BetaHPV E6 and E7 colocalize with NuMa in dividing keratinocytes

Human papillomaviruses (HPVs) of genus betapapillomavirus (betaHPV) are implicated in skin carcinogenesis, but their exact role in keratinocyte transformation is poorly understood. We show an interaction of HPV5 and HPV8 oncoproteins E6 and E7 with the nuclear mitotic apparatus protein 1 (NuMA). Binding of E6 or E7 to NuMA induces little aneuploidy, cell cycle alterations, or aberrant centrosomes. Intracellular localization of NuMA is not altered by E6 and E7 expression in 2D cultures. However, the localization profile is predominantly cytoplasmic in 3D organotypic skin models. Both viral proteins colocalize with NuMA in interphase cells, while only E7 colocalizes with NuMA in mitotic cells. Intriguingly, a small subset of cells shows E7 at only one spindle pole, whereas NuMA is present at both poles. This dissimilar distribution of E7 at the spindle poles may alter cell differentiation, which may in turn be relevant for betaHPV-induced skin carcinogenesis.Peer reviewe

Molecular Mechanisms of Human Papillomavirus Induced Skin Carcinogenesis

Infection of the cutaneous skin with human papillomaviruses (HPV) of genus betapapillomavirus (beta HPV) is associated with the development of premalignant actinic keratoses and squamous cell carcinoma. Due to the higher viral loads of beta HPVs in actinic keratoses than in cancerous lesions, it is currently discussed that these viruses play a carcinogenic role in cancer initiation. In vitro assays performed to characterize the cell transforming activities of high-risk HPV types of genus alphapapillomavirus have markedly contributed to the present knowledge on their oncogenic functions. However, these assays failed to detect oncogenic functions of beta HPV early proteins. They were not suitable for investigations aiming to study the interactive role of beta HPV positive epidermis with mesenchymal cells and the extracellular matrix. This review focuses on beta HPV gene functions with special focus on oncogenic mechanisms that may be relevant for skin cancer development

Role of human papillomavirus (HPV) in the development of skin cancer

The incidence of nonmelanoma skin cancer, the most common cancer in humans, continues to rise. The development of precancerous actinic keratoses and cutaneous squamous cell carcinoma (cSCC) is associated with infection with human papillomavirus (HPV) of genus beta (betaHPV). Persistent betaHPV infections in immunocompetent individuals are generally very well controlled by the immune system and largely asymptomatic. However, immunosuppression results in high levels of betaHPV in the skin and consequently increased viral oncoprotein activity, which in turn leads to a significantly increased risk for skin cancer. However, even in immunocompetent individuals, the risk of cSCC increases with age as a result of accumulated UV-induced DNA damage in the skin. In these patients, the mechanism of betaHPV-dependent carcinogenesis seems to be different from that observed in immunocompromised patients. The underlying mechanism of oncogenesis in immunocompetent patients is currently less well understood. This review summarizes the current research data, which provide compelling evidence that cutaneous papillomaviruses, particularly in interaction with UV light, promote skin carcinogenesis via a hit-and-run mechanism by enhancing the genotoxic effects of UV light in the initial phases of this multistep process. Furthermore, an overview of novel vaccination strategies against papillomaviruses that are currently tested in clinical trials is provided, which could significantly improve the treatment options for high-risk patients in the future

HPV and cancer of the oral cavity

Increased awareness of human papillomavirus (HPV) as an etiological cause of head and neck squamous cell carcinoma has increased the interest in analysis of distinct oral sub-sites. It is currently under debate, whether HPV plays a role in the development of squamous cell carcinoma of the oral cavity (OSCC). The weakness in most published studies is the lack of performing different HPV detection tests combined with analysis for biological activity of the virus. In addition, different sub-sites of the oral cavity had been combined to a single entity, which retrospectively leads to a highly heterogeneous basis of data. In this review we mainly discuss the unclear role of HPV in OSCC development

Establishment and Characterization of Immortalized Gingival Epithelial and Fibroblastic Cell Lines for the Development of Organotypic Cultures

In vitro studies using 3D co-cultures of gingival cells can resemble their in vivo counterparts much better than 2D models that typically only utilize monolayer cultures with short-living primary cells. However, the use of 3D gingival models is still limited through lack of appropriate cell lines. We aimed to establish immortalized cell line models of primary human gingival epithelium keratinocytes (HGEK) and gingival fibroblasts (GFB). Immortalized cell lines (HGEK-16 and GFB-16) were induced by E6 and E7 oncoproteins of human papillomavirus. In addition, 3D multilayered organotypic cultures were formed by embedding GFB-16 cells within a collagen (Col) matrix and seeding of HGEK-16 cells on the upper surfaces. Cell growth was analyzed in both immortalized cell lines and their parental primary cells. The expression levels of cell type-specific markers, i.e. cytokeratin (CK) 10, CK13, CK16, CK18, CK19 for HGEK-16 and Col I and Col II for GFB-16, were evaluated by quantitative real-time polymerase chain reaction (qRT-PCR). Expansion of the primary cultures was impeded at early passages, while the transformed immortalized cell lines could be expanded for more than 30 passages. In 3D cultures, immortalized HGEK formed a multilayer of epithelial cells. qRT-PCR showed that cell-specific marker expression in the 3D cultures was qualitatively and quantitatively closer to that in human gingival tissue than to monolayer cultures. These results indicate that immortalized gingival fibroblastic and epithelial cell lines can successfully form organotypic multilayered cultures and, therefore, may be useful tools for studying gingival tissue in vitro. (C) 2014 S. Karger AG, Base

