MicroRNA-9 (miR-9) induces differentiation of bone marrow mesenchymal stem cells (BMSCs). Autophagy contributes to the efficiency of neuronal differentiation in BMSCs. Does miR-9 activate autophagy?

Lower percentage of cells presenting NSE and MAP-2 (miR-9-, purple) miR-9 induced neuronal cell differentiation in BMSCs Relative miR-9 expression (Real-time PCR) Higher miR-9 expression in cells transfected with MiR-9-1-LV (miR-9+; red) Higher percentage of cells presenting neuronal cell markers NSE and MAP-2 (miR-9+, orange) Lower miR-9 expression in cells transfected with Anti-mmu-miR-9-1 inhibitor (miR-9-; blue) % of neuronal marker positive cells Lower percentage of cells presenting NSE and MAP-2 (miR-9-, purple) Western Blot assay

Autophagy activation by miR-9: Immunofluorescent staining of autophagic marker LC3 in differentiated BMSCs White arrows point at LC3 positive dots Higher expression of miR-9 (miR-9+) LC3 positive dots/cell More positive dots/cell of autophagic marker LC3 (A1; red) Lower expression of miR-9 (miR-9-) Less positive dots/cell of autophagic marker LC3 (A3; blue)

Autophagy activation by miR-9: Western Blot and LC3-II/LC3-I ratio Western blot assay Higher ratio of LC3-II/LC3-I Higher degree of autophagosome formation Higher expression of miR-9 (miR-9+) Higher LC3-II expression and LC3-II/LC3-I ratio (red) Lower expression of miR-9 (miR-9-) Lower LC3-II expression and LC3-II/LC3-I ratio (blue)

Conclusion miR-9 can promote neuronal differentiation in BMSC cells by modulating autophagic activity miR-9 could be used to further the study of the mechanisms of differentiation It could also be used to promote BMSC-based therapy of diseases affecting the nervous system However, the downstream mechanism of this autophagic modulation activity was not explored and necessitates more study