Se llevó a cabo un estudio inmunohistoquímico incluyendo un panel de 18 biomarcadores y un estudio molecular en 81 biopsias de cáncer de cuello uterino (59 carcinomas epidermoides y 19 adenocarcinomas cervicales) diagnosticados en el Servicio de Anatomía Patológica del Hospital Clínico Universitario de Valencia, con la finalidad de realizar la caracterización inmunohistoquímica de esta neoplasia invasora y determinar la presencia de ADN-HPV asociado. El estudio inmunohistoquímico se realizó, en tumores cervicales y tejido cervical benigno como control, mediante la técnica de avidina-biotina, interpretado mediante una escala semicuantitativa. Se determinó la presencia y genotipificación de ADN-HPV, en las muestras neoplásicas, utilizando el método PCR-SPF10/LiPA.El tipo histológico más frecuente fue el carcinoma epidermoide moderadamente diferenciado (43.5%) y el adenocarcinoma endocervical (42.1%). La edad promedio de las pacientes fue 50.67±13.62 años. Los hallazgos inmunohistoquímicos fueron los siguientes: los carcinomas cervicales mostraron un índice proliferativo (Ki-67/MIB-1) entre moderado y alto; presentaron inmunoreactividad positiva para las proteínas celulares reguladoras ciclina D1, pRb, p16, p21, p27 y ciclina E igual a 32.1%, 30%, 79.5%, 52.5%, 53.2% y 7.7% respectivamente. Respecto a las proteínas apoptóticas, la reactividad ante el anticuerpo contra p53 fue de 3.9%, MDM-2 (100%), Bax (67.5%) y Bcl-2 (3.7%). Las moléculas de adhesión celular cadherina-E, CD44s y CD44v3 estaban sobrexpresadas con localización preferentemente citoplasmática en el 76.6%, 90.9% y 97.5%. La inmunorespuesta para la ubicuitina fue de 98.7%, citoqueratinas (AE1/AE3) 97,5%, CEA 89.7% y telomerasa 52.6%. Se encontraron diferencias estadísticamente significativas en la inmunoreactividad de los tumores en comparación con los controles a excepción de la reactividad de p53, Bcl-2 y CEA; también en cuanto a la reactividad de p27, MDM-2, citoqueratinas, localización celular de la cadherina-E y CD44v3 en los carcinomas de diferentes grados de maduración y en el caso de los subtipos histológicos de adenocarcinoma cervical para Bax, telomerasa y CD44v3. Hallamos correlación en la inmunoexpresión de 1) Ki-67 con p16 (p=0.003), p21 (p=0.023), p27 p=0.032), telomerasa (p=0.012) y ubicuitina (p=0.007). 2) pRb y ciclina D1 (p=0.027), CEA (p=0.050) y CD44s (p=0.009); p53 con ciclina D1 (p=0.011) y CEA (p=0.015). 3) p16/p21 (p=0.046), p16/p27 (p=0.027), p21/p27 (p=0.003), p27/Bax (p=0.015), p27/Bcl-2 (p=0.039) y p16/MDM-2 (p=0.013). 4) ciclina D1 con p21 (p=0.000), CEA (p= 0.000) y Cadherina-E (p=0.001). 5) Cadherina-E cn CD44s (p=0.000), CD44v3 (p=0.013), con MDM-2 (p=0.000) y ubicuitina (p=0.026); CD44s/CD44v3 (p=0.000); CD44v3 y p16 (p=0.007), ubicuitina (p=0.012), telomerasa (p=0.025) y citoqueratinas (p=0.009). 6) Citoqueratinas con p27 (p=0.022), Bax (p=0.007) y Bcl-2 (p=0.009). 7) Telomerasa con p16 (p=0.004), p27 (p=0.037), MDM-2 (p=0.002), CD44v3 (p=0.025) y Ki.67 (p=0.012). En cuanto a la presencia de ADN-HPV en las muestras neoplásicas, el 96.3% (78/81) fueron positivas; de estos, el 59% (46/78) presentó infección única y el 41% (32/78) infección múltiple. El tipo viral específico más frecuente fue el HPV16 (55.13%) seguido de los HPV18, HPV58 y HPV X, en igual porcentaje (1.28%), así mismo el HPV16 fue el tipo viral más frecuente tanto en carcinomas epidermoides (55.9%) como en adenocarcinomas cervicales (52.6%). No encontramos diferencias estadísticamente significativas en la inmunoexpresión del panel de biomarcadores estudiados entre la infección única y múltiple. El cáncer de cuello uterino asociado a infección por HPV oncogénico se caracteriza porque muestra inmunoreactividad alterada en las principales vías reguladoras del ciclo celular, modulación de las proteínas apoptóticas, inmunolocalización celular anormal de las moléculas de adhesión celular, así como sobrexpresión de CEA y citoqueratinas, intensa degradación de proteínas y activación de la telomerasa. Los tipos histológicos de cáncer de cuello uterino comparten un proceso de carcinogénesis multifactorial en el que está