Bovine papillomavirus type 4 in oesophageal papillomas of cattle from the south of Italy.

Oesophageal papillomas are known to occur in cattle infected with bovine papillomavirus type 4 (BPV-4), and BPV-4 papillomas may undergo malignant progression in cattle that feed on bracken fern. In the south of Italy, where bracken fern is common, examination of 1133 slaughterhouse cattle aged 4-12 years revealed oesophageal lesions (single or multiple peduncuolated proliferations, or mucosal thickening) in 147 (13%). These two types of lesion were consistent with exophytic and inverted papilloma, respectively. BPV-4 was detected by polymerase chain reaction (PCR) analysis in >60% of the samples in which oesophageal papilloma was diagnosed histopathologically. Nucleotide sequencing of the PCR amplicons confirmed the presence of BPV-4 in the papillomas. This is the first report of such infections in a European country other than Britain.

HPV5b variant in a neoplastic lesion of an Italian patient affected by epidermodysplasia verruciformis.

Epidermodysplasia verruciformis (EV) can be defined as a genetic disorder that determines a lifelong infection of the skin by human Papillomaviruses (HPV). The benign lesions contain different HPVs; whereas in the tumors HPV types 5, 8, and, much less frequently, types 14, 17, 20 and 47 can be detected. Variants of HPV5 have been recognized on the basis of the genetic heterogeneity of the E6 open reading frame. We report a typical case of EV in which the presence and expression of the HPV type 5 were clearly detectable. Direct sequence analysis demonstrated a perfect homology with the sequence of the HPV5b variant. This variant was first isolated in a Japanese patient and thereafter in a Polish one. Its presence in the Italian patient indicates that the same variant can be detected in different geographic areas and therefore that some cellular genes may exert a selection pressure so strong as to induce the emergence of this stable HPV5b infectious variant.

Detection of HPV in genital condylomata: correlation between viral load and clinical outcome.

High DNA viral load in Human Papillomavirus type 16 cervical dysplastic lesions has been proposed as a marker of progression towards malignancy. To evaluate the prognostic value of DNA viral load in infection by HPV type other than HPV-16, a longitudinal study of patients with Anogenital Condylomata (AC) was designed. The study included 40 patients, 25 men and 15 women. Clinical data were collected through standard interview and routine serum analyses. Viral type in lesions was assayed by PCR with general primers (MY09-MY11) and Restriction Fragment Length Polymorphism (RFLP). Viral load was determined by PCR end point determination with type specific primers on serial sample dilutions. The duration of ano-genital lesions upon liquid nitrogen standard cryotherapy or Podofillin chemotherapy was correlated with viral and clinical data. Patients with higher lesional viral load at study entry showed a markedly longer disease clinical course than patients with lower viral loads. The HPV-6 was observed in 31 out of 40 patients (77.5%), much more frequently than HPV-11 (7/40 17.5%), the latter being associated with higher titres of viral load. Two cases (5%) remained undetermined. No HPV-16 or other high-risk type was detected, while HPV-61, not uncommon in Italy, was found once in a double infection carried by an HIV positive patient. No significant association was observed between clinical outcome and demographic and epidemiological data, or risk factors.

Egg yolk antibodies against the E7 oncogenic protein of human papillomavirus type 16.

The E7 oncoprotein is the major transforming protein of Human Papillomavirus type 16 (HPV16) and the most abundant in cervical neoplasia. In this study we report the production of polyclonal antibodies to HPV16 E7 in rabbits and hens. The produced antibodies recognised the denatured and native form of HPV16 E7 protein by Western Blot, and immunoprecipitation. Epitope mapping demonstrated that hen antibodies reacted with a greater number of antigen determinants than the rabbit antibodies. In immunocytochemistry only hen antibodies were able to localize the E7 protein in a HPV positive cell line and in the high-grade squamous intraepithelial lesions, suggesting their possible usefulness in the screening of clinical samples.

The pagetoid variant of urothelial carcinoma in situ of urinary bladder in a cow.

A case of urothelial carcinoma in situ of urinary bladder is reported in a 10-year-old cow naturally grazing on bracken-infested land. The cow suffered from enzootic hematuria for more than 5 years. The presence of bovine papillomavirus type 2 (BPV-2) DNA sequences was detected by polymerase chain reaction. The carcinoma in situ was characterized by the presence of anaplastic cells with amphophilic cytoplasm and pleomorphic nuclei containing granular, irregularly dispersed chromatin. Focal areas within the tumor contained large isolated and/or clustered cells. These cells had pale acidophilic cytoplasm, large nuclei with single or multiple nucleoli, and well-defined borders resembling Paget's cells. Immunohistochemically, all malignant cells were negative for vimentin and S-100 and positive for cytokeratins. In addition, normal and neoplastic cells expressed fragile histidine triad (FHIT) protein; surprisingly, some pagetoid cells did not. FHIT, the tumor suppressor gene at 3p14.2, encodes a protein of 147 amino acids (16.8 kd) with diadenosine triphosphate hydrolase activity and is a common target of deletions in human cancers of epithelial origin. Antibody to laminin detected a continuous epithelial basement membrane, thus clearly showing that neoplastic changes were limited to urothelial cells without invading stromal tissue. To our knowledge, this is the first report of an unusual pattern of spread of urothelial carcinoma in situ in a cow.

