Abstract
Purpose: To investigate the effect of leonurusine hydrochloride (LH) on osteogenic differentiation, and the involvement of Wnt/β-catenin route and runt-associated transcription factor 2 (Runx2) in the process.
 Methods: Mouse osteogenic precursor cell line MC3T3-E1 was used. The cells were assigned to control and three other groups treated with increasing doses of LH, i.e., 0.1, 1, and 10 μmmol/L. Cellular differentiation was determined in the various groups with respect to mRNA expressions of alkaline phosphatase (ALP), number of calcified nodules, Wnt/β-catenin pathway-related genes, viz, 5 (LRP5),β-catenin, GSK-3β, as well as mRNA expressions of related osteogenic differentiation genes, namely. alkaline phosphatase (ALP), alpha 1 type I collagen (Col1-α), Run×2, and osteogenic-related transcription factor antibody (Osterix) using reverse transcription polymerase chain reaction (RT-PCR).
 Results: The secretion levels of ALP in cells exposed to LH doses of 1 and 10 μmmol/L were significantly increased, relative to control values (p < 0.05). There were marked increases in mRNA levels of LRP5 and β-catenin in cells treated with LH doses of 1 and 10 μmmol/L compared with the control group, while GSK-3β mRNA expression levels were significantly lower than that in control group(p < 0.05). However, the expression levels of ALP, Col1-α, Run×2, Osterix mRNA in cells exposed to LH at doses of 1 and 10 μmmol/L were significantly higher than the corresponding expression levels in control cells (p < 0.05).
 Conclusion: Leonurusine hydrochloride may promote osteogenic differentiation and bone mineralization via a mechanism involving activation of the Wnt/β-catenin pathway and up-regulation of Run×2 expression.
 Keywords: Leonurusine hydrochloride, Wnt/β-catenin, Runx2, Osteogenic differentiation
Highlights
Osteoporosis comprises two major pathological types: primary osteoporosis and secondary osteoporosis
Alkaline phosphatase (ALP) levels were significantly higher in cells treated with Leonurine hydrochloride (LH) at doses of 1 and 10 μmmol/L than in control cells (p < 0.05)
The expression levels of LRP5 and β-catenin mRNA were markedly higher in cells exposed to LH doses of 1 and 10 μmmol/L than in control, while GSK-3β levels were markedly reduced, relative to control level
Summary
Osteoporosis comprises two major pathological types: primary osteoporosis and secondary osteoporosis. Primary osteoporosis is more prevalent than secondary osteoporosis. Reduced estrogen levels in postmenopausal women leads to increased bone resorption and decreased bone formation, which in turn leads to disorderly changes in bone microstructure, loss of calcium ions, and significant reduction in bone density and bone strength. The incidence of Current clinical data show that osteoporotic drugs improve the patient's condition to a certain extent, but they do not enhance bone density, nor do they promote bone microstructure recovery in patients [3]. Parathyroid hormone analogues are used for enhancing bone formation, their clinical application is limited by the fact that they may exhibit carcinogenic potential [4]. Clinical research has focused on enhancing femoral bone formation and improving osteoblast differentiation. Leonurine hydrochloride (LH) is extracted from Leonurus sibiricus L., and it exerts multiple medicinal effects [5]
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