involucrada, comúnmente, la presencia del HPV oncogénico como agente causal necesario para el desarrollo de esta neoplasia. / Molecular and inmunohistochemistry (IHC) studies including eighteen biomarkers were performed in eighty-one cervical cancer biopsies with the aim of performing the IHC characterization and to detect the presence of human papillomavirus (HPV).IHC studies were performed using benign cervical tissue as control of expression and results were evaluated using a semi quantitative scale. AND-HPV presence was detect and genotyped in each neoplasic sample by using SPF10/LiPA method.More frequent cancer histologic subtypes were moderately differentiated epidermoid carcinoma (43,5%) and endocervical adenocarcinoma (42,1%). IHC findings obtained for the tested antibodies are described next. Proliferative index inferred from Ki-67/MIB-1 was between moderate and high. Regulating proteins rendered positive results in different percentages: cyclin D1 (32,1%), cyclin E (7,7%), pRB (30%), p16 (79,5%), p21 (52,5%), p27 (53,2%) and apoptotic proteins: p53 (3,9%), MDM-2 (100%), Bax (67,5%) and Bcl-2 (3,7%). Cellular adhesion molecules: cadherin-E, CD44s and CD44v3 were overexpressed with cytoplasmic localization mainly in 76,6%; 90,9% and 97,5%, respectively. Other tumor related molecules were ubiquitin, cytokeratins (AE1/AE3), CEA and telomerase with 98.7%, 97,5%, 89,7% and 52,6%, respectively.Statistical analysis showed significant differences between tumours and controls in all antibodies tested with the exception of p53, BCL2 and CEA. When combining the antibodies we established strech association in the inmunoexpression of them. Presence of AND-HPV was detected in 96,3% of samples showing single infection the 59% and multiple infection the rest of tumours. The most frequent virus type was HPV16 (55,13%) followed by HPV18, HPV58 and HPV X with the same associated frequency (1,28%). HPV16 was the most frequent virus type in both epidermoid and cervical tumours. It was not possible to establish differences between single and multiple infections related to IHC expression profile.Cervical cancer is characterized by the altered expression of IHC patterns in cellular regulatory pathways affecting apoptotic proteins, cellular cycle, adhesion molecules inmunolocalization, CEA and cytoqueratines overexpression and increase of protein degradation and telomerase activity. Histological subtiypes of cervical cancer share a multifactorial complex process including HPV infection as common tumour agent.

The HPV vaccine (Gardasil) is a significant advancement in women’s health that garnered both positive and negative media coverage. Since public opinion and population uptake of the vaccine can be influenced by media coverage, the purpose of this study was to assess the discussion and presentation of risk information about HPV, cervical cancer and the HPV vaccine in Canadian and U.S. newspapers and news magazines. Using directed content analysis, the reporting of fear-inducing messages about HPV, cervical cancer and the HPV vaccine were compared between Canadian and U.S. top-circulating national newspapers between January 2006—December 2007. Significant differences between countries were found in the number of fear messages about cervical cancer (p < 0.05) but not for HPV or the HPV vaccine. Readability was higher than recommended for the public and emotional tone of the articles was progressively negative. Directed content analysis was also used to assess the discussion of risks, fear-inducing messages about HPV, cervical cancer and the HPV vaccine in four high circulating North American news magazines from January 2006—December 2007. Risk messages about HPV and cervical cancer focused on threatening illness or injury and reporting on the HPV vaccine emphasized it being poorly understood by science. Newspaper and news magazine articles on HPV, cervical cancer and the HPV vaccine included fear-inducing messages. Public health officials and health educators need to be aware of media reporting of fear based risk messages in order to alleviate public anxiety and concern about the HPV vaccine.

