Cacilda Tezelli Junqueira Padovani

Local immunosuppression induced by high viral load of human papillomavirus: characterization of cellular phenotypes producing interleukin‐10 in cervical neoplastic lesions

A specific immune response to human papillomavirus (HPV) in the cervical microenvironment plays a key role in eradicating infection and eliminating mutated cells. However, high‐risk HPVs modulate immune cells to create an immunosuppressive microenvironment, and induce these immune cells to produce interleukin 10 (IL‐10). This production of IL‐10, in conjunction with HPV infection, contributes to the appearance of cervical neoplastic lesions. We sought to characterize the IL‐10‐producing cellular phenotype, and investigate the influence of host and HPV factors upon the induction of an immunosuppressive microenvironment. Immunohistochemical analysis demonstrated an increase in IL‐10 production by keratinocytes, macrophages and Langerhans cells in high‐grade cervical lesions and cervical cancer. This increase was more pronounced in patients older than 30 years, and was also correlated with high viral load, and infection with a single HPV type, particularly high‐risk HPVs. Our results indicate the existence of a highly immunosuppressive microenvironment composed of different IL‐10‐producing cellular phenotypes in cervical cancer samples, and samples classified as high‐grade cervical lesions (cervical intraepithelial neoplasia stages II and III). The immunosuppressive microenvironment that developed for these different cellular phenotypes favours viral persistence and neoplastic progression.

Glucocorticoid-induced tumor necrosis factor receptor expression in patients with cervical human papillomavirus infection

INTRODUCTION The progression of human papillomavirus (HPV) infection in the anogenital tract has been associated with the involvement of cells with regulatory properties. Evidence has shown that glucocorticoid-induced tumor necrosis factor receptor (GITR) is an important surface molecule for the characterization of these cells and proposes that GITR ligand may constitute a rational treatment for many cancer types. We aimed to detect the presence of GITR and CD25 in cervical stroma cells with and without pathological changes or HPV infection to better understand the immune response in the infected tissue microenvironment. METHODS We subjected 49 paraffin-embedded cervical tissue samples to HPV DNA detection and histopathological analysis, and subsequently immunohistochemistry to detect GITR and CD25 in lymphocytes. RESULTS We observed that 76.9% of all samples with high GITR expression were HPV-positive regardless of histopathological findings. High GITR expression (77.8%) was predominant in samples with ≥ 1,000 RLU/PCB. Of the HPV-positive samples negative for intraepithelial lesion and malignancy, 62.5% had high GITR expression. High GITR expression was observed in both carcinoma and high-grade squamous intraepithelial lesion (HSIL) samples (p = 0.16). CD25 was present in great quantities in all samples. CONCLUSIONS The predominance of high GITR expression in samples with high viral load that were classified as HSIL and carcinoma suggests that GITR+ cells can exhibit regulatory properties and may contribute to the progression of HPV-induced cervical neoplasia, emphasizing the importance of GITR as a potential target for immune therapy of cervical cancer and as a disease evolution biomarker.

Good agreements between self and clinician-collected specimens for the detection of human papillomavirus in Brazilian patients

Women infected with human papillomavirus (HPV) are at a higher risk of developing cervical lesions. In the current study, self and clinician-collected vaginal and cervical samples from women were processed to detect HPV DNA using polymerase chain reaction (PCR) with PGMY09/11 primers. HPV genotypes were determined using type-specific PCR. HPV DNA detection showed good concordance between self and clinician-collected samples (84.6%; kappa = 0.72). HPV infection was found in 30% women and genotyping was more concordant among high-risk HPV (HR-HPV) than low-risk HPV (HR-HPV). HPV16 was the most frequently detected among the HR-HPV types. LR-HPV was detected at a higher frequency in self-collected; however, HR-HPV types were more frequently identified in clinician-collected samples than in self-collected samples. HPV infections of multiple types were detected in 20.5% of clinician-collected samples and 15.5% of self-collected samples. In this study, we demonstrated that the HPV DNA detection rate in self-collected samples has good agreement with that of clinician-collected samples. Self-collected sampling, as a primary prevention strategy in countries with few resources, could be effective for identifying cases of HR-HPV, being more acceptable. The use of this method would enhance the coverage of screening programs for cervical cancer.

