Langerhans cell homeostasis and activation is altered in hyperplastic human papillomavirus type 16 E7 expressing epidermis

It has previously been shown that expression of human papillomavirus type 16 (HPV) E7 in epidermis causes hyperplasia and chronic inflammation, characteristics of pre-malignant lesions. Importantly, E7-expressing epidermis is strongly immune suppressed and is not rejected when transplanted onto immune competent mice. Professional antigen presenting cells are considered essential for initiation of the adaptive immune response that results in graft rejection. Langerhans cells (LC) are the only antigen presenting cells located in normal epidermis and altered phenotype and function of these cells may contribute to the immune suppressive microenvironment. Here, we show that LC are atypically activated as a direct result of E7 expression in the epidermis, and independent of the presence of lymphocytes. The number of LC was significantly increased and the LC are functionally impaired, both in migration and in antigen uptake. However when the LC were extracted from K14E7 skin and matured in vitro they were functionally competent to present and cross-present antigen, and to activate T cells. The ability of the LC to present and cross-present antigen following maturation supports retention of full functional capacity when removed from the hyperplastic skin microenvironment. As such, opportunities are afforded for the development of therapies to restore normal LC function in hyperplastic skin

The effect of different types of online reviews on Instagram regarding users’ choice of a restaurant

The digital transformation in the hospitality and tourism industry has led to a multitude of upheavals and the development of new communication channels. Social media platforms have been widely used for the purpose of sharing information between consumers about products and services. More users share their experiences with restaurants with an online audience that goes far beyond their personal contacts. Online reviews provide valuable information for potential customers to facilitate their purchase decision. Additionally, restaurant managers benefit from feedback systems, as they are able to assess strengths and weaknesses of their services. Previous studies have shown that electronic Word-of-Mouth has had a significant effect on consumers’ decision-making process in the hospitality industry. The restaurant sector in particular is one of the most strongly influenced by social media platforms such as Instagram. This research study focuses on the effect of different types of online reviews on Instagram regarding users’ choice of a restaurant. An empirical approach was chosen to evaluate the effects of reviews on Instagram on consumer behavior in the restaurant sector. A survey designed to gain insights into user`s responses to relevant Instagram content collected 316 valid responses. The results show that content showcasing food quality, service quality and restaurant atmosphere has a significant and positive impact on trust, customer perceived value and thereby, purchase intention. This study provides valuable information to restaurants, identifies areas of research that can help them understand the power of Instagram and take advantage of online reviews on Instagram as a new marketing tool.A transformação digital na indústria hoteleira e turística tem levado a uma multiplicidade de convulsões e ao desenvolvimento de novos canais de comunicação. Mais usuários compartilham suas experiências com restaurantes com um público online que vai muito além de seus contatos pessoais. As análises online fornecem informações valiosas para os potenciais clientes para facilitar a sua decisão de compra. Além disso, os gerentes de restaurantes se beneficiam de sistemas de feedback, pois são capazes de avaliar os pontos fortes e fracos de seus serviços. Estudos anteriores mostraram que a palavra electrónica boca a boca teve um efeito significativo no processo de tomada de decisão dos consumidores na indústria hoteleira. Este estudo de pesquisa foca o efeito de diferentes tipos de revisões on-line na Instagram sobre a escolha dos usuários de restaurantes. Uma abordagem empírica foi escolhida para avaliar os efeitos das críticas sobre o Instagram na escolha de um restaurante. Uma pesquisa concebida para obter insights sobre as respostas dos usuários aos conteúdos relevantes do Instagram coletou 316 respostas válidas. Os resultados mostram que o conteúdo que mostra a qualidade dos alimentos, a qualidade do serviço e a atmosfera do restaurante tem um impacto significativo e positivo na confiança, no valor percebido pelo cliente e, portanto, na intenção de compra. Este estudo fornece informações valiosas aos restaurantes, identifica áreas de pesquisa que podem ajudá-los a entender o poder da Instagram e aproveitar as análises on-line da Instagram como uma nova ferramenta de marketing

Epidermal langerhans cells rapidly capture and present antigens from C-type lectin-targeting antibodies deposited in the dermis

Antigen-presenting cells can capture antigens that are deposited in the skin, including vaccines given subcutaneously. These include different dendritic cells (DCs) such as epidermal Langerhans cells (LCs), dermal DCs, and dermal langerin DCs. To evaluate access of dermal antigens to skin DCs, we used mAb to two C-type lectin endocytic receptors, DEC-205/CD205 and langerin/CD207. When applied to murine and human skin explant cultures, these mAbs were efficiently taken up by epidermal LCs. In addition, anti-DEC-205 targeted langerin CD103 and langerin CD103 mouse dermal DCs. Unexpectedly, intradermal injection of either mAb, but not isotype control, resulted in strong and rapid labeling of LCs in situ, implying that large molecules can diffuse through the basement membrane into the epidermis. Epidermal LCs targeted in vivo by ovalbumin-coupled anti-DEC-205 potently presented antigen to CD4 and CD8 T cells in vitro. However, to our surprise, LCs targeted through langerin were unable to trigger T-cell proliferation. Thus, epidermal LCs have a major function in uptake of lectin-binding antibodies under standard vaccination conditions