The HPV vaccine (Gardasil) is a significant advancement in women’s health that garnered both positive and negative media coverage. Since public opinion and population uptake of the vaccine can be influenced by media coverage, the purpose of this study was to assess the discussion and presentation of risk information about HPV, cervical cancer and the HPV vaccine in Canadian and U.S. newspapers and news magazines. Using directed content analysis, the reporting of fear-inducing messages about HPV, cervical cancer and the HPV vaccine were compared between Canadian and U.S. top-circulating national newspapers between January 2006—December 2007. Significant differences between countries were found in the number of fear messages about cervical cancer (p < 0.05) but not for HPV or the HPV vaccine. Readability was higher than recommended for the public and emotional tone of the articles was progressively negative. Directed content analysis was also used to assess the discussion of risks, fear-inducing messages about HPV, cervical cancer and the HPV vaccine in four high circulating North American news magazines from January 2006—December 2007. Risk messages about HPV and cervical cancer focused on threatening illness or injury and reporting on the HPV vaccine emphasized it being poorly understood by science. Newspaper and news magazine articles on HPV, cervical cancer and the HPV vaccine included fear-inducing messages. Public health officials and health educators need to be aware of media reporting of fear based risk messages in order to alleviate public anxiety and concern about the HPV vaccine.

Le cancer du col utérin est un des cancers les plus prévalents dans le monde et représente une cause majeure de mortalité par cancer chez la femme. Le dépistage de ce cancer est actuellement basé sur l’examen clinique, la colposcopie, la cytologie du col utérin et l’histopathologie dont la reproductibilité inter observateur est médiocre surtout en ce qui concerne les lésions de bas grade (CIN 1). Bien qu’amélioré par de nouveaux marqueurs (p16INK4A), la recherche de nouveaux marqueurs spécifiques des lésions du col utérin nécessite l’élucidation des mécanismes de la cancérogenèse parmi lesquels les altérations épigénétiques, but de notre recherche. Nous avons évalué au cours des différentes étapes de la cancérogenèse du col utérin la méthylation globale de l'ADN par immunohistochimie et HPLC. L’hyperméthylation des îlots de CpG de trois gènes candidats impliqués dans la signalisation cellulaire (CDH13, DAPK1, TWIST1) a été étudié par une technique de PCR – méthylation spécifique (MSP) après modification de l’ADN au bisulfite de sodium. L’étude immunohistochimique à l'aide d’anticorps dirigés contre la 5-MeCyd objective une hypométhylation globale de l’ADN tardive n’apparaissant que dans les cancers invasifs. Ces résultats ont été confirmés par HPLC. Par contre, l’hyperméthylation des îlots de CpG des gènes candidats étudiés est présente dés le stade lésions précancéreuses. Ces résultats suggèrent qu’au cours de la cancérogenèse du col utérin, l’hypométhylation globale et l’hyperméthylation des îlots de CpG sont deux mécanismes indépendants. L’étude du profil de méthylation des régions promotrices des gènes CDH13, DAPK1, TWIST1 pourrait constituer un marqueur potentiel des lésions précancéreuses et cancéreuses du col de l’utérus / Background: The uterine cervix cancer is one of the main cancer among women worldwide and represents an important cause of death in women especially in developing countries. The screening of this cancer is based on clinic tests, colposcopy, cytology and histopathology which have a low inter observer reproducibility, specially concerning CIN1 lesions. Although improved by new markers (p16INK4A), the discovery of new specific markers requires the elucidation of mechanisms of carcinogenesis such as epigenetic alterations, aim of our study. Material and methods: We evaluated during uterine cervix carcinogenesis global DNA methylation by immunohistochemistry and HPLC. CpGs islands hypermethylation of three genes implied in cellular signalizing (CDH13, DAPK1, TWIST1) were evaluated using specific methylation PCR (MSP) after bisulfite DNA modification. Results: Immunohistochemistry study using anti 5-MeCyd antibodies objectives a late global DNA hypomethylation appearing only in invasive uterine cervix carcinoma. These results were confirmed by HPLC. However, CpGs islands hypermethylation of the promoter of CDH13, DAPK1 and TWIST1 genes studied were detected in precancerous lesions. Conclusions: Our results suggest that during uterine cervix carcinogenesis, global DNA hypomethylation and gene hypermethylation were two independent mechanisms. The methylation profile of the promoter regions of CDH13, DAPK1 and TWIST1 genes would represent a potential marker for the diagnosis of the precancerous and cancerous uterine cervix lesions

