Beta-Ecdysone Protects Mouse Osteoblasts from Glucocorticoid-Induced Apoptosis In Vitro

Abstract

Glucocorticoid-induced osteoporosis is a common form of secondary osteoporosis. Glucocorticoids affect both bone formation and resorption, and prolonged glucocorticoid exposure can suppress osteoblast activities. beta-Ecdysone, found in many plants, is involved in protein synthesis, carbohydrate and lipid metabolism, and immunologic modulation. Here, we evaluated the effects of beta-ecdysone on osteoblast viability by assessing apoptosis following treatment with excess glucocorticoids. Mouse bone marrow stromal cells were induced to differentiate and grow into osteoblasts, and then treated with 10 µM glucocorticoid and 10, 1, or 0.1 µM beta-ecdysone. The expression levels of osteoblast growth and differentiation factors (runt-related transcription factor 2, osteogenic protein-1, and alkaline phosphatase), apoptosis-related genes (transformation-related protein 53, ataxia telangiectasia mutated protein, caspase-3, and caspase-8), and Akt1 and phospho-Akt (Thr308) were then assessed via alkaline phosphatase staining, acridine orange-propidium iodide staining, annexin V/PI apoptosis assay, real-time RT-PCR, and Western blot analyses. Notably, treatment with 10 µM glucocorticoid resulted in reduced osteoblast viability and the specific activity of alkaline phosphatase as well as reduced runt-related transcription factor 2, osteogenic protein-1, and alkaline phosphatase mRNA expression in vitro, indicating that glucocorticoid inhibited osteogenic differentiation. Moreover, glucocorticoid treatment yielded increased transformation-related protein 53, ataxia telangiectasia mutated protein, caspase-3, and caspase-8 expression and decreased Akt1 and phospho-Akt levels, indicating glucocorticoid-induced apoptosis. Meanwhile, beta-ecdysone inhibited glucocorticoid function, preserving the expression of Akt1 and phospho-Akt and reducing the expression of transformation-related protein 53, ataxia telangiectasia mutated protein, caspase-3, and caspase-8. Thus, beta-ecdysone prevented glucocorticoid-induced osteoblast apoptosis in vitro. These data highlight the potential for beta-ecdysone as a treatment for preventing the effects of glucocorticoid on bone growth.

Key words

glucocorticoid - osteoporosis - beta-ecdysone - apoptosis - gene expression

^* These authors contributed equally to this manuscript.

• References

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