Enhancing design method training with insights from educational research–improving and evaluating a training course for a qualitative modelling method

Abstract This study presents an approach for identification and elimination of challenges in modelling in embodiment design. These challenges can be caused either by the modelling method or the corresponding training course. To investigate the efficacy of a modelling method, first challenges of the corresponding training course need to be addressed. The study is conducted at a training course of the modelling method of the Contact and Channel Approach. A situation analysis of the training course is conducted in three application with 45 participants. Based on the findings, the training course is improved through application of insights from educational research that correspond to the identified challenges. A concluding evaluation takes place with 20 participants. The improvement of the training course takes place based on identification of challenges in the four areas of didactic elements, content structure, visualization and practical modelling in evaluations. Modularization is needed for purposeful training of different target groups. An issue regarding the practical modelling indicates a clearer view on the efficacy of the modelling method. Article highlights Identification of challenges in a training course for qualitative modelling in embodiment design through free text evaluation in three applications. Clustering of the evaluation results enabled identification of suitable findings from educational research to eliminate challenges in the training course. Conflicts of objectives regarding content and time can be addressed by modularization, however, this increases the effort needed for investigations

Specific protein antigen delivery to human Langerhans cells in intact skin

Immune modulating therapies and vaccines are in high demand, not least to the recent global spread of SARS-CoV2. To achieve efficient activation of the immune system, professional antigen presenting cells have proven to be key coordinators of such responses. Especially targeted approaches, actively directing antigens to specialized dendritic cells, promise to be more effective and accompanied by reduced payload due to less off-target effects. Although antibody and glycan-based targeting of receptors on dendritic cells have been employed, these are often expensive and time-consuming to manufacture or lack sufficient specificity. Thus, we applied a small-molecule ligand that specifically binds Langerin, a hallmark receptor on Langerhans cells, conjugated to a model protein antigen. Via microneedle injection, this construct was intradermally administered into intact human skin explants, selectively loading Langerhans cells in the epidermis. The ligand-mediated cellular uptake outpaces protein degradation resulting in intact antigen delivery. Due to the pivotal role of Langerhans cells in induction of immune responses, this approach of antigen-targeting of tissue-resident immune cells offers a novel way to deliver highly effective vaccines with minimally invasive administration

Multi-Epitope DC Vaccines with Melanoma Antigens for Immunotherapy of Melanoma

Background/Objectives: The revolution for the treatment of melanoma came with the approval of checkpoint blockade antibodies. However, a substantial proportion of patients show primary or secondary resistance to this type of immunotherapy, indicating the need for alternative therapeutic strategies. Dendritic cells (DCs) of the skin are prime targets for vaccination approaches due to their potential to prime naïve T cells and their accessibility. This study aimed to develop and evaluate novel vaccines targeting the C-type lectin receptor DEC-205 to deliver melanoma-associated antigenic peptides to skin DCs. Methods: We cloned MHC-I-restricted peptides from the glycoprotein (gp)10025–33 and Tyrosinase-related protein (trp)2180–188 into the DEC-205 antibody sequence with modified peptide cutting sites from the OVA257–264 SIINFEKL peptide. We tested their potential to induce CD8+ T cell responses in both in vitro and in vivo settings. Tumor growth inhibition was evaluated in the transplantable B16.OVA melanoma murine model using a multi-epitope DC-based vaccine combining both peptides. Results: The cross-presentation of both gp100 and trp2 peptides was confirmed in vivo when peptide sequences were flanked by the OVA257–264 peptide cutting sites. Moreover, the combination of both antigenic peptides into a multi-epitope DC vaccine was required to inhibit B16.OVA melanoma growth. Conclusions: Our findings suggest that a DC-targeted vaccination approach using multiple epitopes deriving from melanoma antigens could represent a promising strategy for melanoma therapy

Dendritic cell-targeted therapy expands CD8 T cell responses to bona-fide neoantigens in lung tumors

