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Immunological function of Langerhans cells in HIV in vitro and ex vivo infection
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Abstracts / Journal of Dermatological Science 84 (2016) e89–e180
chronic renal failure and atopic dermatitis. We have shown that repeated intra-dermal injection of antigen to dorsal skin of IgEtransgenic mice caused basophil-dependent, papulo-nodular skin lesions mimicking prurigo. Local cytokine profiles were characterized by elevated levels of interleukin (IL)-4, IL-13, IL-18, IL-31, IL-33, and TSLP. Nuclear translocation of pSTAT6 was observed in epidermal keratinocytes and dermal cells. Unexpectedly, however, STAT 6 (-/-) mice showed significant exacerbation of skin reactions compared with wild-type mice when treated with exogenous IgE and repeated antigen challenge. These were histopathologically characterized by fewer macrophages having M2-type phenotype and a larger number of basophils with a significant epidermal hyperplasia in STAT6 (-/-) mice than in wild-type mice. Local expression of TSLP and epidermal growth factor-like molecule amphiregulin (AREG) was remarkable in STAT6 (-/-) mice compared with WT mice. AREG was expressed by basophils in the lesional skin. In addition, bone marrow-derived basophils stimulated with TSLP and IgE plus antigen produced AREG. Thus, exacerbated prurigo-like lesions in the absence of STAT6 might be attributable, in part, to two possible mechanisms: impaired generation of inhibitory M2type macrophages and overproduction of basophil-derived AREG, leading to the promoted inflammatory processes and epidermal proliferation. http://dx.doi.org/10.1016/j.jdermsci.2016.08.462 P10-03[C07-07] Immunological function of Langerhans cells in HIV in vitro and ex vivo infection Takamitsu Matsuzawa 1,∗ , Tatsuyoshi Kawamura 1 , Youichi Ogawa 1 , Hiroaki Mitsuya 2 , Shinji Shimada 1 1
Dermatology, University of Yamanashi, Yamanashi, Japan 2 Department of Infectious Diseases and Department of Hematology, Kumamoto University School of Medicine, Kumamoto, Japan Langerhans cells (LCs) are suspected to be initial cellular targets for HIV infection. However, it is still unknown whether HIVinfected LCs induce HIV-specific CTL and regulatory T (Treg) cells, both of which play central roles in controlling HIV replication in infected individuals. To investigate the inducibility of HIVspecific CTL by HIV-infected LCs, naive CD8+ T cells obtained from HLA-A0201 healthy donors (n = 3) were cocultured with HIVBaL infected autologous monocyte derived LCs (mLCs). 7days later, the primed CD8+ T cells were restimulated by HIVBaL infected autologous mLCs for 7days. HIVBaL-infected mLCs could induce HIV gag tetramer+ CD8+ T cells. In addition, IFN␥ production of tetramer+ cells was significantly higher than that of tetramer-cells. Importantly, when we examined the inducibility of HIV-specific CTL by HIV-infected epidermal LCs by using an ex vivo skin explant model, similar results to mLCs were obtained (n = 2). Next, to investigate the inducibility of Treg cells by HIV-infected LCs, naive CD4+ T cells were cocultured with HIVBaL-infected autologous epidermal LCs, that emigrated from skin explants, for 6days. Treg cells have recently been separated into three subpopulations; FoxP3hi CD45RA- , FoxP3lo CD45RA+ , and FoxP3lo CD45RA- cells. Interestingly, the number of FoxP3hiCD45RA-Treg cells, but not other two subpopulations, induced by HIV-infected LCs was significantly lower than that by uninfected LCs. Moreover, in vitro suppression assay showed that Treg cells induced by HIV-infected LCs or uninfected LCs exhibited comparable suppressive function. Taken together, HIV-infected LCs may play crucial roles in eliciting
beneficial anti-HIV immune responses in the initial phase of HIV infection and leading to the incubation period from HIV infection until AIDS. http://dx.doi.org/10.1016/j.jdermsci.2016.08.463 P10-04[C07-06] EpCAM expressed by epidermal langerhans cells modulates immunization to an epicutaneously-applied protein antigen Takeshi Ouchi 1,2,∗ , Mark C. Udey 1 1
