Study On Differentiation Characteristics Of Langerhans Cells Derived From Cord Blood And Their Mechanisms Of Immune Response To A Microbial Metabolite IAId

Epidermal Langerhans cells(LC)play a critical role in regulating immune response of the skin as antigen-presenting cells.However,the development of LCs and their function in sensing external stimuli are still unclear.In this study,our research is divided into two parts.The first part is mainly about the study of differentiation characteristics of LCs by using CD34+ cord blood cell-derived LC model.In the second part,we explore the response of LCs to a skin microbial metabolite.Using flow cytometry,we first analyzed the surface phenotype of different LC subsets,CD 1 a+CD207-,CD 1 a+CD20710 and CD1ahiCD207hi.CD1a+CD207loLCs and CD1ahiCD207hiLCs differentiated from CD1a+CD207-DCs exhibited phenotypic and functional characteristics similar to resident LCs and migratory LCs,respectively.CD207 mediated the recognition and uptake of Der p2 by LCs specifically,in which CD207hiLCs played a dominant role.By single-cell RNA sequencing,we further demonstrated the heterogeneity of LC population.Here,we similarly identified two subpopulations,CD207+Type 1 LCs and CD207lo/-Type 2 LCs,that showed a one-to-one correspondence with CD207hiLCs and CD207loLCs distinguished by flow cytometry.Differentiation of Type 1 LCs was regulated by PU.1,while that of Type 2 LCs was regulated by Id2.In summary,our research uncovers the heterogeneity of cord blood-derived LCs at both transcriptome and protein levels.The finding of LC subsets with different phenotypes and functions can benefit us to better study the development,differentiation and function of LCs.Atopic derrmatitis(AD)is a common inflammatory skin disease in which microbiota and their metabolites are involved in the pathogenesis.Indole-3-aldehyde(IAId),as a microbial metabolite of tryptophan,was found significantly decreased on the surface skin of AD patients.However,little is known about the cellular and molecular mechanisms between IAId and skin inflammation.In the second part,we investigated the effects of IAId on cord blood-derived LCs.The data indicated that IAId induced LC maturation,and acted as a negative regulator by suppressing the production of TNF-a and IL-4,while up-regulating IL-12p70 and IL-10.LCs activated by IAId inhibited CD4+T cell proliferation.Moreover,the levels of IL-10 and IFN-y secreted by CD4+T cells were significantly higher in IAId-treated group,whereas the secretion of IL-6 decreased,consequently regulated Thl/Th2 balance.Aryl hydrocarbon receptor(AhR)signaling pathway was measured,and a significant increase was found in the expression of CYP1A1,CYP1B1 and indole-2,3-dioxygenase(IDO)upon IAId treatment.The expression of IL-10 and IL-10 receptor decreased after the application of an AhR antagonist,suggesting that AhR was required for the effects of IAId.Furthermore,we found that IAId exerted its effect on LCs by down-regulating high affinity receptor for IgE,FcεRI,not only in mRNA and protein levels,but through promoting ubiquitination and subsequent degradation.We then confirmed that regulation of FcεRI expression was in an AhR-dependent pathway by using AhR siRNA transfected into LCs and the antagonist.Taken together,we show the important role and mechanism of IAId in negative regulation of immune response through LCs for the first time.It provides new strategies for the treatment of skin disorders such as AD with application of particular microbial metabolites.