In this study, anal brushings were examined for HPV-DNA, the spectrum of HPV genotypes involved, and type-specific viral load to show, at baseline, determinants of HPV infection in HIV-positive individuals that could constitute potential risk factors for progression.
To our knowledge, these are the first determinations of type-specific quantitative viral loads from low-risk HPV and from a number of high-risk HPV (HPV 53, 58, 59, 66) in male anal samples. The HPV copy numbers in samples were expressed per nanogram of total input DNA, to take into account the variable amounts of cells taken during brushing of the anal canal. There were wide variations between determinations, consistent with previous results from cervical samples [11,18]. Interestingly, in many samples, low-risk HPV had quite high DNA copy numbers, similar to high-risk HPV. This finding could be explained by increased HPV persistency in anal versus cervical infections, as stated in a recent report in HIV-infected women . Indeed, high viral load has been related to HPV persistency in cervical lesions and to an increased risk of progression towards cervical cancer precursors [10–12]; however, only the HPV 16 viral load seems to be predictive of future squamous abnormalities at a stage when a Papanicolaou smear tested negative .
The elevated level of HPV-DNA also found in low-risk HPVs in anal samples is in keeping with papers reporting that infections with low-risk HPVs were an independent risk factor for anal intraepithelial neoplasia [27–29]. Moreover, HPV copy number mean values did not differ significantly between HIV-positive and HIV-negative patients, even though the high variability in the measurements could have hidden a trend. Taken together, these observations apparently contrast with the classic view of HIV infection as a high-risk condition for developing HPV-related malignancies; however, they are in keeping with reports stating that restoration of immunity during HAART did not lower the relative risk of developing high-grade lesions and cancer [2,30]. Indeed, the role of the immune system seems to be exerted at different sites and stages during HPV's natural history and, in recent years, the local milieu of anogenital mucosa has been recognized as playing a pivotal role in HPV infections . As seen in HIV-seropositive women, immune functions in epithelial tissues are not fully accounted for by circulating CD4 T-cell counts and HIV-RNA levels [31,32] and are very complex to monitor. Several studies have reported somewhat conflicting data when measuring inflammatory markers, numbers of CD4 and CD8 T cells, macrophages, Langerhans cells in the cervix of HIV-infected women, either HPV coinfected or not [33–35]; no such studies have been undertaken in anal mucosa. Other possible explanations for the diminished capability of HIV-infected individuals to resolve HPV infections could be a lack of immune response to specific HPV antigens  or differential regulation of HPV-DNA expression due to HIV proteins [37,38].
In conclusion, HPV-DNA tests, and eventually HPV load measurements, should also be performed in university clinics in the absence of dysplastic lesions in anal canal cells, at least in HIV-positive homosexual men. Information on HPV genotypes in subsequent tests can discriminate between persistent and reinfections and may lead to the identification of the high-risk and low-risk genotypes in anal infections in men.
The present study was supported by grants to G.A. from ISS 2006 # 30G.43 and ANLAIDS 2006 # 455.
Role of each of the authors in the study: Alessandra Pierangeli was responsible for design of the study, execution of the HPV-DNA experiments and writing of the manuscript; Carolina Scagnolari for execution of the TaqMan experiments, analysis of the data and revising of the manuscript; Anna Marta Degener for design of the HPV-DNA test and analysis of the sequencing data; Mauro Bucci for execution of the HPV-DNA tests; Antonio Ciardi for execution of the cytological tests; Elisabetta Riva for design and execution of the HPV plasmids for the TaqMan experiments; Marileda Indinnimeo for performing anal brushings and collecting HIV-negative patient data; Giuseppe Mancini for performing HIV test and analysing of the HIV-positive patient data; Gabriella D'Ettorre for contributing to design of the study, to selection of HIV patients, and to revise the manuscript; Vincenzo Vullo for selection of HIV patients, analysis of clinical data, and revising of the manuscript; and Guido Antonelli for design of the study, writing of the manuscript, and grants owner.
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