Dermatology Branch, Center for Cancer Research, National Cancer Institute, NIH, Japan 2 Department of Dermatology, Keio University School of Medicine, Tokyo, Japan Langerhans cells (LC) are required for efficient formation of Type 2 (IgG1 and IgE) Ab in response to foreign proteins that are applied to skin without adjuvant. Immunization is mediated via LC that extend dendrites through tight junctions (TJ) to acquire topically applied Ag and subsequently migrate to regional lymph nodes (LN). EpCAM (CD326) is expressed at high levels by LC. We previously reported that EpCAM promotes LC dendrite mobility, as well as LC migration within epidermis and from epidermis to regional LN in response to contact sensitizers. We also demonstrated that, in intestinal epithelial cells, EpCAM regulates the levels of expression and distribution of selected claudins. We hypothesized that in the absence of EpCAM, acquisition of Ag by LC might be impaired and topical immunization to proteins might be reduced. To test this, we studied conditional KO mice with EpCAM-deficient LC (cKO mice). Control (WT) and cKO LC dendrites docked with and penetrated the epidermal TJ with equal efficiencies. LC from both strains internalized surface biotinylated self-proteins. Unexpectedly, topical immunization of cKO mice with ovalbumin (Ova) led to the accumulation of increased numbers of LC in regional LN. This was accompanied by enhanced proliferation of adoptively transferred Ova- reactive OT-II cells and enhanced formation of Ova-specific Ab in cKO mice. These results indicate that EpCAM regulates topical immunization to protein acting via LC, but immunization is potentiated by EpCAM deficiency rather than inhibited. We propose that EpCAM-dependent mechanisms that regulate LC migration may differ in the setting of limited inflammation associated with topical immunization with protein as compared with the intense inflammation triggered by strong contact sensitizers. http://dx.doi.org/10.1016/j.jdermsci.2016.08.464 P10-05[C07-05] Skin resident CD8+ T cells display low cytotoxic potential in healthy skin and fail to fully mature in primary melanoma lesions Judith A. Seidel 1,4,∗ , Sian H. Henson 1 , Natalie Riddell 1 , Milica Vukmanovic-Stejic 1 , Malcolm H. Rustin 2 , Frank Nestle 3 , Katie Lacy 3 , Arne N. Akbar 1 1
University College London, London, United Kingdom 2 Royal Free Hospital, London, United Kingdom 3 Kings College London, London, United Kingdom 4 Kyoto University, Japan Melanoma lesions develop and spread in the skin but local tissue resident CD8+ T cells fail to contain the malignant cells.
Abstracts / Journal of Dermatological Science 84 (2016) e89–e180
chronic renal failure and atopic dermatitis. We have shown that repeated intra-dermal injection of antigen to dorsal skin of IgEtransgenic mice caused basophil-dependent, papulo-nodular skin lesions mimicking prurigo. Local cytokine profiles were characterized by elevated levels of interleukin (IL)-4, IL-13, IL-18, IL-31, IL-33, and TSLP. Nuclear translocation of pSTAT6 was observed in epidermal keratinocytes and dermal cells. Unexpectedly, however, STAT 6 (-/-) mice showed significant exacerbation of skin reactions compared with wild-type mice when treated with exogenous IgE and repeated antigen challenge. These were histopathologically characterized by fewer macrophages having M2-type phenotype and a larger number of basophils with a significant epidermal hyperplasia in STAT6 (-/-) mice than in wild-type mice. Local expression of TSLP and epidermal growth factor-like molecule amphiregulin (AREG) was remarkable in STAT6 (-/-) mice compared with WT mice. AREG was expressed by basophils in the lesional skin. In addition, bone marrow-derived basophils stimulated with TSLP and IgE plus antigen produced AREG. Thus, exacerbated prurigo-like lesions in the absence of STAT6 might be attributable, in part, to two possible mechanisms: impaired generation of inhibitory M2type macrophages and overproduction of basophil-derived AREG, leading to the promoted inflammatory processes and epidermal proliferation. http://dx.doi.org/10.1016/j.jdermsci.2016.08.462 P10-03[C07-07] Immunological function of Langerhans cells in HIV in vitro and ex vivo infection Takamitsu Matsuzawa 1,∗ , Tatsuyoshi Kawamura 1 , Youichi Ogawa 1 , Hiroaki Mitsuya 2 , Shinji Shimada 1 1
