The stimulatory impact of d-δ-Tocotrienol on the differentiation of murine MC3T3-E1 preosteoblasts
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Osteoblasts and osteoclasts play essential and opposite roles in maintaining bone homeostasis. Osteoblasts fill cavities excavated by osteoclasts. The mevalonate pathway provides essential prenyl pyrophosphates for the activities of GTPases that promote differentiation of osteoclasts but suppress that of osteoblasts. Preclinical and clinical studies suggest that mevalonate suppressors such as statins increase bone mineral density and reduce risk of bone fracture. Tocotrienols down-regulate 3-hydroxy-3-methylglutaryl coenzyme A (HMG CoA) reductase, the rate-limiting enzyme in the mevalonate pathway. In vivo studies have shown the bone-protective activity of tocotrienols. We hypothesize that d-δ-tocotrienol, a mevalonate suppressor, induces differentiation of murine MC3T3-E1 preosteoblasts. Alizarin staining showed that d-δ-tocotrienol (0–25 μmol/L) induced mineralized nodule formation in a concentration-dependent manner in MC3T3-E1 preosteoblasts. d-δ-Tocotrienol (0–25 μmol/L), but not d-α-tocopherol (25 μmol/L), significantly induced alkaline phosphatase activity, an indicator of preosteoblast differentiation. The expression of differentiation marker genes including BMP-2 and VEGFα was stimulated dose dependently by d-δ-tocotrienol (0–25 μmol/L). Concomitantly, Western blot analysis showed that d-δ-tocotrienol down-regulated HMG CoA reductase. d-δ-Tocotrienol (0–25 μmol/L) had no impact on the viability of MC3T3-E1 preosteoblasts following 48-h incubation, suggesting lack of cytotoxicity at these doses. Tocotrienols and other mevalonate suppressors have potential in maintaining bone health.
KeywordsTocotrienol Preosteoblasts Differentiation Mevalonate Alizarin ALP Ras HMG CoA reductase BMP-2 VEGFα
- HMG CoA
3-Hydroxy-3-methylglutaryl coenzyme A
Small guanosine triphosphate-binding protein
Bone morphogenetic protein
Vascular endothelial growth factor
We thank Dr Jay Cao from USDA for providing technical assistance in the culture of MC3T3-E1 cells.
This work was partially supported by the Agriculture and Food Research Initiative Grant 2009-02941 from the USDA National Institute for Food and Agriculture, Texas Department of Agriculture Food and Fiber Research Program, and Texas Woman’s University Research Enhancement Program.
Compliance with ethical standards
Conflict of interest
The authors declare that they have no conflict of interest.
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