The Role And Mechanism Of CD207 In The Immune Response Mediated By Langerhans Cells To Skin Flora

Background In recent years,the research on skin flora has gradually become a new hot spot.The interaction between bacteria,viruses and fungi on the surface of the skin and host has a important significance for the maintenance of the physiological functions of the skin and is involved in the development of a variety of skin diseases.The skin flora plays an important role in the skin immune system.The animal experiments showed that the skin flora can affect the distribution and quantity of T cells in the skin,promote the inflammatory factor to release,enhance the immunity to pathogenic bacteria infection;on the other hand,skin flora can enhance the number of Treg cells in local skin draining lymph nodes,strengthen the skin immune tolerance and maintain the balance between immune response and immune tolerance.Our previous studies showed that the mice skin flora is reduced after using wide-spectrum antibiotics,then paint the sensitizing agent on the skin,observe difference of the contact hypersensitivity reaction in different groups of mice.The contact hypersensitivity increased significantly after the skin bacteria decreased,suggesting that the regulation of skin flora may be involved in the skin immune response,the specific mechanism in it needs to further study. Langerhans cells(LCs)classified in dendritic cells reside in the outer skin,LCs are in charge of recognising,uptaking,processing and presentating antigen,the LCs are matured after activation then migrate gradually to the local drainage lymph nodes and activate T cells in local draining lymph nodes,then mediate immune response to antigen.In recent years,it has been found that LC plays an important role in the immune tolerance of the skin,which is the most important one in the study of LCs' function.In the skin,LCs can induce Treg cells to produce IL-10,and play a role of immunosuppression.However,the specific role of LCs mediated immune response in the skin is still unclear,and further research is needed.Studies have shown that LCs do not express TLR2,4,5,which can recogize bacterial antigens,so they can not recognize the extracellular bacteria,and can not produce related immunity to bacteria.However,in recent years,the studies found that C type lectin receptor CD207 have been clarified that CD207 has the function of recognising microbios including HIV virus,mycobacteria,fungi,but whether CD207 residing in skin playing a role in recognising skin resident bacteria is still not clear.Aims: to further explicit the role of skin flora for regulating contact hypersensitivity,to clear the effect and the mechanism of LCs mediated by skin flora to regulate contact hypersensitivity,in which the role and mechanism of CD207 receptor on the surface of LCs.Methods: 1 15 wild type C57BL/6 mice were randomly divided into emulsion base group,mupirocin group and Staphylococcus epidermidis group,5 mice in each group.After shaving,emulsion base group mice were painted with emulsion base,mupirocin group mice were painted with mupirocin to establish bacteria reduction model,staphylococcus epidermidis group mice were applied with mupirocin to establish bacteria reduce model and then added staphylococcus epidermidis to save the flora.The three groups of mice were then sensitized and challenged by oxazolone,then observing the effect of CHS response on mice after changing of skin flora.2 Using CD207-DTA transgenic mice and wild type C57BL/6 mice,5 mice in each group,the mice were shaved and painted with emulsion base or mupirocin respectively to establish the skin flora reduced model,and to observe the response of CHS after changing of the skin flora when epidermal LCs loss.3 The mouse CD207 plasmid was transfected into HEK293 T cells,and staphylococcus epidermidis were incubated with that.Staphylococcus epidermidis were incubated with recombinant mouse CD207 protein,and we observe whether CD207 could combine Staphylococcus epidermidis.4 15 wild type C57BL/6 mice were randomly divided into emulsion base group,mupirocin group and Staphylococcus epidermidis group,5 mice in each group.After shaving,emulsion base group mice were painted with emulsion base,mupirocin group mice were painted with mupirocin to establish bacteria reduction model,staphylococcus epidermidis group mice were applied with mupirocin to establish the skin flora reduced model and then added staphylococcus epidermidis to save the flora,we observed the mechanism of that LCs and skin flora regulate the skin immune response,including the expression of CD80/86/273/274 and the expression of IL-1,IL-6,IFN-?,TNF-?and Rank L in the skin,and the secretion of IL-10 after adding bacteria.5 Using wild-type and CD207-DTA mice,5 mice in each group,tape strip sticking,causing the skin mild injury,adding Staphylococcus epidermidis,the differences of HE were observed about epidermal thickness in two group mices after same treatment.Result: 1 The skin flora inhabit contact hypersensitivity:using wild type mice after applying mupirocin,the changing of mice ear thickness and tissue sections showed that the skin inflammation was significantly increased;and after applying staphylococcus epidermidis to save,the changing of ear thickness and HE tissues showed that the mice skin inflammation significantly reduce,suggesting that skin flora can inhabit the response of CHS.2 LCs mediates the regulation of CHS response by skin flora:using CD207-DTA mice,the influence of the changing of the flora on CHS was not seen when epidermal LCs were lost,suggesting that LCs mediates the regulation of skin flora on the response of CHS.3 CD207 molecules can bind to staphylococcus epidermidis:HEK293T cells transfected with mouse CD207 plasmid were co-cultured with CTC staining staphylococcus epidermidis,and confocal laser scanning results showed that both of them were co-localized,and some of the staphylococcus epidermidis were embedded in the CD207 positive transfected cell membrane.Using staphylococcus epidermidis as the carrier,staphylococcus epidermidis was co-incubated with recombinant mouse CD207 protein,and it was found that recombinant mouse CD207 protein could bind to staphylococcus epidermidis.4 The regulation mechanism of immune response by skin flora and LCs:in wild type mice,combined with the CHS mouse model,after applying mupirocin and staphylococcus epidermidis,we found that CD80,CD86,CD273 and CD274 of CD207 postive cells were increased in skin draining lymph nodes when the skin flora is reduction,while the expression of IL-1? IL-6 and other cytokine were increased in the local skin lesions.However,the expression of CD80,CD86,CD273,CD274 and these inflammatory factors such as IL-1 and IL-6 in the local skin lesions decreased after adding staphylococcus epidermidis.It was also found that the ability of LCs to secrete IL-10 increased after the addition of staphylococcus epidermidis.5 LCs promotes inflammation of the skin after skin barrier damaged:using CD207-DTA mice,shaving and sticking with tape strip,then staphylococcus epidermidis was added on the back of mice for 1 week,the experimental area of skin was preparated for HE slice,the epidermal thickness after measurement in CD207-DTA group increased weakly than that in wild-type mice,suggesting that LCs could promote the skin inflammation when the skin barrier was damaged.Conclusion: Through the above research,we found that skin flora can regulate CHS inflammatory response by using the flora reduced model and flora resaved model and by using CD207-DTA transgenic mice which were LC constitutive deletion,we found that LC participates in the process of skin flora regulating skin inflammatory response;in vitro experiments showed that SE can bind to CD207 molecules,suggesting that LC can recognized SE through CD207.In addition,flow cytometry results showed ability of LC secretion of IL-10 cytokines was enhanced after adding SE,further proved that LC may play an important role in the regulation of immune response by skin flora.When the epidermal barrier is destroyed,SE can promote epidermal thickening,and LC play a role in it.