Expression of endothelin 1 and endothelin A receptor in HPV-associated cervical carcinoma: new potential targets for anticancer therapy.

Human papillomaviruses (HPV) are associated with cervical cancer and interact with growth factors that may enhance malignant transformation of cervical carcinoma cells. Endothelin-1 (ET-1) is released from HPV transfected keratinocytes and induces increased growth response in these cell lines in comparison with normal cells. In the present study several cervical carcinoma cell lines have been analyzed to investigate the expression of ET-1 and its receptors as well as their involvement in tumor growth. All HPV-positive cancer cells secreted ET-1 and expressed mRNA for ET-1 and its receptors, whereas a HPV-negative carcinoma cell line expressed only the ETBR mRNA and didn't secrete ET-1. Binding studies showed that HPV-associated cells expressed an increased number of functional ETAR. ET-1 stimulated a marked dose-dependent increase in [3H]-thymidine incorporation with respect to the normal cells whereas ET-3 and ETBR agonists had no effect. In HPV-positive cancer cells, a specific antagonist of ETAR inhibited the proliferation induced by ET-1 and substantially reduced the basal growth rate of unstimulated cervical tumor cells, whereas the ETBR antagonist had no effect. These results demonstrate that ET-1 participates in the progression of neoplastic growth in HPV-associated carcinoma, in which ETAR are increased and could be targeted for antitumor therapy.

Low prevalence of islet autoimmunity in adult diabetes and low predictive value of islet autoantibodies in the general adult population of northern Italy.

AIMS/HYPOTHESIS: To assess the prevalence of islet autoimmunity in adult-onset diabetes mellitus and the predictive value of islet autoantibodies in the general adult population of northern Italy. METHODS: A sample of 2076 people aged 40 years or more participating in the population-based Cremona Study and classified in 1990 as having diabetes mellitus, impaired and normal glucose tolerance according to WHO criteria after an oral glucose tolerance test, were tested for antibodies to glutamic acid decarboxylase and IA-2. RESULTS: Increased concentrations of glutamic acid decarboxylase antibodies were found in 4 (2.8 %) of 143 participants with known diabetes and none of 50 with previously unknown diabetes, 1 (0.65%) of 153 with impaired and 18 (1.0%) of 1718 with normal glucose tolerance. The increased prevalence of these antibodies in subjects with known diabetes was not statistically significant. Protein tyrosine phosphatase IA-2-antibodies were found in only four subjects, two of whom also had glutamic acid decarboxylase antibodies, all with normal glucose tolerance. After 8 years of follow-up, none of 21 non-diabetic subjects with either glutamic acid decarboxylase or IA-2-antibodies had developed diabetes and only a slight deterioration from normal to impaired fasting glucose was observed in 3 of 15 subjects with previous normal glucose tolerance. CONCLUSION/INTERPRETATION: This study has shown that in northern Italy the prevalence of adult autoimmune diabetes in the general adult population is 0.19% (95 % CI 0.05-0.5); that autoimmune diabetes represents only a minority of all cases of adult diabetes; and that islet autoantibodies are not a high-risk factor for diabetes development in adults with normal glucose tolerance over 8 years of follow-up.

The E5 oncoprotein of human papillomavirus type 16 enhances endothelin-1-induced keratinocyte growth.

Human keratinocytes express ETA receptors and produce endothelin-1 (ET-1), which stimulates growth response. Previously, we reported that a twofold increase in ETA receptors is present in human papillomavirus type 16 (HPV16) immortalized keratinocytes and that ET-1 induces enhanced proliferative response in these cell lines compared to normal cells. The present studies examine whether the E5 gene of HPV16 is responsible for the enhanced activity of ET-1 in HPV-transfected keratinocytes. The presence of the E5 gene in growth factor-starved keratinocytes induced the DNA synthesis and enhanced the mitogenic activity of ET-1 or epidermal growth factor. The selection of primary keratinocytes in growth factor-free medium with the addition of ET-1 as a growth factor showed that E5-transfected keratinocytes were able to grow and to form a higher number of larger colonies with respect to untransfected cells. This effect seems to be related to the interaction of E5 with the mitogenic signaling pathway of ET-1 rather than to an increase in the expression of the receptors for ET-1. In conclusion, our data demonstrate that E5 enhances ligand signaling in keratinocytes outside the EGF pathway by the amplification of the proliferative effect of ET-1/ETA receptor signaling.

HPV 16 E7 antibody levels in cervical cancer patients: before and after treatment.

Antibody response to HPV16 E7 oncoprotein may represent a marker of cervical cancer. A HPV16 GST-E7 fusion protein was used in a Western Blot assay to analyse the HPV16 E7 antibody response in 30 patients before and after treatment for cervical carcinoma (stage IIB or IIIB). Patients were treated with three courses of cisplatin/bleomycin therapy followed by surgery, or with surgery alone. Thirteen out of 30 patients had serum antibodies to HPV16 E7 antigen. Three months after chemotherapy little or no change in antibody titre was detected. In contrast, after surgery, a significant decrease in antibody titre was observed in 9/10 patients. In two cases the titre declined to zero 3 and 9 months after treatment, respectively. These results confirm the usefulness of studying anti-HPV16 E7 antibody profile in cervical cancer patients and suggest that the serum response correlates with tumour burden.