A Primary Exploration Of Immunomodulatory Effects Mechanisms Of Ganoderma Atrum Polysaccharides On Dendritic Cells

The history of Ganoderma atrum being used in traditional medicine and functional food has been for thousands of years in China.Polysaccharides were found to be the main active ingredient of Ganoderma atrum.PSG-1 was a polysaccharide component extracted from Ganoderma atrum.The present study was designed to investigate the immunomodulatary effects and mechanisms of PSG-1 on dendritic cells(DCs)induced in vitro.The main research results obtained in this dissertation were concluded as follows:1.PSG-1 has direct immunomodulatory effects on DCs.PSG-1 increased cell surface molecule expression of MHC-II,CD80 and CD86,and enhanced the production of TNF-?,IL-12 p70,IL-6,IL-10,RANTES and MIP-1a in DCs.PSG-1-treated DCs promoted the proliferation of splenic T lymphocyte of mouse in mixed lymphocyte reaction.MAPK pathway is a key pathway for PSG-1 to regulate the immune function of DCs.2.Based on RNA-Seq technology,transcriptional sequence information of PSG-1 stimulated DCs was obtained.The total quantity of sequencing data of treatment group and control group were 6350423700 bp,6749478634 bp,6922562700 bp,6501046800 bp,6608047500 bp and 7174258200 bp respectively.The quantity of clean data in control group was 6174227174 bp,6552993304 bp and 6687389756 bp respectively.The quantity of clean data in PSG-1 group was 6260597776 bp,6272268250 bp and 6830301050 bp respectively.The quality control analysis of the sequencing data and the quality analysis of the expression quantity showed that the results of RNA-seq sequencing were of good quality.3.Genome database of Ensembl was used as a reference,and the percentage of sequencing results matching to mouse genome was up to 98.37%.There were 2231 differentially expressed genes between the two groups,in which 1014 genes were Up-regulated and 1217 genes were down-regulated.In KEGG pathway analysis results,240 signal pathways were involved in PSG-1 stimulated DCs.In the most significantly changed 20 signaling pathways,the immune related pathways not only involved the MAPK pathway,but also inclued PI3K/Akt pathway and JAK-STAT pathway.In addition,transformations in cytokine receptor interaction,ECM receptor interaction,complement,adhesion and adhesion molecules and other pathways indicated that PSG-1 changed secretory function,adhesion function and migration ability of DCs.4.In the RNA-Seq data,a part of differentially expressed genes which involved in the NF-?B and PI3K/Akt signaling pathway were selected for further verification,and the RNA-Seq results were consistent with the RT-q PCR results.Secretions of cytokines(TNF-? and IL-6)and surface molecules(MHC II molecules,CD80 and CD86)of DCs via PSG-1 inducing were rapidly reduced by NF-?B and PI3K/Akt pathway inhibitors,which showed that the results of RNA-Seq were reliable and reproducible.5.PSG-1 has indirect immunomodulatory effects on DCs in the case of intestinal epithelial cell barrier.Based on a Caco-2-DCs co-culture model,it was found that expression of MHC II molecules,CD80 and CD86 on DCs and production of TNF-? and IL-6 of DCs were indirectly promoted by high dose of PSG-1.But PSG-1 would not affect the permeability of Caco-2 monolayer.