葛根素在体外促进成骨分化和抑制成脂分化。
Puerarin promotes osteogenesis and inhibits adipogenesis in vitro.
机构信息
Translational Medicine R&D Center, Institute of Biomedical and Health Engineering, Shenzhen Institutes of Advanced Technology, Chinese Academy of Sciences, Shenzhen, China.
出版信息
Chin Med. 2013 Aug 21;8(1):17. doi: 10.1186/1749-8546-8-17.
BACKGROUND
Puerarin (daidzein 8-C-glucoside) has potential on preventing osteoporosis. This study aims to investigate the effects of puerarin on osteogenesis and adipogenesis in vitro.
METHODS
CCK-8 assay, alkaline phosphatase (ALP) activity and Alizarin Red S were used to measure the effects of puerarin on proliferation, osteoblastic differentiation, and mineralization in osteoblast-like MC3T3-E1 cells. The effects of puerarin on adipogenesis were measured by Oil Red O staining and intracellular triglyceride level in preadipocyte 3T3-L1 cells. The mRNA and protein levels of osteogenesis- and adiopogenesis-related factors were detected by qRT-PCR and western blot, respectively. Further, the secreted osteocalcin levels and nuclear translocation of β-catenin were detected by ELISA and immunofluorescence assay, respectively.
RESULTS
As to osteogenesis, puerarin could stimulate proliferation (1 μM, P = 0.012; 10 μM, P = 0.015; 20 μM, P = 0.050), ALP activity (20 μM, P = 0.008) and calcium nodule formation (20 μM, P = 0.011) in a dose-dependent manner. Puerarin (20 μM) promoted osteocalcin secretion (P = 0.004) and the protein expression of both osteopontin (P = 0.001) and osteoprotegerin (P = 0.003). As to adipogenesis, puerarin suppressed adipocytes formation and intracellular triglyceride level (P = 0.001). In addition, puerarin (20 μM) decreased the mRNA and protein levels of CCAAT/enhancer binding protein α (P = 0.001, P = 0.002), proliferator-activated receptor γ (P = 0.005, P = 0.003), and adipocyte lipid-binding protein 4 (P = 0.001, P = 0.001). Moreover, phosphorylation of AKT1-Ser437 (10 μM, P = 0.003; 20 μM, P = 0.007) and GSK-Ser9 (10 μM, P = 0.005; 20 μM, P = 0.003), and the nuclear translocation of β-catenin (10 μM, P = 0.006; 10 μM, P = 0.002) were increased in 3T3-L1 cells treated by puerarin.
CONCLUSION
Puerarin promoted osteogenesis and inhibited adipogenesis in vivo, and Akt/GSK-3β/β-catenin signaling pathway was involved in the suppression of adipogenesis.
背景
葛根素（大豆苷元 8-C-葡萄糖苷）具有预防骨质疏松的潜力。本研究旨在探讨葛根素在体外成骨和成脂中的作用。
方法
用 CCK-8 法、碱性磷酸酶（ALP）活性和茜素红 S 检测葛根素对成骨样 MC3T3-E1 细胞增殖、成骨分化和矿化的影响。用油红 O 染色和前脂肪细胞 3T3-L1 细胞内三酰甘油水平检测葛根素对成脂的影响。用 qRT-PCR 和 Western blot 分别检测成骨和成脂相关因子的 mRNA 和蛋白水平。进一步通过 ELISA 和免疫荧光法检测骨钙素的分泌和β-连环蛋白的核转位。
结果
在成骨作用方面，葛根素可呈剂量依赖性地刺激增殖（1 μM，P=0.012；10 μM，P=0.015；20 μM，P=0.050）、ALP 活性（20 μM，P=0.008）和钙结节形成（20 μM，P=0.011）。葛根素（20 μM）促进骨钙素分泌（P=0.004）和骨桥蛋白（P=0.001）和骨保护素（P=0.003）的蛋白表达。在成脂作用方面，葛根素抑制脂肪细胞形成和细胞内三酰甘油水平（P=0.001）。此外，葛根素（20 μM）降低 CCAAT/增强子结合蛋白 α（P=0.001，P=0.002）、过氧化物酶体增殖物激活受体 γ（P=0.005，P=0.003）和脂肪细胞脂质结合蛋白 4（P=0.001，P=0.001）的 mRNA 和蛋白水平。此外，用葛根素处理的 3T3-L1 细胞中 AKT1-Ser437 的磷酸化（10 μM，P=0.003；20 μM，P=0.007）和 GSK-Ser9（10 μM，P=0.005；20 μM，P=0.003）以及β-连环蛋白的核转位（10 μM，P=0.006；10 μM，P=0.002）增加。
结论
葛根素促进体内成骨和抑制成脂，并涉及 Akt/GSK-3β/β-连环蛋白信号通路抑制成脂。
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