Establishment of an oral infection model resembling the periodontal pocket in a perfusion bioreactor system

Periodontal infection involves a complex interplay between oral biofilms, gingival tissues and cells of the immune system in a dynamic microenvironment. A humanized in vitro model that reduces the need for experimental animal models, while recapitulating key biological events in a periodontal pocket, would constitute a technical advancement in the study of periodontal disease. The aim of this study was to use a dynamic perfusion bioreactor in order to develop a gingival epithelial-fibroblast-monocyte organotypic co-culture on collagen sponges. An 11 species subgingival biofilm was used to challenge the generated tissue in the bioreactor for a period of 24 h. The histological and scanning electron microscopy analysis displayed an epithelial-like layer on the surface of the collagen sponge, supported by the underlying ingrowth of gingival fibroblasts, while monocytic cells were also found within the sponge mass. Bacterial quantification of the biofilm showed that in the presence of the organotypic tissue, the growth of selected biofilm species, especially Campylobacter rectus, Actinomyces oris, Streptococcus anginosus, Veillonella dispar, and Porphyromonas gingivalis, was suppressed, indicating a potential antimicrobial effect by the tissue. Multiplex immunoassay analysis of cytokine secretion showed that interleukin (IL)-1 beta, IL-2, IL-4, and tumor necrosis factor (TNF)-alpha levels in cell culture supernatants were significantly up-regulated in presence of the biofilm, indicating a positive inflammatory response of the organotypic tissue to the biofilm challenge. In conclusion, this novel host-biofilm interaction organotypic model might resemble the periodontal pocket and have an important impact on the study of periodontal infections, by minimizing the need for the use of experimental animal models

Simultaneous Induction of Benign Condyloma and High-grade Anal Dysplasia Induced by Low-risk Human Papillomavirus Type 42

Infections with human papillomaviruses (HPV) induce a heterogeneous spectrum of cutaneous and mucomembraneous lesions. Epithelial lesions caused by HPV-types of the genus alpha (e.g. anogenital warts/condylomata acuminata, intraepithelial neoplasias and invasive cancers) predominantly occur in the anogenital region. Depending on their oncogenic potential, alpha-HPVs can be divided into high-risk (e.g. HPV16 or HPV18) and low-risk types (e.g. HPV6 or HPV11). In contrast to high-risk HPVs, low-risk HPVs are only rarely capable of inducing high-grade dysplasias or anogenital cancers (< 1-10%) (1-3). We report here an immunocompetent patient with simultaneous development of benign condylomata acuminata and high-grade anal dysplasia induced by the same low-risk HPV-type, HPV42

Inactivation of Polyomavirus SV40 as Surrogate for Human Papillomaviruses by Chemical Disinfectants

Human papillomaviruses (HPV) are non-enveloped DNA viruses infecting cutaneous and mucosal squamous epithelia. Sexually transmitted HPV-types that are carcinogenic to humans such as HPV16 can induce cervical and other anogenital cancers. Virus transmission through fomites such as inadequately disinfected gynecological equipment is a further potential transmission route. Since HPV cannot be easily grown in cell culture, polyomavirus SV40 has been used as a surrogate virus when testing the virucidal activity of chemical disinfectants. So far, studies that have compared the virucidal activity of different disinfectants against HPV and SV40 are lacking. Here, we evaluated the susceptibility of HPV16 pseudovirus and SV40 to seven active biocidal substances using quantitative suspension tests. Ethanol, glutaraldehyde (GTA), dodecyldipropylentriamin (DPTA), and ortho-phthalaldehydes (OPA) were able to reduce the infectivity of HPV16 pseudovirus > 99.99% after 5 min. In contrast, isopropanol, peracetic acid (PAA), and quaternary ammonium compounds with alkylamines (QAC) only led to a slight or no reduction in infectivity. Concerning SV40, only GTA (60 min contact time), PAA, and OPA had virus-inactivating effects. In conclusion, the virucidal activity of three out of seven disinfectants tested was different for HPV16 pseudovirus and SV40. In this study, SV40 was shown to be a reliable surrogate virus for HPV when testing isopropanol-, GTA-, QAC-, and OPA-based disinfectants

Acquired lymphangioma circumscriptum in high-grade penile intraepithelial neoplasia

Lymphangioma circumscriptum (LC) is a rare, benign vascular malformation induced by abnormal lymphatic vessels of the skin. LC might be either congenital or acquired, and is predominantly located on the trunk, buttock, axillary region, or thighs. Penile LC is rare. This case report describes a patient with acquired LC associated with high-grade penile intraepithelial neoplasia induced by human papillomavirus type 66. As the patient had multifocal lesions on the glans penis and prepuce we decided to perform circumcision, followed by electrocoagulation of the lesions on the glans. Electrocautery should be considered as a first choice for treatment of LC located at surgically challenging regions such as the glans penis