Role Of Langerhans Cells And Skin Microbiome In Skin Sensitization And(or) Tolerance

Skin is one of the most important immune organs for human and mammals. It cansense and transmit the stimulation of the external environment, thus mediate and direct thesubsequent immune response. Latest study reported that skin immune system not onlypromote the activation of the immune response, but also induce immune tolerance. Ourprevious studies found that protein antigens applied to intact skin inhibited the subsequentimmune response induced by subcutaneous immunization using the same antigen.However, the exact mechanisms of skin-induced tolerance is not clear yet. Uptake ofexternal Ag depends on LCs and LCs has long been thought to have positive regulatoryfunction for promoting immune activation. However, a series of recent studies havepointed out that the major function of LCs might be maintaining skin homeostasis[1,2]. Themechanism of LCs’ function may be related to regulatory T cells (Treg)[3]. Factors affecting skin sensitization/tolerance include: skin barrier integrity, antigenproperties, antigen presenting cells, skin microbiome, etc. In addition to the LCs, we focuson skin microbiome to explore the mechanism of skin-induced immune response. In recentyears, epidemiological and clinical studies have shown that with the development ofsterile and disinfection methods, occurence of traditional infectious disease graduallydecreased, but incidence rates of allergic and autoimmune disease are gradually increased,suggesting that the human flora have an important influence on immune system[4].Previous studies mainly focused on the intestinal flora, and a series of important progresshave been made, revealing the important role of flora in the immune system. However, thecorrelation between skin microbiome, immune system and human diseases has just beennoticed and remain a fresh research field. The few studies at present have suggested thatskin microbiome participate in the immune response of the skin. The latest researchshowed that skin microbiome can promote immune cell activation, maintaining thesecretion of IL-17A, IL-1and other cytokines. These effects may not only promote theanti-infection immunity of skin, but also participate in arthritis, asthma, psoriasis and otherskin diseases[5]. But the exact effect and the mechanism of skin microbiome in skinsensitization/tolerance are unclear.Based on previous works, the present study used LCs defecient mice, Langerin-DTAmice, and intact-skin-induced skin-tolerance model to explore the function of LCs in skintolerance. At the same time, we applied topical antibiotics to mouse skin to producebacteria-reduction model, and then prepared contact hypersensitivity model, to analyze theeffect and mechanism of skin resident bacteria on skin-sensitization. We anticipated touncover the effect and mechanism of LCs and skin resident bacteria on skinsensitization/tolerance, and to reveal new functions of the skin immune system, providingnew ideas for the treatment of a variety of diseases through the skin in future.Objectives: Explore the role of LCs in skin-tolerance induced by protein antigen;Reveal the effect and mechanism of skin microbiome in skin contact hypersensitivity. Methods: Langerin-DTA mice and control mice were used to produce skin-tolerancemodel by applying protein antigen OVA onto intact skin, and serum OVA specific IgG1,IgG2aand total IgE were detected by ELISA. Antibiotics were used topically to Balb/cmice to eliminate skin microbiome, and reduction of skin bacteria was confirmed by skintissue culture. Then CHS model was prepared, and ear swelling and tissue inflammationwere compared afterward. For mechanism study, number of LCs, Treg cells in skindraining lymph nodes were analyzed.Results:(1)Role of LCs in protein antigens induced skin-tolerance: After application ofprotein antigen OVA onto the intact skin of wild mice, OVA specificIgG1, IgG2aand totalIgE after subcutaneous OVA immunization were much lower than those mice pretreatedwith PBS onto the skin; In Langerin-DTA+mice that are deficient in LCs, the serum OVAspecificIgG1, IgG2aand total IgE after SC immunization were significantly higher thanthat of Langerin-DTA-mice and close to PBS mice, suggesting that the mice can notachieve the OVA induced skin-tolerance after deletion of LCs.(2) Influence of the skin microbiome on CHS model: Topically application ofmupirocin on healthy skin of mice showed no alteration on the skin organization structureand there is no sign of inflammation, but the number of skin bacterial colony after tissueculture was much lower than that of control. The ear thickness of the oxazolone-inducedCHS model in Mupirocin group, Fusidic Acid group and Neomycin Sulfate group weremuch greater than that in the Emulsion bases group. The ear thickness in the Mupirocingroup, after sensitization by oxazolone, DNFB, or TNCB respectively, is much greaterthan that in the Emulsion bases group. These data indicated that the CHS was enhanced bythe reduction of the skin microbiome.(3) Mechanism of the effect of skin microbiome on CHS: Number of LCs in skindraining lymph nodes in Mupirocin group was higher than that of Emulsion bases groupafter sensitization. Number of Treg in skin draining lymph nodes in Mupirocin group was higher than Emulsion bases group after sensitization. These data indicated that LCs mightinduce the production of Treg to act against the robust sensitization in skin microbiomereduced skin.Conclusions: LCs played an important role in the process of skin tolerance; Skinresident bacteria reduction could increase the skin allergic reactions. Skin resident bacteriaplayed an important role in immune responses of skin.