MicroRNA-195-5p regulates carbon monoxide releasing molecule-3-induced
osteogenic differentiation of rat bone marrow mesenchymal stem cells by
targeting Wnt3a
Abstract
Background and Purpose: Bone marrow-derived mesenchymal stem cells
(BMSCs) are potential in promoting bone regeneration for their
multipotential differentiation capacity. Our previous study showed that
carbon monoxide releasing molecule-3 (CORM-3) promoted the osteogenic
differentiation of rat BMSCs. However, the mechanism was not clearly
understood. MicroRNAs (miRNAs) play a critical role in regulating the
osteogenic differentiation of BMSCs. We therefore investigated the role
of miR-195-5p in CORM-3-induced osteogenic differentiation. Experimental
Approach: The rat BMSCs were transfected with miR-195-5p mimics,
miR-195-5p inhibitor, pcDNA3.1-Wnt3a, Wnt3a siRNA or their corresponding
controls. The rat BMSCs osteogenic differentiation was assessed by
quantitative real-time polymerase chain reaction, Western blot and
alizarin red staining. In addition, dual luciferase assay was used for
the verification of targeting relationship between miR-195-5p and Wnt3a.
Key Results: miR-195-5p was down-regulated during the CORM-3-induced
osteogenic differentiation of rat BMSCs. Overexpression of miR-195-5p
inhibited CORM-3-induced osteogenic differentiation of rat BMSCs,
evidenced by significantly decreased mRNA and protein expressions of
runt-related transcription factor 2 and osteopontin, and matrix
mineralization demonstrated. Whereas, inhibition of miR-195-5p
expression enhanced osteogenic differentiation. miR-195-5p directly
targeted Wnt3a. Overexpression of Wnt3a increased CORM-3-induced
osteogenic differentiation of rat BMSCs, the opposite effect was
observed in Wnt3a-deficient cells. Moreover, the inhibitory effect of
miR-195-5p overexpression on CORM-3-induced osteogenic differentiation
was rescued by Wnt3a overexpression. Conclusion and Implications: These
results demonstrated that miR-195-5p may negatively regulate
CORM-3-induced osteogenic differentiation of rat BMSCs by targeting
Wnt3a, which provided insight into new mechanism of CORM-3, and
theoretical basis for bone regeneration.