Glutamine-alpha-KG axis affects dentin regeneration and regulates odontogenic differentiation of mesenchymal adult stem cells via IGF2 m6A modification
Background: Multi-lineage differentiation of mesenchymal adult stem cells (m-ASCs) is crucial for tissue regeneration and accompanied with metabolism reprogramming, among which dental-pulp-derived m-ASCs has obvious advantage of easy accessibility. Stem cell fate determination and differentiation are closely related to metabolism status in cell microenvironment, which could actively interact with epigenetic modification. In recent years, glutamine-alpha-ketoglutarate(alpha-KG) axis was proved to be related to aging, tumorigenesis, osteogenesis etc., while its role in m-ASCs still lack adequate research evidence.Methods: We employed metabolomic analysis to explore the change pattern of metabolites during dental-pulp-derived m-ASCs differentiation. A murine incisor clipping model was established to investigate the influence of alpha-KG on dental tissue repairment. shRNA technique was used to knockdown the expression of related key enzyme-dehydrogenase 1(GLUD1). RNA-seq, m6A evaluation and MeRIP-qPCR were used to dig into the underlying epigenetic mechanism.Results: Here we found that the glutamine-alpha-KG axis displayed an increased tendency along with the odontogenic differentiation of dental-pulp-derived m-ASCs, same as expression pattern of GLUD1. Further, the key metabolite alpha-KG was found able to accelerate the repairment of clipped mice incisor and promote dentin formation. The promotion role of alpha-KG was also proved to be dose dependent, while the inhibition of glutamine-derived alpha-KG level via GLUD1 knockdown had the opposite effect. Further, extracellular matrix (ECM) function and PI3k-Akt signaling pathway was screened out to be widely involved in the process with insulin-like growth factor 2 (IGF2) participation. Inhibition of glutamine-alpha-KG axis may affect IGF2 translation efficiency via m6A methylation and can be significantly rescued by alpha-KG supplementation.Conclusion: Our findings indicate that glutamine-alpha-KG axis may epigenetically promote odontogenic differentiation of dental-pulp-derived m-ASCs and dentin regeneration, which provide a new research vision of potential dental tissue repairment therapy method or metabolite-based drug research.