The effect of 3-hydroxybutyrate on the in vitro differentiation of murine osteoblast MC3T3-E1 and in vivo bone formation in ovariectomized rats

Abstract

3-hydroxybutyrate (3HB), one of the degradation products of microbial biopolyesters polyhydroxyalkanoates (PHA), is a high energy metabolic substrate in animals. This study evaluated the effects of 3HB on growth of osteoblasts in vitro and on anti-osteoporosis in vivo. Alkaline phosphatase (ALP) assay, Van Kossa assay and Alizarin S red staining were used to study in vitro differentiation of murine osteoblast MC3T3-E1 cells. The intensity of in vitro cell differentiation measured in ALP was in direct proportion to the concentration of 3HB when it was lower than 0.01 g/L. Calcium deposition, a strong indication of cell differentiation, also showed an obvious increase with increasing 3HB concentration from 0–0.1 g/L, evidenced by Alizarin red S staining and Van Kossa assay. RT-PCR also showed significantly higher expression of osteocalcin (OCN) mRNA in MC3T3-E1 cells after 3HB administration. In vivo study using female Wistar rats (3 months old, n=80) allocated into normal, sham-operated or ovariectomized (OVX) group that led to decreasing bone mineral density (BMD), bone histomorphometry and biomechanics compared with normal and sham groups, had demonstrated that 3HB increased serum ALP activity and calcium deposition, decreased serum OCN, prevented BMD reduction resulting from OVX. All these led to enhanced femur maximal load and bone deformation resistance, as well as improved trabecular bone volume (TBV%). In conclusion, 3HB monomer containing PHA can be effective bone growth stimulating implant materials.