Role of m⁶A Reader YTHDC2 in Differentiation of Human Bone Marrow Mesenchymal Stem Cells

  Objective  To study the regulatory effect of YTH domain-containing protein 2 (YTHDC2), a member of N⁶-methyladenosine (m⁶A) readers, on the osteogenic or adipogenic differentiation of human bone marrow mesenchymal stem cells (hBMSCs).

  Methods  YTHDC2 expression was knocked down by small interfering RNA (siRNA) in vitro. Osteogenic differentiation and adipogenic differentiation of hBMSCs were induced after YTHDC2 knockdown in order to study the changes in the differentiation phenotype of hBMSCs. Alkaline phosphatase staining (ALP staining) and alizarin red S staining were performed to examine osteogenic activity and calcium-nodular formation. Nile red staining was performed to examine lipid-droplet formation. Real-time quantitative polymerase chain reaction (RT-qPCR) was used to assess the expression of osteogenesis and adipogenesis-related genes. RNA-sequencing was performed to identify the transcriptome changes after YTHDC2 knockdown and to explore the potential regulatory mechanism by which YTHDC2 regulated the diferentiation of hBMSCs.

  Results  In this study, we found that siRNA-induced YTHDC2 knockdown resulted in increased ALP activity and calcium-nodular formation of hBMSCs during osteogenic differentiation, and significantly upregulated the expression of osteogenesis-related genes. In addition, the lipid-droplet formation capacity of hBMSCs was decreased during adipogenic differentiation. The expression of adipogenesis-related genes was significantly down-regulated. Gene-set enrichmen analysis of RNA-seq data showed that YTHDC2 was significantly correlated with ribosome function and mRNA-translation-related signaling pathways.

  Conclusion  The findings indicate that YTHDC2 knockdown can promote the osteogenic differentiation of hBMSCs and inhibit the adipogenic differentiation. YTHDC2 knockdown may cause changes in ribosome function.