The Role And Mechanism Of MiR-155-5p Regulating The Transdifferentiation Of Bone Marrow Mesenchymal Stem Cell To Gastric Cancer Tissue Derived Msc Like Cell

Objective: To elucidate the rugulation mechanism for transdifferation of bone marrow mesenchymal stem cell(BM-MSC) into gastric cancer tissue derived MSC-like cell(GC-MSC) in gastric cancer microenvironment, and to explore new targets for the treatment of gastric cancer.Methods: The phenotype and function differences of BM-MSC and GC-MSC were determined by immunofluorescence staining of α-SMA and FAP, qRT-PCR for cytokine expression, transwell migration and invasion assay, colony formation assay and nude mice subcutaneous tumor bearing assay for gastric cancer cells. The miR-155-5p expression of BM-MSC and GC-MSC were detected by qRT-PCR. The mi RNA-155-5p mimics was transfected into GC-MSC and miR-155-5p inhibitor was transfected into BM-MSC to analyze the effect of mi R-155-5p on transdifferation of BM-MSC into GC-MSC. Target genes of miR-155-5p were predicted by TargetScan and picTar software and validated by luciferase reporter assay. Inhibitors or si-RNAs of target genes were used to elucidate the mechanism of miR-155-5p inhibition promoting BM-MSC to obtain GC-MSC like phenotype and function. The expression of target genes of MSCs were detected in diffuse type and intestinal type gastric cancer tissue samples and gastritis to analyze the relationship between target genes and gastric cancer classification by immunohistochemical method. Isolated GC-MSC and GCN-MSC from gastric cancer and adjacent cancer tissues and detect the expression of target genes by western blot. Use target gene inhibitor or si-RNAs for downstream cytokines to treat GC-MSC and detect its phenotypic and functional changes to clarify the function of miR-155-5p regulation related signaling molecules on maintain the phenotype and function of GC-MSC.Results: GC-MSC exhibited the classic phenotype of reactive stroma cells, a stronger gastric cancer promoting capacity and lower expression of miR-155-5p compared to BM-MSC. Inhibition of mi R-155-5p by transfecting miR-155-5p inhibitor induced a phenotypical and functional transition of BM-MSC into GC-MSC-like cell, and the reverse experiment deprived GC-MSC of tumor-promoting phenotype and function. NF-kappa B p65(NF-κB p65) and inhibitor of NF-kappa B kinase subunit epsilon(IKBKE/IKKε) were identified as targets of mi R-155-5p and important for mi R-155-5p inhibitor activating NF-κB p65 in the transition. Inactivation of NF-κB p65 by si-NF-κB p65、si-IKBKE or pyrrolidine dithiocarbamic acid(PDTC) significantly blocked the effect of mi R-155-5p inhibitor on BM-MSC. The expression of IKBKE, NF-κB and phosphor-NF-κB of MSC in diffuse gastric cancer tissue were more abundance than in intestinal type of gastric cancer tissue, with positive rates of 83.5%, 80.8% and 65.0%, respectively. However, the positive rate of the three protein of MSC in gastritis tissue were only 19.3%, 0% and 4%. In GC-MSC, the expression of miR-155-5p was downregulated and NF-κB p65 protein was increased and activated. NF-κB p65 inactivation by PDTC or knockdown of its downstream cytokines reversed the phenotype and function of GC-MSC.Conclusions: Our findings indicate that compared to BM-MSC, GC-MSC exhibites the classic phenotype of reactive stroma cells, a stronger gastric cancer promoting capacity and lower expression of mi R-155-5p. miR-155-5p downregulation induces BM-MSC to acquire a GC-MSC-like phenotype and function depending on NF-κB p65 activation, which suggests a novel mechanism underlying the cancer associated MSC remodeling in the tumor microenvironment and provides an effective target and approach for gastric cancer therapy.