Author(s): Dr. Nakul S Chandak and Dr. Pavan Kumar Guntaka

Abstract: Background: Although autophagy is crucial for the self-renewal and differentiation of stem cells, it is yet unknown how it affects bone marrow mesenchymal stem cells (BMSCs). This work used rapamycin (RAPA), a traditional autophagy agonist with osteo-regulatory effects, to examine the relationship between autophagy and osteogenic differentiation. Methods: After osteo induction (0, 7, 14, and 21 d), real-time quantitative polymerase chain reaction, western blotting, and immune-fluorescence were used to examine the autophagy of rat BMSCs (RT-qPCR). Alizarin red staining, alkaline phosphatase tests, and RT-qPCR/Western blotting quantification of bone sialoprotein, type 1 collagen, alkaline phosphatase, osteopenia, and Runt-related transcription factor 2 mRNA and protein levels were also used to assess osteogenic differentiation.Results: During osteogenic differentiation, the BMSCs' baseline autophagy level steadily reduced due to changes in BECN1, the ratio of lapidated (LC3-II) to unelucidated (LC3-I) microtubule-associated protein 1 light chain 3, and the expression of the specific autophagic target p62. On the other hand, it grew when RAPA concentration increased. Additionally, whereas 5 nM RAPA inhibited osteogenesis on days 14 and 21, 2 nM RAPA increased BMSC osteogenic differentiation on days 7 and 14. The autophagy inhibitor 3-methyladenine may reduce RAPA's ability to increase osteogenesis in BMSCs.Conclusions: During osteogenic differentiation, the baseline autophagy level of the BMSC reduced with time. Yet, a suitable RAPA concentration encouraged BMSC osteogenic differentiation through activation of autophagy.

DOI: 10.22271/27078345.2021.v3.i2c.145

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