The role of protein C receptor(PROCR)in the whole glucan particle(WGP)induced maturation of mouse bone marrow-derived dendritic cells
Objective To investigate the effects of protein C receptor(PROCR)on the maturation and immune functions of mouse bone marrow-derived dendritic cells(BMDCs)induced by whole glucan particle(WGP).Methods The GSE2197 dataset was downloaded from the Gene Expression Omnibus(GEO)database of the National Center of Biotechnology Information(NCBI).By analyzing the microarray data of this dataset,differentially expressed genes(DEGs)were identified.Immune-related genes were obtained from the Immunology Database and Analysis Portal(IMMPORT)and intersected with DEGs to acquire immune-related differential genes.Key genes and relevant signaling pathways were identified and analyzed using specialized bioinformatics algorithms,including R packages.PCR was used to detect the differential genes in BMDCs cultured with or without WGP.In vitro,we extracted bone marrow from the tibias and fibulas of mice and induced the bone marrow cells to differentiate into BMDCs using FMS-like tyrosine kinase 3 ligand(Flt-3 L).After induction,we transfected these BMDCs and subsequently stimulated them with WGP.Flow cytometry was utilized to analyze the expression of surface molecules CD40,CD80,CD86,major histocompatibility complex Ⅰ/Ⅱ(MHC-Ⅰ/Ⅱ),and programmed cell death 1-ligand 1(PD-L1)to evaluate the immunophenotype of BMDCs.ELISA was used to measure the secretion levels of cytokines interleukin 12p40(IL-12p40),IL-6 and IL-10 in the supernatants.In T cell experiments,CD8+and CD4+T cells were isolated from the lymph nodes and spleens of OT-Ⅰ and OT-Ⅱ mice using magnetic bead sorting kits.These T cells were co-cultured with specific antigen ovalbumin(OVA)and the previously prepared BMDCs.Finally,flow cytometry was used to assess T cell proliferation and differentiation to evaluate the role of BMDCs in antigen-specific T cell responses.Results Analysis revealed that the PROCR gene was among the immune-related genes differentially expressed in WGP-induced BMDCs compared to CpG-stimulated BMDCs(GSE2197).Knockdown of the PROCR gene resulted in reduced surface expression of MHC-Ⅰ and increased secretion of IL-10 in WGP-stimulated BMDCs.Additionally,the ability of BMDCs to drive CD8+T cells to produce interferon γ(IFN-γ)was significantly weakened.Conclusion PROCR regulates the expression of MHC-Ⅰ and the secretion of IL-10 during the maturation of BMDCs induced by WGP,which in turn affects the proliferation and differentiation of CD8+IFN-γ+T cells in BMDCs.
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