[Simultaneous detection and typing of human papillomavirus in cervical biopsies using PCR-reverse hybridization]. / Simultanea ricerca e tipizzazione del papillomavirus umano in biopsie cervicali mediante PCR-ibridazione inversa.

INTRODUCTION: Genotyping of Human papillomavirus (HPV) is an important step in the clinical evaluation of the oncogenic risk associated with HPV infection of cervical mucosa. The purpose of this work was to develop a fast PCR-reverse-hybridization assay (PCR-RH) for the simultaneous detection and genotyping of anogenital HPVs. METHODS: HPV DNA from cervical biopsies was amplified by consensus primer-PCR. Digoxigenin-labeled PCR products were hybridized to type-specific probes anchored to the surface of plastic microwells and revealed by an ELISA system. RESULTS: The method was tested on 115 clinical samples (81 koilocytic atypias, 11 CIN1, 10 CIN2, 12 CIN3 and 1 squamous carcinoma). HPV DNA was found in 56.7% koilocytic atypias, in 90.9% of CIN1 and in 100% of CIN2 and higher-grade lesions. Thus, PCR-RH is sensitive, rapid, easy-to-perform and readily applicable to the routine analysis of a large number of samples.

The enhancing effect of fasting/refeeding on the growth of nodules selectable by the resistant hepatocyte model in rat liver.

Caloric restriction causes a generalized decrease in growth rate and has been shown to delay the development of both spontaneous and induced neoplasia. In contrast to chronic food restriction, the extreme condition of fasting/refeeding is associated with an overall increase in cell turnover in several organs, including liver, compared with regular feeding. The present study was therefore designed to investigate the effect of complete food withdrawal followed by refeeding on the growth of hepatocyte nodules in initiated rat liver. Male Fischer 344 rats were given a single dose of diethylnitrosamine (DEN, 200 mg/kg i.p.) and then, starting 1 wk later, they were exposed to one or three cycles of fasting (3 days) followed by refeeding (11 days). The control group was fed continuously. Seven weeks after DEN administration all rats were subjected to the resistant hepatocyte model (2-acetylaminofluorene coupled with CCl4) and 2 weeks later 2/3 partial hepatectomy (PH) was performed. All animals were killed 2 weeks after surgery. At PH rats given one cycle of fasting/refeeding had significantly larger glutathione S-transferase 7-7-positive hepatic lesions compared with controls (mean area 0.73 +/- 0.04 versus 0.50 +/- 0.05 mm2, P < 0.025; mean percent area 25.6 +/- 3.2 versus 12.4 +/- 0.9, P < 0.005), while no significant change was observed in their number. The observed differences were more pronounced with three cycles of fasting/refeeding. A similar pattern of results was obtained at the time of killing. It is concluded that fasting/refeeding can exert a positive effect on the growth of rat hepatocyte foci and nodules, in contrast to the general inhibitory effect on carcinogenesis caused by food restriction.

Influence of dehydroepiandrosterone on G-6-PD activity and 3H-thymidine uptake of human lymphocytes in vitro.

Dehydroepiandrosterone (DHEA) was found to inhibit experimental cancer development in mouse and rat lung, colon and mammary gland. Since DHEA is a potent inhibitor of mammalian G-6-PD, the hypothesis that the compound could inhibit cell proliferation through an inhibition of the pentose phosphate pathway has been formulated. We studied the effects of DHEA on the proliferation in vitro of human lymphocytes induced by several mitogens (PHA, ConA and PWM), measuring 3H-thymidine uptake. DHEA inhibited 3H-thymidine uptake of mitogen-stimulated cells from both G-6-PD+ and G-6-PD- (mediterranean type deficiency) individuals in a dose-dependent and reversible fashion. The inhibitory effect was found even if DHEA was added to cells in the last hours of culture, simultaneously with the addition of 3H-thymidine. These data suggest that the inhibition of thymidine uptake induced by DHEA on human lymphocytes probably does not depend on the inhibition of G-6-PD.