Phytoestrogen—rich Pueraria mirifica (PM) tuberous extract is a promising candidate for the development of anti—osteoporosis drugs for postmenopausal women, but its action has never been validated in humans or in non—human primates, which are more closely related to humans than rodents. In vitro study of non—human primate osteoblasts is thus fundamental to prepare for in vivo studies of phytoestrogen effects on primate bone. This study aimed to establish a culture system of baboon primary osteoblasts and to investigate the effects of PM extract and its phytoestrogens on these cells. Primary osteoblasts from adult baboon fibulae exhibited osteoblast characteristics in regard to proliferation, differentiation, mineralization, and Puerarin estrogen receptor expression. They responded to 17β—estradiol by increased proliferation rate and mRNA levels of alkaline phosphatase (ALP), type I collagen, and osteocalcin. After being exposed for 48 h to 100 μg/ml PM extract, 1000 nM genistein, or 1000 nM puerarin, primary baboon osteoblasts markedly increased the rate of proliferation and mRNA levels of ALP and type I collagen without changes in Runx2, osterix, or osteocalcin expression. PM extract, genistein, and puerarin also decreased the RANKL/OPG ratio, suggesting that they could decrease osteoclast—mediated bone resorption. However, neither PM extract nor its phytoestrogens altered calcium deposition in osteoblast culture. In conclusion, we have established baboon primary osteoblast culture, which is a new tool for bone research and drug discovery. Furthermore, the present results provide substantial support for the potential of PM extract and its phytoestrogens to be developed as therapeutic agents against bone fragility.
In conclusion, we have provided strong support for the potential of tuberous extract of PM from the Northern Thailand and its phytoestrogens (genistein and puerarin) to be developed as therapeutic agents against bone loss in postmenopausal women. PM extract, genistein, and puerarin could enhance proliferation and expression of ALP and type I collagen in primary baboon osteoblasts, which might, in turn, promote bone formation. A decrease in the mRNA level of RANKL further suggested that PM extract was capable of suppressing osteoclast—mediated bone resorption. Further investigation is, however, required to compare the osteogenic efficacy of PM extract with extracts from other related species, such as Chinese P. lobata and Pueraria thomsonii. The establishment of primary baboon osteoblast culture has also provided a new tool for studying osteoblast biology as well as for phytomedical and translational research. Thus, the baboon osteoblasts can be useful in the evaluation of candidate therapeutic agents intended for treatment of metabolic bone diseases in humans, such as postmenopausal osteoporosis and age—related osteoporosis.
Wacharaporn Tiyasatkulkovita,b, Suchinda Malaivijitnondb,* ,
Narattaphol Charoenphandhuc,d, Lorena M. Haville , Allen L. Forde, John L. VandeBerge
aBiological Sciences Program, Faculty of Science, Chulalongkorn University, Bangkok, Thailand
bDepartment of Biology, Faculty of Science, Chulalongkorn University, Bangkok, Thailand
cDepartment of Physiology, Faculty of Science, Mahidol University, Bangkok, Thailand
dCenter of Calcium and Bone Research (COCAB), Faculty of Science, Mahidol University, Bangkok, Thailand
eSouthwest National Primate Research Center and Department of Genetics, Texas Biomedical Research Institute, San Antonio, TX 78245, USA
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