Dermatology, University of Yamanashi, Yamanashi, Japan 2 Department of Infectious Diseases and Department of Hematology, Kumamoto University School of Medicine, Kumamoto, Japan Langerhans cells (LCs) are suspected to be initial cellular targets for HIV infection. However, it is still unknown whether HIVinfected LCs induce HIV-specific CTL and regulatory T (Treg) cells, both of which play central roles in controlling HIV replication in infected individuals. To investigate the inducibility of HIVspecific CTL by HIV-infected LCs, naive CD8+ T cells obtained from HLA-A0201 healthy donors (n = 3) were cocultured with HIVBaL infected autologous monocyte derived LCs (mLCs). 7days later, the primed CD8+ T cells were restimulated by HIVBaL infected autologous mLCs for 7days. HIVBaL-infected mLCs could induce HIV gag tetramer+ CD8+ T cells. In addition, IFN␥ production of tetramer+ cells was significantly higher than that of tetramer-cells. Importantly, when we examined the inducibility of HIV-specific CTL by HIV-infected epidermal LCs by using an ex vivo skin explant model, similar results to mLCs were obtained (n = 2). Next, to investigate the inducibility of Treg cells by HIV-infected LCs, naive CD4+ T cells were cocultured with HIVBaL-infected autologous epidermal LCs, that emigrated from skin explants, for 6days. Treg cells have recently been separated into three subpopulations; FoxP3hi CD45RA- , FoxP3lo CD45RA+ , and FoxP3lo CD45RA- cells. Interestingly, the number of FoxP3hiCD45RA-Treg cells, but not other two subpopulations, induced by HIV-infected LCs was significantly lower than that by uninfected LCs. Moreover, in vitro suppression assay showed that Treg cells induced by HIV-infected LCs or uninfected LCs exhibited comparable suppressive function. Taken together, HIV-infected LCs may play crucial roles in eliciting
beneficial anti-HIV immune responses in the initial phase of HIV infection and leading to the incubation period from HIV infection until AIDS. http://dx.doi.org/10.1016/j.jdermsci.2016.08.463 P10-04[C07-06] EpCAM expressed by epidermal langerhans cells modulates immunization to an epicutaneously-applied protein antigen Takeshi Ouchi 1,2,∗ , Mark C. Udey 1 1
Dermatology Branch, Center for Cancer Research, National Cancer Institute, NIH, Japan 2 Department of Dermatology, Keio University School of Medicine, Tokyo, Japan Langerhans cells (LC) are required for efficient formation of Type 2 (IgG1 and IgE) Ab in response to foreign proteins that are applied to skin without adjuvant. Immunization is mediated via LC that extend dendrites through tight junctions (TJ) to acquire topically applied Ag and subsequently migrate to regional lymph nodes (LN). EpCAM (CD326) is expressed at high levels by LC. We previously reported that EpCAM promotes LC dendrite mobility, as well as LC migration within epidermis and from epidermis to regional LN in response to contact sensitizers. We also demonstrated that, in intestinal epithelial cells, EpCAM regulates the levels of expression and distribution of selected claudins. We hypothesized that in the absence of EpCAM, acquisition of Ag by LC might be impaired and topical immunization to proteins might be reduced. To test this, we studied conditional KO mice with EpCAM-deficient LC (cKO mice). Control (WT) and cKO LC dendrites docked with and penetrated the epidermal TJ with equal efficiencies. LC from both strains internalized surface biotinylated self-proteins. Unexpectedly, topical immunization of cKO mice with ovalbumin (Ova) led to the accumulation of increased numbers of LC in regional LN. This was accompanied by enhanced proliferation of adoptively transferred Ova- reactive OT-II cells and enhanced formation of Ova-specific Ab in cKO mice. These results indicate that EpCAM regulates topical immunization to protein acting via LC, but immunization is potentiated by EpCAM deficiency rather than inhibited. We propose that EpCAM-dependent mechanisms that regulate LC migration may differ in the setting of limited inflammation associated with topical immunization with protein as compared with the intense inflammation triggered by strong contact sensitizers. http://dx.doi.org/10.1016/j.jdermsci.2016.08.464 P10-05[C07-05] Skin resident CD8+ T cells display low cytotoxic potential in healthy skin and fail to fully mature in primary melanoma lesions Judith A. Seidel 1,4,∗ , Sian H. Henson 1 , Natalie Riddell 1 , Milica Vukmanovic-Stejic 1 , Malcolm H. Rustin 2 , Frank Nestle 3 , Katie Lacy 3 , Arne N. Akbar 1 1
University College London, London, United Kingdom 2 Royal Free Hospital, London, United Kingdom 3 Kings College London, London, United Kingdom 4 Kyoto University, Japan Melanoma lesions develop and spread in the skin but local tissue resident CD8+ T cells fail to contain the malignant cells.