Regulating Osteogenic Fate: How Dexamethasone Targets the CAST-CAPN1-ATP5A1 Axis in BMSCs

Osteoporosis is a common systemic metabolic disease, leading to increased bone fragility and risk of fractures. Research has shown that Adenosine triphosphate (ATP) synthase, H+transporting, mitochondrial F1 complex, alpha subunit 1 (ATP5A1), a crucial component in ATP production, is inhibited in dexamethasone (DEX)-induced osteoblasts. Therefore, this study aimed to investigate the molecular mechanism underlying the inhibitory impact of DEX on osteogenic differentiation in rat bone marrow mesenchymal stem cells (BMSCs).

Rat BMSCs were treated with varying concentrations of DEX for 14 days, followed by subsequent analyses. The expression levels of calpastatin (CAST), calpain 1 (CAPN1), and ATP5A1 were assessed using quantitative reverse transcription polymerase chain reaction (qRT-PCR) and Western blotting analyses. Furthermore, osteogenic marker proteins and ATP activity were evaluated employing Western blotting analysis and enzyme-linked immunosorbent assay (ELISA). Moreover, to determine the regulatory role of DEX on the CAST-CAPN1 axis, overexpression plasmids for CAST (oe-CAST) and CAPN1 (oe-CAPN1) were constructed. Additionally, osteogenic differentiation and ATP activity in BMSCs were analyzed using qRT-PCR, Western blotting, Alizarin Red S staining, and ELISA.

With increasing concentrations of DEX, the expression of the CAST-CAPN1-ATP5A1 axis in BMSCs was significantly altered (p < 0.05). DEX downregulated the levels of osteogenic markers, including Runt-Related Transcription Factor 2 (RUNX2), alkaline phosphatase (ALP), and osteopontin (OPN), while reducing ATP activity (p < 0.05). However, oe-CAST partially mitigated the inhibitory effects of DEX on osteogenic differentiation and ATP activity (p < 0.05). In contrast, oe-CAPN1 exacerbated the effects of DEX and reversed the regulatory impact of CAST (p < 0.05).

DEX inhibits osteogenic differentiation and reduces ATP activity in BMSCs by modulating the CAST-CAPN1 axis.