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Skin mononuclear phagocytes (MNPs) provide the first interactions of invading viruses with the immune system. In addition to Langerhans cells (LCs), we recently described a second epidermal MNP population, Epi-cDC2s, in human anogenital epidermis that is closely related to dermal conventional dendritic cells type 2 (cDC2) and can be preferentially infected by HIV. Here we show that in epidermal explants topically infected with herpes simplex virus (HSV-1), both LCs and Epi-cDC2s interact with HSV-1 particles and infected keratinocytes. Isolated Epi-cDC2s support higher levels of infection than LCs in vitro, inhibited by acyclovir, but both MNP subtypes express similar levels of the HSV entry receptors nectin-1 and HVEM, and show similar levels of initial uptake. Using inhibitors of endosomal acidification, actin and cholesterol, we found that HSV-1 utilises different entry pathways in each cell type. HSV-1 predominantly infects LCs, and monocyte-derived MNPs, via a pH-dependent pathway. In contrast, Epi-cDC2s are mainly infected via a pH-independent pathway which may contribute to the enhanced infection of Epi-cDC2s. Both cells underwent apoptosis suggesting that Epi-cDC2s may follow the same dermal migration and uptake by dermal MNPs that we have previously shown for LCs. Thus, we hypothesize that the uptake of HSV and infection of Epi-cDC2s will stimulate immune responses via a different pathway to LCs, which in future may help guide HSV vaccine development and adjuvant targeting., Author summary Here we describe the first interactions of herpes simplex virus type 1 (HSV-1) with the human immune system as it enters the human genital skin. It was previously thought that this initial interaction of HSV-1 was only with ‘Langerhans cells’ (LCs) but we describe another important interaction with a new immune cell, Epi-cDC2s, which can also be infected by HIV and much more efficiently than LCs. Thus, there are now two of these types of immune cells resident in human epidermis. Therefore the way in which sexually transmitted and skin infecting viruses establish infection needs to be re-examined, including for HSV. We compared how these two cell types interacted with HSV-1 and showed Epi-cDC2s are more susceptible to HSV-1 infection than LCs and take up the virus via a different entry pathway. We speculate these cells may then carry HSV fragments to the dermis and then to lymph nodes to stimulate a protective immune response. These findings suggest similar routes may apply for chickenpox and cowpox viruses which also enter via the skin. Studying this pathway to establishment of natural immunity to HSV, may help guide the development of a successful vaccine.

Herpes simplex viruses (HSV) types 1 and 2 are ubiquitous. They both cause genital herpes, occasionally severe disease in the immunocompromised, and facilitate much HIV acquisition globally. Despite more than 60 years of research, there is no licensed prophylactic HSV vaccine and some doubt as to whether this can be achieved. Nevertheless, a previous HSV vaccine candidate did have partial success in preventing genital herpes and HSV acquisition and another immunotherapeutic candidate reduced viral shedding and recurrent lesions, inspiring further research. However, the entry pathway of HSV into the anogenital mucosa and the subsequent cascade of immune responses need further elucidation so that these responses could be mimicked or improved by a vaccine, to prevent viral entry and colonization of the neuronal ganglia. For an effective novel vaccine against genital herpes the choice of antigen and adjuvant may be critical. The incorporation of adjuvants of the vaccine candidates in the past, may account for their partial efficacy. It is likely that they can be improved by understanding the mechanisms of immune responses elicited by different adjuvants and comparing these to natural immune responses. Here we review the history of vaccines for HSV, those in development and compare them to successful vaccines for chicken pox or herpes zoster. We also review what is known of the natural immune control of herpes lesions, via interacting innate immunity and CD4 and CD8 T cells and the lessons they provide for development of new, more effective vaccines.

