin carcinomas of epidermodysplasia verruciformis.

Recent polymerase chain reaction data have shown that most human papillomavirus (HPV) genotypes associated with epidermodysplasia verruciformis (EV) are widespread; however, HPV5 associated with EV skin carcinomas has only rarely been detected in non-EV patients. To identify the reservoir of this virus, we examined 335 sera from different groups of patients for the presence of HPV5 antibodies by an enzyme-linked immunosorbent assay test based on HPV5 virus-like particles. The prevalence of antibodies reacting with HPV5 virus-like particles was found to be significantly higher in psoriatic patients (24.5%) than in other groups (2-5%), including patients with atopic dermatitis and renal transplant recipients. Analysis of scrapings of lesional and uninvolved skin by a nested polymerase chain reaction method, using degenerate EV HPV primers, disclosed HPV DNA in 91.7% of 48 psoriatic skin samples and 35.5% of 31 atopic dermatitis specimens. Eleven EV HPV genotypes, most frequently HPV5 and HPV36, and a putative novel genotype (PsoX1) were identified in psoriasis. Five EV HPV genotypes and two putative novel genotypes (ADX1 and ADX2) were detected in atopic dermatitis patients. HPV5 was not found in atopic dermatitis patients. Using type specific primers, HPV5, HPV36, and HPV1 were found in 89.4%, 84.2%, and 42.1% of specimens from psoriatic patients, whereas HPV36 was detected in 22.5% of specimens from atopic dermatitis patients. HPV16 was never detected. On the whole, 27 HPV5 and 13 HPV36 DNA variants were disclosed after sequencing amplification products. Our data confirm that EV HPV are widespread and point to psoriasis as a reservoir for HPV5. Whether HPV5 is involved in the pathogenesis of psoriasis remains to be determined.

A possible vertical transmission of human papillomavirus genotypes associated with epidermodysplasia verruciformis.

Epidermodysplasia verruciformis (EV) is characterized by an abnormal genetic susceptibility to a group of related human papillomavirus (HPV) genotypes, including the oncogenic HPV5 and HPV8. The mode of transmission of these viruses remains unknown. In view of the rare incidence of EV, we had a unique opportunity to perform a virologic study of the amniotic fluid and placenta from an EV patient infected with HPV5, HPV8, several other EV HPV, and HPV3. The child was born by cesarean section and the amniotic fluid specimen was taken prior to rupture of membranes. Analysis of the amniotic fluid and placenta specimens by a nested polymerase chain reaction method, using degenerate EV HPV primers or type-specific HPV primers, disclosed the presence of the variants of EV HPV5, HPV8, HPV24, and HPV36, and of HPV3 detected in the skin lesions of the patient. HPV5, HPV8, HPV24, and HPV3 were also detected in the placenta. No viral sequences were detected in peripheral blood mononuclear cells collected 2 y and 6 mo before cesarean section, rendering an hematogenous transmission unlikely. The same HPV variants were also detected in cervical scrapes taken from the patient, which may suggest an ascending infection of the placenta. This first report of the detection of EV HPV in amniotic fluid, placenta, and cervical scrapes from an EV patient renders vertical transmission of EV HPV likely.

A new type of human papillomavirus associated with oral focal epithelial hyperplasia.

Lesions from 10 patients suffering from focal epithelial hyperplasia (FEH) of the oral mucosa, including those of 4 Greenlandic Eskimos, were investigated for the presence of human papillomavirus (HPV) DNA sequences by blot hybridization experiments. Two distinct HPVs were detected in the DNA extracted from these lesions, and their genomes were molecularly cloned and characterized. One of these HPVs, detected in 4 patients, was found to be identical with HPV13, whose association with FEH was already known. The other one, detected in 6 patients, was only weakly related to HPV13 and to the other HPVs associated with lesions of the mucous membranes, and constituted a new HPV type, tentatively named HPV32. Lesions from other types of oral papillomas, obtained from 14 additional patients, were also analyzed. Human papillomavirus DNA sequences were detected in the DNA preparations extracted from 5 specimens: HPV6 DNA in a condyloma and in a papilloma, 2 as yet uncharacterized HPV DNAs in 2 papillomas, and HPV32 DNA in a papilloma which showed histologic similarities to FEH. Thus, it seems likely that FEH of the oral mucosa is a disease associated with 2 specific HPVs--HPV13 and HPV32.

TGF beta-1 and TNF alpha expression in the epidermis of patients with epidermodysplasia verruciformis.

