P058 Inhibition of microRNA-29a in human bone marrow mesenchymal stem cells augments their immunosuppressive properties
A. Oikonomopoulos*1, C. Polytarchou2, M. Hatziapostolou2, G. Koukos2, D.W. Hommes1
1UCLA, Medicine, Division of Digestive Diseases, Center for Inflammatory Bowel Diseases, Los Angeles, United States, 2UCLA, Medicine, Division of Digestive Diseases, Center for Systems Biomedicine, Los Angelos, United States
Mesenchymal stem cells (MSC) offer new therapeutic strategies for refractory IBD due to their potent immunosuppressive properties that can be augmented by stimulation (priming) with pro-inflammatory cytokines. MSC have been proven beneficial in various immunological disorders, including graft versus host disease, ulcerative colitis and Crohn's disease. However, the exact regulatory mechanisms of the MSC immunosuppressive properties remain elusive. MicroRNAs (miRNA) are non-coding RNAs that regulate gene expression, participating in the pathogenesis of several disorders, IBD included. The role of miRNA on the immunosuppressive properties of MSC remains unknown. We hypothesized that miRNA may be involved in the molecular regulation of the immunosuppressive properties of adult BMMSC and that manipulation of miRNA levels might potentiate the salutary properties of BMMSC.
Human donor derived bone marrow MSC (BMMSC) (n=3) were primed with IFN-γ (50ng/mL, 24 hours). MiRNA profiling (800 miRNAs) was performed by Nanostring nCounter hybridization assays (n=3). Cytokine expression was determined in primed BMMSC via multiplex immunoassays (Bioplex, 40 cytokines) (n=3). MSC immunomodulatory properties were accessed via co-culture assays (n=3) of resting and primed BMMSC with human CFSE-loaded peripheral blood mononuclear cells (PBMC). Gene expression was determined by quantitative RT-PCR. Transfection of BMMSC with miRNA-29a oligonucleotides (inhibitors) or with scramble was performed by lipofectamine assays.
MiRNA-29a was found to be the most robustly expressed miRNA in rested (non-primed) BMMSC and it was the most highly up-regulated following IFN-γ priming (1.7-fold). MiRNA-29a was found to be 2-fold up-regulated in independent samples of IFN-γ primed BMMSC by conventional real-time PCR. Cytokine profiling revealed that IFN-γ induces the expression of numerous immunomodulators including IDO1, CCL5, TNFSF10, CCL2, CXCL16, CCL7, CCL13, CX3CL1, CXCL9, and CCL8. Inhibition of miRNA-29a altered the expression of various cytokines such as IDO1, CCL5, CCL2 and TNFSF10. Finally, co-culture assays between resting and primed BMMSC with human PBMC revealed that inhibition of miRNA-29a in IFN-γ primed BMMSC augments their immunosuppressive properties in comparison to scramble transfected counterparts.
Our data strongly support the notion that miRNA-29a is a novel regulator of the immunosuppressive properties of human BMMSC. MiRNA-29a regulates the expression of numerous cytokines in BMMSC. Understanding the molecular circuits controlling the immunomodulatory functions of MSC will facilitate the design of cell therapy trials for IBD and other inflammatory diseases.