Herpes Simplex Virus (HSV) is a highly prevalent sexually transmitted infection that aside from causing cold sores and genital lesions, causes complications in the immunocompromised and has facilitated a large proportion of HIV acquisition globally. Despite decades of research, there is no prophylactic HSV vaccine ready for use in humans, leaving many questioning whether a prophylactic vaccine is an achievable goal. A previous HSV vaccine trial did have partial success in decreasing acquisition of HSV2 - promising evidence that vaccines can prevent acquisition. However, there is still an incomplete understanding of the immune response pathways elicited by HSV after initial mucosal infection and how best to replicate these responses with a vaccine, such that acquisition and colonisation of the dorsal root ganglia could be prevented. Another factor to consider in the rational design of an HSV vaccine is adjuvant choice. Understanding the immune responses elicited by different adjuvants and whether lasting humoral and cell-mediated responses are induced is important, especially when studies of past trial vaccines found that a sufficiently protective cell-mediated response was lacking. In this review, we discuss what is known of the immune control involved in initial herpes lesions and reactivation, including the importance of CD4 and CD8 T cells, and the interplay between innate and adaptive immunity in response to primary infection, specifically focusing on the viral relay involved. Additionally, a summary of previous and current vaccine trials, including the components used, immune responses elicited and the feasibility of prophylactic vaccines looking forward, will also be discussed.

ObjectiveVaccination against hepatitis B virus (HBV) confers protection from subsequent infection through immunological memory that is traditionally considered the domain of the adaptive immune system. This view has been challenged following the identification of antigen-specific memory natural killer cells (mNKs) in mice and non-human primates. While the presence of mNKs has been suggested in humans based on the expansion of NK cells following pathogen exposure, evidence regarding antigen-specificity is lacking. Here, we demonstrate the existence of HBV-specific mNKs in humans after vaccination and in chronic HBV infection.DesignNK cell responses were evaluated by flow cytometry and ELISA following challenge with HBV antigens in HBV vaccinated, non-vaccinated and chronic HBV-infected individuals.ResultsNK cells from vaccinated subjects demonstrated higher cytotoxic and proliferative responses against autologous hepatitis B surface antigen (HBsAg)-pulsed monocyte-derived dendritic cells (moDCs) compared with unvaccinated subjects. Moreover, NK cell lysis of HBsAg-pulsed moDCs was significantly higher than that of hepatitis B core antigen (HBcAg)-pulsed moDCs (non-vaccine antigen) or tumour necrosis factor α-activated moDCs in a NKG2D-dependent manner. The mNKs response was mediated by CD56dim NK cells coexpressing CD57, CD69 and KLRG1. Further, mNKs from chronic hepatitis B patients exhibited greater degranulation against HBcAg-pulsed moDCs compared with unvaccinated or vaccinated patients. Notably, mNK activity was negatively correlated with HBV DNA levels.ConclusionsOur data support the presence of a mature mNKs following HBV antigen exposure either through vaccination or infection. Harnessing these antigen specific, functionally active mNKs provides an opportunity to develop novel treatments targeting HBV in chronic infection.

Herpes simplex keratitis is commonly caused by Herpes simplex virus type 1, which primarily infects eyelids, corneas, or conjunctiva. Herpes simplex virus type 1-through sophisticated interactions with dendritic cells (DCs), a type of antigen-presenting cell)-initiates proinflammatory responses in the cornea. Corneas were once thought to be an immune-privileged region; however, with the recent discovery of DCs that reside in the cornea, this long-held conjecture has been overturned. Therefore, evaluating the clinical, preclinical, and cell-based studies that investigate the roles of DCs in corneas infected with Herpes simplex virus is critical. With in vivo confocal microscopy, animal models, and cell culture experiments, we may further the understanding of the sophisticated interactions of Herpes simplex virus with DCs in the cornea and the molecular mechanism associated with it. It has been shown that specific differentiation of DCs using immunohistochemistry, flow cytometry, and polymerase chain reaction analysis in both human and mice tissues and viral tissue infections are integral to increasing understanding. As for in vivo confocal microscopy, it holds promise as it is the least invasive and a real-time investigation. These tools will facilitate the discovery of various targets to develop new treatments.