Presence of highly oncogenic human papillomavirus in the oral mucosa of asymptomatic men

OBJECTIVES The aim of this study was to identify highly oncogenic forms of human papillomavirus in the oral mucosa of asymptomatic men. METHODS In this study, we analyzed samples of exfoliated cells from the oral cavity of 559 asymptomatic men. DNA-human papillomavirus was detected using the consensus primers PGMY09/11; viral genotyping was performed using type-specific PCR and restriction fragment length polymorphism. RESULTS DNA-human papillomavirus was detected in 1.3% of the study participants and of those 42.8% were infected by more than one type of virus. Viral types included HPV6, 11, 89 (low oncogenic risk), and HPV52, 53 (high oncogenic risk). Increased vulnerability to human papillomavirus infection was observed in individuals aged over 26 years, among those who reported oral sex practices, and in those who have had more than 16 sexual partners since first engaging in sexual intercourse. CONCLUSIONS There was a low prevalence of human papillomavirus detection in the oral mucosa of asymptomatic men. Highly oncogenic human papillomavirus types and infection by more than one viral type was observed. Oral sex practices and a large number of sexual partners may increase the risk of acquiring human papillomavirus infection.

Predominant overexpression of CD25/FOXP3, IFN-γ, and suppressive cytokines in high-grade lesion samples infected with human papillomavirus

Introduction: Human papillomavirus (HPV) persistent infection is the leading cause of cervical cancer and its precursor lesions, and the inappropriate immune response is among the factors that contribute to viral persistence. This may be influenced by regulatory T (Treg) cells and the production of immunosuppressive cytokines, such as transforming growth factor beta (TGF-β) and interleukin-10 (IL-10). Objective: We established the profile of the predominant response, Th1 or immunosuppressive response, in the tissue microenvironment, by detecting interferon-gamma (IFN-γ), TGF-β, and IL-10, as well as the co-expression of IL-2 receptor alpha (CD25) and forkhead box P3 (FOXP3). Methods: Seventy-four samples from uterine cervix biopsies that underwent HPV deoxyribonucleic acid (DNA) detection and histopathology analysis were immunostained to detect CD25/FOXP3, IFN-γ and suppressive cytokines in lymphocytes. Results: The microenvironment of high-grade squamous intraepithelial lesion (HSIL) samples with high numbers of viral particles (≥ 10,000 copies/ml) contained high numbers of CD25/FOXP3+, TGF-β+, IL-10+, and IFN-γ+ cells. Conclusion: The co-expression of CD25/FOXP3 and the expression of TGF-β, and IL-10 in HSIL samples suggest the existence of Treg cells in these locations, although IFN-γ expression was observed in several cells in these samples. Our data suggest that this cytokine could be related to immunosuppressed microenvironment maintenance, favoring the persistent HPV infection and the progression to carcinoma.

Molecular epidemiology of the human papillomavirus infection in self-collected samples from young women

The prevalence of human papillomavirus (HPV) infection is the highest in young, sexually active women less than 35 years of age. Direct diagnosis of infection by enabling genotyping methods is important considering that the viral types are divided into high (HR‐HPV) and low (LR‐HPV) oncogenic risk. This study aimed to evaluate the epidemiological and molecular characteristics of HPV infection in self‐collected samples from young women. A cross‐sectional study of 245 sexually active students (18 to 35 years of age) was undertaken with self‐collected samples. Extracted DNA was analyzed by polymerase chain reaction (PCR) with the PGMY 09/11 and PC04/GH20 primers for the detection of HPV DNA and the β‐globin gene, respectively. Viral genotyping was performed by type‐specific PCR (TS‐PCR) and restriction fragment length polymorphism (RFLP). Of the 236 valid samples, 68 (28.9%) were positive for HPV DNA, as genotyped by TS‐PCR and RFLP. The HR‐HPV were most prevalent, especially HPV‐16, ‐31, ‐33, and ‐45, and the most prevalent LR‐HPV were HPV‐6 and ‐83. Multi‐type HPV infections were detected in 17 (25%) samples. HPV infection was statistically more prevalent among younger women with lower educational levels and who had more partners in the past 2 years. A high prevalence of HPV infection was found in the age group examined, especially HR‐HPV types, as well as the presence of risk behaviors associated with HPV infection were observed. Considering these results, vaccinating females before the onset of sexual activity in Brazil should be emphasized. J. Med. Virol. 86:266–271, 2014.

Type-specific Human papillomavirus infection among heterosexual males examined by peniscopy

Men are the primary link in the human papillomavirus (HPV) epidemic chain. They act as both bearers and transmitters of HPV, contributing substantially to the increase in HPV incidence and the increased risk of cervical cancer. Less frequently, HPV causes the development of penis or anal cancer.1 The results of previous HPV studies in men have shown inconsistent prevalence ranging from 0% to –73%.2 The aim of this study was to determine whether there were any associations between peniscopy (a suggested test for HPV detection), the frequency of HPV DNA detection, and the primary viral types present in men who sought …

Detection of regulatory T cell phenotypic markers and cytokines in patients with human papillomavirus infection: BONIN et al.