In epidermodysplasia verruciformis (EV), the infection with specific human papillomaviruses (HPV) might be under control of the local immunosurveillance mechanisms related to cytokines produced by epidermal cells. We have investigated by in situ hybridization the expression of mRNA coding for TGF beta-1 and TNF alpha in the skin of patients with EV (n = 4) as compared to the skin lesions of patients with other premalignant (actinic keratosis; n = 5) or malignant (squamous cell carcinoma; n = 4) skin lesions, and to the skin of healthy individuals (n = 5). The expression of TGF beta-1 and TNF alpha mRNA was higher in the epidermis of EV patients as compared to the control skin from healthy individuals. The increased expression of mRNA for both cytokines was confirmed by northern blot analysis of RNA isolated from the skin lesions of the patient with EV. No specific signals for TGF beta-1 and TNF alpha were detected in actinic keratosis, and in cases of squamous cell carcinomas only single neoplastic cells were positive for TGF beta-1. It is conceivable that in EV TGF beta-1 and TNF alpha can be involved in the regulation of the growth and differentiation of HPV-infected keratinocytes and in the persistence of HPV-induced skin lesions.

Evidence for a nonallelic heterogeneity of epidermodysplasia verruciformis with two susceptibility loci mapped to chromosome regions 2p21-p24 and 17q25.

Epidermodysplasia verruciformis is a rare genodermatosis associated with a high risk of skin cancer. This condition is characterized by an abnormal susceptibility to specific related human papillomavirus genotypes, including the oncogenic HPV5. Epidermodysplasia verruciformis is usually considered as an autosomal recessive disease. We recently mapped a susceptibility locus for epidermodysplasia verruciformis (EV1) to chromosome 17qter within the 1 cM interval between markers D17S939 and D17S802. We report here the genotyping for 10 microsatellite markers spanning 29 cM around EV1 in two consanguineous epidermodysplasia verruciformis families from Colombia (C2) and France (F1) comprising five patients and two patients, respectively. Using homozygosity mapping, linkage with 17qter markers was observed for family C2 only. Multipoint linkage analysis yielded maximum multipoint LOD-score values above 10 between markers D17S1839 and D17S802 encompassing the EV1 locus. A genome-wide search performed in family F1 yielded evidence for linkage between epidermodysplasia verruciformis and the chromosomal 2p marker D2S365. Nine additional microsatellite markers spanning 15 cM in this region were analyzed. Assuming an autosomal recessive inheritance with a complete penetrance, the expected maximum two-point LOD-score value of 1.8 was obtained for three markers and multipoint linkage analysis yielded a maximum LOD-score value of 3. 51 between markers D2S2144 and D2S392. Haplotype analysis allowed to map a candidate region for a second epidermodysplasia verruciformis susceptibility locus (EV2) within the 8 cM interval between markers D2S171 and D2S2347 of the 2p21-p24 region. In contrast, linkage with 2p markers was excluded for family C2 and for the three families in which we mapped EV1 previously. The disclosure of two susceptibility loci for epidermodysplasia verruciformis provides evidence for a nonallelic heterogeneity in this disease.

Human papillomavirus (HPV) types specific of epidermodysplasia verruciformis detected in warts induced by HPV3 or HPV3-related types in immunosuppressed patients.

Epidermodysplasia verruciformis (EV) is characterized by an abnormal genetic predisposition to infection with specific types of human papillomavirus (HPV). Specific defects of the cell-mediated immunity and/or of the control of HPV infection in keratinocytes are assumed to be involved in the development of the disease. As a model to test this hypothesis, we have studied the prevalence of EV-specific HPV in skin warts of 56 immunosuppressed patients. All main types of cutaneous HPV (HPV1, 2, 3, 4, 10, and 28) responsible for skin warts in the general population were detected by blot hybridization. EV-specific HPV (HPV5, 20, and 23) were detected in three patients. Four additional patients were found infected with HPV49, first characterized in the course of this study, and found to be related to EV HPV. A most important finding was that HPV5, 20, 23, and 49 were always codetected with HPV3 or the related types HPV10 and 28. None of the specimens showed the typical clinical morphology of EV lesions. In none of these specimens was the specific cytopathic effect of EV recognized; instead that of HPV3 and related types was seen. No evidence for productive EV HPV DNA replication was obtained for the three specimens that could be further analyzed by in situ hybridization. Our data suggest that HPV3 infection favors infection with EV HPV in immunosuppressed patients but that the full expression of EV HPV is usually restricted as in the general population.