Cross-presentation by type 1 DCs (cDC1) is critical to induce and sustain antitumoral CD8 T cell responses to model antigens, in various tumor settings. However, the impact of cross-presenting cDC1 and the potential of DC-based therapies in tumors carrying varied levels of bona-fide neoantigens (neoAgs) remain unclear. Here we develop a hypermutated model of non-small cell lung cancer in female mice, encoding genuine MHC-I neoepitopes to study neoAgs-specific CD8 T cell responses in spontaneous settings and upon Flt3L + αCD40 (DC-therapy). We find that cDC1 are required to generate broad CD8 responses against a range of diverse neoAgs. DC-therapy promotes immunogenicity of weaker neoAgs and strongly inhibits the growth of high tumor-mutational burden (TMB) tumors. In contrast, low TMB tumors respond poorly to DC-therapy, generating mild CD8 T cell responses that are not sufficient to block progression. scRNA transcriptional analysis, immune profiling and functional assays unveil the changes induced by DC-therapy in lung tissues, which comprise accumulation of cDC1 with increased immunostimulatory properties and less exhausted effector CD8 T cells. We conclude that boosting cDC1 activity is critical to broaden the diversity of anti-tumoral CD8 T cell responses and to leverage neoAgs content for therapeutic advantage

Targeted delivery of a vaccine protein to Langerhans cells in the human skin via the C-type lectin receptor Langerin

Human skin is a preferred vaccination site as it harbors multiple dendritic cell (DC) subsets, which display distinct C-type lectin receptors (CLR) that recognize pathogens. Antigens can be delivered to CLR by antibodies or ligands to boost antigen-specific immune responses. This concept has been established in mouse models but detailed insights into the functional consequences of antigen delivery to human skin DC in situ are sparse. In this study, we cloned and produced an anti-human Langerin antibody conjugated to the EBV nuclear antigen 1 (EBNA1). We confirmed specific binding of anti-Langerin-EBNA1 to Langerhans cells (LC). This novel LC-based vaccine was then compared to an existing anti-DEC-205-EBNA1 fusion protein by loading LC in epidermal cell suspensions before coculturing them with autologous T cells. After restimulation with EBNA1-peptides, we detected elevated levels of IFN-γ- and TNF-α-positive CD4+ T cells with both vaccines. When we injected the fusion proteins intradermally into human skin explants, emigrated skin DC targeted via DEC-205-induced cytokine production by T cells, whereas the Langerin-based vaccine failed to do so. In summary, we demonstrate that antibody-targeting approaches via the skin are promising vaccination strategies, however, further optimizations of vaccines are required to induce potent immune responses

Skin dendritic cells in melanoma are key for successful checkpoint blockade therapy.

BACKGROUND: Immunotherapy with checkpoint inhibitors has shown impressive results in patients with melanoma, but still many do not benefit from this line of treatment. A lack of tumor-infiltrating T cells is a common reason for therapy failure but also a loss of intratumoral dendritic cells (DCs) has been described. METHODS: We used the transgenic tg(Grm1)EPv melanoma mouse strain that develops spontaneous, slow-growing tumors to perform immunological analysis during tumor progression. With flow cytometry, the frequencies of DCs and T cells at different tumor stages and the expression of the inhibitory molecules programmed cell death protein-1 (PD-1) and T-cell immunoglobulin and mucin-domain containing-3 (TIM-3) on T cells were analyzed. This was complemented with RNA-sequencing (RNA-seq) and real-time quantitative PCR (RT-qPCR) analysis to investigate the immune status of the tumors. To boost DC numbers and function, we administered Fms-related tyrosine 3 ligand (Flt3L) plus an adjuvant mix of polyI:C and anti-CD40. To enhance T cell function, we tested several checkpoint blockade antibodies. Immunological alterations were characterized in tumor and tumor-draining lymph nodes (LNs) by flow cytometry, CyTOF, microarray and RT-qPCR to understand how immune cells can control tumor growth. The specific role of migratory skin DCs was investigated by coculture of sorted DC subsets with melanoma-specific CD8+ T cells. RESULTS: Our study revealed that tumor progression is characterized by upregulation of checkpoint molecules and a gradual loss of the dermal conventional DC (cDC) 2 subset. Monotherapy with checkpoint blockade could not restore antitumor immunity, whereas boosting DC numbers and activation increased tumor immunogenicity. This was reflected by higher numbers of activated cDC1 and cDC2 as well as CD4+ and CD8+ T cells in treated tumors. At the same time, the DC boost approach reinforced migratory dermal DC subsets to prime gp100-specific CD8+ T cells in tumor-draining LNs that expressed PD-1/TIM-3 and produced interferon γ (IFNγ)/tumor necrosis factor α (TNFα). As a consequence, the combination of the DC boost with antibodies against PD-1 and TIM-3 released the brake from T cells, leading to improved function within the tumors and delayed tumor growth. CONCLUSIONS: Our results set forth the importance of skin DC in cancer immunotherapy, and demonstrates that restoring DC function is key to enhancing tumor immunogenicity and subsequently responsiveness to checkpoint blockade therapy