Upregulation of osteoblastic differentiation marker mRNA expression in osteoblast—like UMR106 cells by puerarin and phytoestrogens from Pueraria mirifica

ABSTRACT

Phytoestrogens have attracted attention for their potential in the prevention of postmenopausal osteoporosis. Recently, phytoestrogen—rich herb Pueraria mirifica has been demonstrated to possess an osteogenic effect on bone in ovariectomized rats, but its underlying cellular mechanism was not known. Here, we investigated the effects of P. mirifica extract and its major isoflavone compound, puerarin, on cell viability, cell proliferation and the expression of differentiation markers in rat osteoblast—like UMR106bcells. After exposure to 17β—estradiol (E2), genistein, Pueraria mirifica extract and puerarin, proliferation but not viability 0fUMR106 cells was markedly decreased. Quantitative real—time PCR revealed that Pueraria mirifica extract and puerarin significantly increased the mRNA expression of alkaline phosphatase (ALP) and osteoprotegerin, but not Runx2, osterix or osteocalcin. Puerarin also decreased the mRNA expression of receptor activator of nuclear factor—KB ligand, an osteoclastogenic factor, suggesting that it could induce bone gain by enhancing osteoblast differentiation and suppressing osteoclast function. Furthermore, after an exposure to high affinity estrogen receptor(ER) antagonist (ICI182780), the E2—, genistein—, Pueraria mirifica extract— and puerarin—induced upregulation of ALP expressions were completely abolished. It could be concluded that Pueraria mirifica extract and puerarin induced osteoblast differentiation rather than osteoblast proliferation in an ER—dependent manner. The present findings, therefore, corroborated the potential benefit of Pueraria mirifica extract and puerarin in the prevention and treatment of postmenopausal osteoporosis.

In conclusion, the present study provided corroborative evidence for the first time that Pueraria mirifica extract and its major isoflavone, puerarin, were likely to enhance bone formation by promoting osteoblast differentiation, as indicated by the upregulation of ALP mRNA expression. Moreover, both Pueraria mirifica extract and puerarin may suppress osteoclast activity since they predominantly upregulated OPG expression, thereby decreasing the RANKL/OPG ratio. The present mRNA expression study also suggested that the effect of Pueraria mirifica extract on osteoblasts was mainly due to the action of puerarin because they similarly increased the mRNA levels of ALP and OPG, but not Runx2, osterix or osteocalcin. However, puerarin might have greater anti—osteoporotic potency than Pueraria mirifica extract as puerarin directly decreased the mRNA level of RANKL, which was the major osteoclastogenic factor. Although our study focused on changes at the transcriptional level, the present findings have underlined the potential of Pueraria mirifica extract and puerarin in prevention of bone loss, and the use of puerarin as an anti-osteoporotic drug would be an avenue worth exploring.

Wacharaporn Tiyasatkulkovit^a,c,d, Narattaphol Charoenphandhu^a,b,*, Kannikar Wongdee ^a,e,
Jirawan Thongbunchoo^a, Nateetip Krishnamra^a,b, Suchinda Malaivijitnond^c
aCenter of Calcium and Bone Research (COCAB), Faculty of Science, Mahidol University, Bangkok, Thailand
^bDepartment of Physiology, Faculty of Science, Mahidol University, Bangkok, Thailand
^cPrimate Research Unit, Department of Biology, Faculty of Science, Chulalongkorn University, Bangkok, Thailand
^dBiological Sciences Program, Faculty of Science, Chulalongkorn University, Bangkok, Thailand
^eOffice of Academic Management, Faculty of Allied Health Sciences, Burapha University, Chonburi, Thailand

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