Human papillomavirus (HPV) is the most common sexually transmitted infection in the United States, but few studies have examined the progression from HPV infection to disease in men. Genital condyloma are the most common clinical manifestation of HPV infection. Though not associated with mortality, condyloma are a source of emotional distress, and treatment is often painful with a high recurrence rate. The aims of this study were to examine the distribution of HPV types present on the surface of condyloma, estimate the incidence of condyloma overall and after type-specific HPV infections, assess the sociodemographic and sexual behavior factors independently associated with incident condyloma, and examine the concordance between HPV types detected on the surface and in the tissue of condyloma. Participants included 2,487 men from the United States, Brazil, and Mexico who were enrolled in the prospective HPV in Men (HIM) Study and followed every six months for up to four years. At each study visit men completed a computer-assisted-self-administered risk factor questionnaire and samples of healthy penile skin were obtained to test for HPV DNA. A trained clinician examined men for the presence of condyloma and swabbed the surface of lesions to test for HPV DNA. Men were followed for a median of 17.9 months and 112 incident condyloma were identified. Thirty-four external genital lesions were also biopsied to test for HPV within the lesion tissue. PCR was used to test for HPV DNA and Linear Array was used to genotype 13 oncogenic and 24 non-oncogenic HPV types in samples obtained from swabbing the lesion surface. The LiPa assay was used to genotype 20 HPV types in biopsy samples. The Kaplan-Meier method was used to estimate incidence and Cox proportional hazards models were used to examine factors independently associated with incident condyloma. Using biopsy samples as the gold standard, sensitivity and specificity were calculated to examine concordance between HPV types detected on the surface and within the tissue of condyloma. Condyloma incidence was 2.35 per 1,000 person-years. HPV 6 (43.8%), 11 (10.7%), and 16 (9.8%) were the most common types detected on condyloma. The probability of developing condyloma within 24-months of an incident HPV 6/11 infection was 14.6% (95% confidence interval (CI): 7.5-21.1). The median time to condyloma development was 17.1 months (95% CI: 12.4-19.3), with the shortest time to detection observed among men with incident HPV infections with types 6/11 only (6.2 months; 95% CI: 5.6-24.2). Factors associated with condyloma were incident HPV 6/11 infection (hazard ratio (HR) = 12.42; 95% CI: 3.78-40.77), younger age (HR = 0.43; 95% CI: 0.26-0.77; 45-70 vs. 18-30 years), high lifetime number of female partners (HR = 5.69; 95% CI: 1.80-17.97); 21 or greater vs. 0), and sexual behaviors in the previous three months including infrequent condom use (HR = 2.44; 95% CI: 1.16-5.14;

This dissertation research explores how values regarding sexuality, morality, responsibility, protection, trust, and risk — expressed through parent, daughter, and healthcare provider relationships and interactions — inform parental decisions regarding the Gardasil® vaccine. In particular, the research examines the competing and conflicting meanings that parents and providers ascribe to vaccination and how actors position the vaccine within a wider set of negotiated, value–laden discourses. Because these narratives are situated within a larger structural field that shapes the landscape in which providers and parents interact, relevant historical and structural factors, including vaccine policy, cost, and compensation are discussed.

