Objective: To investigate the development of a minimal traditional Chinese medicine (TCM) formula using selected TCM ingredients and evaluating their biological activity with bone-specific in vitro tests. Finally, determining if the minimal formula can maintain bone mineral density (BMD) in a low bone mass (LBM)/osteoporosis (OP) model system.
Methods and results: Sixteen different TCM plant extracts were tested for estrogenic, osteogenic and osteoclastic activities. Despite robust activation of the full-length estrogen receptors α and β by Psoralea corylifolia and Epimedium brevicornu, these extracts do not activate the isolated estrogen ligand binding domains (LBD) of either ERα or ERβ; estrogen (17-β estradiol) fully activates the LBD of ERα and ERβ. E. brevicornu and Drynaria fortunei extracts activated cyclic AMP response elements (CRE) individually and when combined these ingredients stimulated the production of osteoblastic markers Runx2 and Bmp4 in MC3T3-E1 cells. E. brevicornu, Salvia miltiorrhiza, and Astragalus onobrychis extracts inhibited the Il-1β mediated activation of NF-κβ and an E. brevicornu/D. fortunei combination inhibited the development of osteoclasts from precursor cells. Further, a minimal formula containing the E. brevicornu/D. fortunei combination with or without a third ingredient (S. miltiorrhiza, Angelica sinensis, or Lycium barbarum) maintained bone mineral density (BMD) similar to an estradiol-treated control group in the ovariectomized rat; a model LBM/OP system.
Conclusion: A minimal formula consisting of TCM plant extracts that activate CRE and inhibit of NF-κβ activation, but do not behave like estrogen, maintain BMD in a LBM/OP model system.
Keywords: Anti-inflammatory; BMD, bone mineral density; BSA, bovine serum albumin; Bmp4, bone morphogenic protein 4; CRE, cyclic adenosine monophosphate response element; CREB, cyclic adenosine monophosphate response element binding protein; DEXA, dual-energy X-ray absorptiometry; DMSO, Dimethyl sulfoxide; Drynaria fortunei; E2, estradiol; ER, estrogen receptor; ERE, estrogen response element; Epimedium brevicornu; Estrogenic; FBS, fetal bovine serum; Fsk, forskolin; Hprt, hypoxanthine-guanine phosphoribosyl-transferase; IL-1, interleukin 1; LBD, ligand binding domain; LBM, low bone mass; M-CSF, macrophage colony-stimulating factor; MAPK, mitogen activated protein kinase; NF-κβ, nuclear factor kappa beta; OP, osteoporosis; Osteoporosis; PTH, parathyroid hormone; PTHrp, PTH related peptide; RANKL, receptor activator of nuclear factor kappa beta ligand; RLU, relative luminescence unit; ROI, region of interest; Runx2, runt-related transcription factor 2; SFM, serum free media; TCM, traditional Chinese medicine; TNFα, tumor necrosis factor alpha; TRAP, tartrate-resistant acid phosphatase; UAS, upstream activating sequence; cAMP, cyclic adenosine monophosphate; qPCR, quantitative polymerase chain reaction.