Infection with human papillomavirus (HPV) is the main cause of cervical cancer. Viral persistence is considered the main risk factor for neoplastic progression and evidence suggests that regulatory T cells (Treg) play an important role in the failure of viral elimination. The aim of this study was to detect phenotypic markers of Treg and cytokines interleukin (IL)‐10 and transforming growth factor (TGF)‐β, in the cervical microenvironment of HPV‐infected patients. One hundred and one samples of uterine cervix embedded in paraffin were analyzed. We used immunohistochemistry to examine the coexpression of the CD25/FOXP3 and CD4/TGF‐β markers, and the expression of GITR and IL‐10 in cells present in the cervical stroma. We detected a microenvironment composed of high proportions of CD25+FOXP3+, CD4+TGFβ+, IL‐10+, and GITR+ cells in samples with high viral loads and severe lesions of HPV‐infected patients. The abundance of these markers, indicative of the presence of Treg cells and immunosuppressive cytokines, was significantly associated with severe lesions and elevated viral loads in the examined samples. These results suggest that Treg cells may be involved in maintaining a microenvironment favorable for viral persistence and neoplastic progression. Our findings support those of previous studies that suggested that these markers could be used to predict HPV persistence and neoplastic progression, and as potential targets for immune response modulation.

Human papillomavirus and coinfections with Chlamydia trachomatis, Gardnerella vaginalis, and Trichomonas vaginalis in self-collected samples from female sex workers in the Central-Western region of Brazil

Introduction: Human papillomavirus (HPV) is intimately associated with cervical cancer, and the presence of coinfections, such as with Chlamydia trachomatis, Gardnerella vaginalis and Trichomonas vaginalis, may potentiate or facilitate HPV infection. Female sex workers are considered vulnerable to the acquisition of these infections due to exposure to risk factors. Objective: To determine HPV infection, viral types and coinfections in self-collected samples from female sex workers. Methods: Self-collected samples from female sex workers, of vaginal canal and uterine cervix, were subjected to HPV-deoxyribonucleic acid (DNA) detection, viral genotyping by type-specific polymerase chain reaction (PCR), restriction fragment length polymorphism (RFLP), and the detection of coinfection. Results: HPV-DNA was detected in 19.4% of the samples, and HPV 31, 6, and 53 were the most frequently detected types. There was a predominance of highrisk oncogenic HPV (HR-HPV) and a strong presence of simultaneous infections with multiple HPV types (84.6%). Coinfections with both HPV and C. trachomatis, and HPV and G. vaginalis were detected. The variables that were statistically associated with HPV infection and the presence of multiple infections were non-use of condoms and non-compliance with regular cervical cytology screening. Conclusion: The results highlight the importance of more comprehensive studies among vulnerable populations, aiming to establish measures to raise awareness about the risks of contracting sexually transmitted infections, as well as to support future studies for introducing HPV vaccines with wider coverage of viral types.

S100, CD68, and MHC class II molecule expression in cervical high- and low-grade HPV-induced lesions

INTRODUCTION Some human papillomavirus (HPV) types are involved in malignant processes in the cervical epithelium, with 99% of cases attributed to oncogenic HPV infection. This study aimed to detect S100, CD68, and major histocompatibility complex class II (MHC-II) molecules in cervical uterine epithelial samples in patients with high- and low-grade lesions induced by HPV. METHODS Fifty-eight samples from patients who were confirmed positive or negative for high-risk oncogenic HPV DNA, had histopathological diagnosis of cervical intraepithelial neoplasia (CIN) of grades I, II, or III, or were negative for intraepithelial lesion or malignancy were subjected to immunohistochemistry reaction to S100 protein, CD68, and MHC-II (HLA-DR alpha chain). RESULTS The presence of MHC-II predominated in samples exhibiting histopathological alterations (p < 0.05). S100 detection was more numerous in carcinoma samples (CIN III) (75%). Presence of this protein correlated significantly (p < 0.05) with histopathological findings and viral load. CONCLUSIONS A small expression of CD68 was observed, which may be explained by the observation in our study having been made on random microscopic fields and not on specific areas. The findings, such as the presence of S100 protein and MHC-II expression in samples with histological alterations, could suggest that the immune system fails to control HPV replication at the early stages of infection. Further studies with larger prospective data are necessary to confirm this result.