Introduction The mechanism by which immunity to herpes simplex virus (HSV) is initiated is not completely defined. HSV initially infects the stratified squamous epithelium of the anogenital mucosa prior to entering nerve endings. We have recently reported that topical application of HSV-1 to human foreskin explants results in infection of epidermal Langerhans cells (LCs) which then emigrate into the dermis where they expressed the maturation marker CD80 and formed large cell clusters with BDCA3+ subsets of DC-SIGN+ and dermal dendritic cells (DCs) and. HSV-expressing LC fragments were observed inside the dermal DCs/macrophages. No other infected epidermal cells interacted with the dermal DCs (Kim et al. Plos Pathogens, 2015). Methods Therefore, we isolated LCs and dermal DCs from large abdominal skin specimens by collagenase digestion and flow sorting. LCs were pulsed with fluorescent tagged HSV and co-cultured with a subset of (BDCA3+) dermal DCs. Results All infected LCs showed markers of apoptosis at 18 hr p.i. Approximately 50% of BDCA3+ DCs co-localised with infected LCs and in some cases fragments of infected LCs were observed within the dermal DC cytoplasm. Such colocalizaton of HSV antigen bearing LCs and dermal DC subsets, was also detected within biopsies of initial genital herpes lesions. The mechanism of interaction of apoptotic LCs and dermal DCs, and uptake by phagocytosis are being determined. Conclusion Thus, a viral antigen relay takes place where HSV infected LCs slowly die by apoptosis during migration to the dermis and are taken up by dermal DCs by phagocytosis for subsequent antigen presentation. This provides a rationale for targeting these dermal DCs for mucosal or perhaps intradermal HSV immunisation. Disclosure of interest statement No pharmaceutical grants were received in the development of this study.

The mechanism by which immunity to Herpes Simplex Virus (HSV) is initiated is not completely defined. HSV initially infects mucosal epidermis prior to entering nerve endings. In mice, epidermal Langerhans cells (LCs) are the first dendritic cells (DCs) to encounter HSV, but it is CD103+ dermal DCs that carry viral antigen to lymph nodes for antigen presentation, suggesting DC cross-talk in skin. In this study, we compared topically HSV-1 infected human foreskin explants with biopsies of initial human genital herpes lesions to show LCs are initially infected then emigrate into the dermis. Here, LCs bearing markers of maturation and apoptosis formed large cell clusters with BDCA3+ dermal DCs (thought to be equivalent to murine CD103+ dermal DCs) and DC-SIGN+ DCs/macrophages. HSV-expressing LC fragments were observed inside the dermal DCs/macrophages and the BDCA3+ dermal DCs had up-regulated a damaged cell uptake receptor CLEC9A. No other infected epidermal cells interacted with dermal DCs. Correspondingly, LCs isolated from human skin and infected with HSV-1 in vitro also underwent apoptosis and were taken up by similarly isolated BDCA3+ dermal DCs and DC-SIGN+ cells. Thus, we conclude a viral antigen relay takes place where HSV infected LCs undergo apoptosis and are taken up by dermal DCs for subsequent antigen presentation. This provides a rationale for targeting these cells with mucosal or perhaps intradermal HSV immunization., Author Summary Herpes Simplex Virus (HSV) is a highly prevalent virus that causes cold sores and genital herpes but also increases the chance of contracting HIV by several folds. In fact, most new cases of HIV in Africa occur in people infected with HSV. Thus, a protective HSV vaccine would have a large impact on public health. Currently, the process by which immunity to HSV is generated is incompletely understood. Paradoxically, the first immune cells to become infected, Langerhans cells in the epidermis, are not the cells that initiate the immune response, while the dermal dendritic cells thought to be responsible for initiating the immune response are not likely to be infected. Here, we have shown, in human skin models and genital herpes lesion biopsies, an interaction between these dendritic cells that could relay HSV to the lymph node. HSV is taken up by the epidermal Langerhans cells that then migrate into the dermis, die and are taken up by another subset of dermal dendritic cells—the homologs of those in mice which stimulate HSV-specific T cells in the lymph node. Thus, a mucosal or intradermal vaccine targeting these two dendritic cells may be required.