High-risk human papillomaviruses (HPVs) are associated with nearly all cases of cervical cancer and also contribute to other types of anogenital and oropharyngeal cancers. The high-risk HPV E6 oncoprotein contributes to malignant progression in part by the targeted degradation of the tumor suppressor p53. The activation of growth factor and nutrient sensing pathways including receptor protein tyrosine kinases (RPTKs) and mTORC1 may also support cellular transformation. Moreover, previous studies suggested that HPV16 E6 activates mTORC1. We are particularly interested in understanding the mechanisms by which HPV E6 activates mTORC1 and the function of mTORC1 activation in HPV infection. Here we show that high-risk HPV16 E6 activates mTORC1 signaling and increases cap dependent translation through an increase in S6K signaling and an increase in 4E-BP1 phosphorylation. Mechanistically we found that HPV16 E6 activates AKT under conditions of nutrient deprivation. The combined approach of phospho-tyrosine immunoprecipitations and Western blot identified HPV16 E6 mediated activation of a subset of receptor protein tyrosine kinases. HPV16 E6 activates RPTKs at least in part by increasing the internalization of phosphorylated and activated receptor species. The signaling adaptor protein Grb2 associates with HPV16 E6, and Grb2 knockdown abrogated HPV16 E6 mediated mTORC1 activation. We hypothesize that Grb2 may be important in relaying E6 mediated RPTK activation to downstream signaling cascades. In this dissertation we also evaluate mTORC1 signaling and cap dependent translation in cells expressing HPV16 E6 mutants and E6 proteins from other HPV types. Binding to p53 and the association with proteins that contain an LXXLL motif are important for HPV16 E6 mediated mTORC1 activation. An increase in mTORC1 activation and cap dependent translation is shared between high-and low-risk mucosal, but not cutaneous HPV E6 proteins. Association with proteins through their LXXLL binding motif is also important for low-risk mucosal HPV E6 activation of mTORC1 and cap dependent translation. Shared mucosal E6 activation of mTORC1 indicates that mTORC1 may be important for the viral lifecycle in mucosal epithelia. However, it does not rule out the possibility that together with other properties of high-risk HPV E6 proteins, mTORC1 activation may promote transformation.

The aim of this research was to study the correlation between tissue tumor marker expression and HR-HPV infection, smoking, hormonal contraceptive use and sex steroids in women with cervical intraepithelial neoplasia or normal epithelium. The study investigated the expression of 11 tumor markers in cervical biopsies obtained from 228 women with different diagnoses ranging from normal cervical epithelium to various stages of CIN. 188 women were recruited at our colposcopy clinic (out-patient surgery, Department of Obstetrics and Gynecology, Falun Hospital) for laser cervical conization or a directed punch biopsy, either because of a vaginal smear (Pap smear) that showed cytological findings suggesting CIN, or because of repeated findings showing atypical squamous cells of undetermined significance (ASCUS). For 40 volunteers, punch biopsies were taken from the normal cervical epithelium. The time period for this study was 2005-2007. Study I :  228 women, of whom 116 were tested, 64 were positive to HR-HPV. The results showed that Ki67 tumor cell proliferation index was the only marker that independently correlated to both the presence of HR-HPV and the severity of cervical lesions. Study II:  228 women, of whom 83 were smokers (36, 9%). Smokers showed lower expression of p53, FHIT (tumor suppressor markers) and interleukin-10 .Higher expression of Cox-2 and Ki-67 (tumor proliferation markers). Study III:  195 women who were premenopausal. There was increased p53 expression (tumor suppressor) in the progestin-IUD users compared to non-users. Decreased IL-10 expression (immunological marker) was observed in both COC users and any progestin-only users. Study IV: Serum from 80 premenopausal women was available. The main finding was that the increased levels of serum progesterone and estradiol were associated with increased Cox-2 expression (proliferation marker). Serum progesterone and estradiol levels influence cellular and extracellular proteins which have been associated with neoplastic development in normal epithelium and CIN. Conclusion: The results of these studies support previous epidemiological findings on the role of smoking, contraceptive use and sex steroids as co-factors in development of CIN and that tumor marker expression varies in different grades of CIN.

The most common sexually transmitted infection for both males and females is the human papillomavirus (HPV). HPV is responsible for nearly all cervical cancers. Currently, an HPV vaccine is available; however, HPV vaccination rates for US adolescents are dismal. School nurses serve as the person connecting medical and school communities, and are a critical component in assisting families traverse the medical and educational systems. Thus, there is reason to assume school nurses can be key opinion leaders regarding the HPV vaccine. The purpose of this study was to: (1) explain how the Diffusion of Innovations (DOI) theory explains school nurses' roles as opinion leaders regarding the HPV vaccine; (2) document current literature regarding healthcare providers' perspectives and practice regarding the HPV vaccine; and (3) evaluate school nurses' knowledge, attitudes, perceptions of being an opinion leader and professional practice regarding the HPV vaccine for youth. DOI states opinion leaders influence the rate of an innovation (e.g., the HPV vaccine). We argue school nurses are opinion leaders for the HPV vaccine because of their unique leadership position through their cross-disciplinary understanding of the educational and health systems. The systematic literature review included 28 studies of healthcare providers. The main barrier, vaccine cost, was identified in 12 reports. Additionally, females and older adolescents were more frequently vaccinated than males and younger adolescents. To examine school nurses' knowledge, attitudes, perceptions and professional practice regarding the HPV vaccine, the study included a sample of 413 school nurses. Structural equation modeling revealed knowledge influenced attitudes, attitudes affected perceptions and professional practices, and perceptions predicted professional practice. Furthermore, the perceptions variable was found to be a partial mediator in the model. Practitioners designing programs to engage school nurses in disseminating HPV vaccine education may benefit from questioning whether their programs might be emphasizing non-crucial elements for influencing vaccine dissemination practice (e.g., knowledge) and de-emphasizing influential elements such as attitudes and perceptions.

The overall aim of this thesis is to provide a better understanding of knowledge, attitudes, consent, and decision-making regarding Human papillomavirus (HPV) vaccination, seen from the perspectives of concerned parties – high school students, school nurses, and parents. Two quantitative studies were performed: one descriptive cross-sectional study and one quasi-experimental intervention study. Qualitative studies using focus group interviews and individual interviews were also performed. High school students’ knowledge about HPV and HPV prevention was low but their attitudes toward HPV vaccination were positive. An educational intervention significantly increased the students’ knowledge regarding HPV and HPV prevention. Their already positive attitudes toward condom use and HPV vaccination remained unchanged. The students wanted to receive more information about HPV from school nurses. The school nurses were also positive to HPV vaccination but identified many challenges concerning e.g. priorities, obtaining informed consent, culture, and gender. They saw an ethical dilemma in conflicting values such as the child’s right to self-determination, the parents’ right to make autonomous choices on behalf of their children, and the nurse’s obligation to promote health. They were also unsure of how, what, and to whom information about HPV should be given. Parents, who had consented to vaccination of their young daughters, reasoned as follows: A vaccine recommended by the authorities is likely to be safe and effective, and the parents were willing to do what they could to decrease the risk of a serious disease for their daughter. Fear of unknown adverse events was overweighed by the benefits of vaccination. Parents also saw the school nurse as an important source of HPV information. Conclusions: Positive attitudes toward HPV vaccination despite limited knowledge about HPV, are overarching themes in this thesis. School nurses have a crucial role to inform about HPV prevention. It is important that the concerned parties are adequately informed about HPV and its preventive methods, so that they can make an informed decision about vaccination. A short school-based intervention can increase knowledge about HPV among students. From a public health perspective, high vaccination coverage is important as it can lead to a reduced number of HPV-related disease cases.