title: Chlorogenic Acid Prevents Osteoporosis by Shp2/PI3K/Akt Pathway in Ovariectomized Rats.
abstract: Cortex Eucommiae is used worldwide in traditional medicine, various constituents of Cortex Eucommiae, such as chlorogenic acid (CGA), has been reported to exert anti-osteoporosis activity in China, but the mechanism about their contribution to the overall activity is limited. The aims of this study were to determine whether chlorogenic acid can prevent estrogen deficiency-induced osteoporosis and to analyze the mechanism of CGA bioactivity. The effect of CGA on estrogen deficiency-induced osteoporosis was performed in vivo. Sixty female Sprague-Dawley rats were divided randomly among a sham-operated group and five ovariectomy (OVX) plus treatment subgroups: saline vehicle, 17α-ethinylestradiol (E2), or CGA at 9, 27, or 45 mg/kg/d. The rats' femoral metaphyses were evaluated by micro-computed tomography (μCT). The mechanism of CGA bioactivity was investigated in vitro. Bone mesenchymal stem cells (BMSCs) were treated with CGA, with or without phosphoinositide 3-kinase (PI3K) inhibitor LY294002. BMSCs proliferation and osteoblast differentiation were assessed with 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2-H-tetrazolium bromide (MTT) and alkaline phosphatase, with or without Shp2 interfering RNA (RNAi). The results display that CGA at 27 and 45 mg/kg/day inhibited the decrease of bone mineral density (BMD) that induced by OVX in femur (p< 0.01), significantly promoted the levels of bone turnover markers, and prevented bone volume fraction (BV/TV), connectivity density (CoonD), trabecular number (Tb.N), trabecular thickness (Tb.Th) (all p< 0.01) to decrease and prevented the trabecular separation (Tb.Sp), structure model index (SMI)(both p< 0.01) to increase. CGA at 1 or 10 μM enhanced BMSC proliferation in a dose-dependent manner. CGA at 0.1 to 10 μM increased phosphorylated Akt (p-Akt) and cyclin D1. These effects were reversed by LY294002. CGA at 1 or 10 μM increased BMSC differentiation to osteoblasts (p< 0.01), Shp2 RNAi suppressed CGA-induced osteoblast differentiation by decreasing Shp2, p-Akt, and cyclin D1. This study found that CGA improved the BMD and trabecular micro-architecture for the OVX-induced osteoporosis. Therefore, CGA might be an effective alternative treatment for postmenopausal osteoporosis. CGA promoted proliferation of osteoblast precursors and osteoblastic differentiation of BMSCs via the Shp2/PI3K/Akt/cyclin D1 pathway.
authors.affiliation: Orthopaedic Department, The Second Affiliated Hospital of NanChang University, NanChang, JiangXi, China.|Regeneration and Rehabilitation Engineering Research Institute on Bone and Nerve of JiangXi, NanChang, JiangXi, China.|Orthopaedics Research Institute of Jiangxi, NanChuang University, NanChang, JiangXi, China.
authors.forename: Rong Ping
authors.initials: RP
authors.lastname: Zhou
authors.affiliation: Regeneration and Rehabilitation Engineering Research Institute on Bone and Nerve of JiangXi, NanChang, JiangXi, China.|Rehabilitation Department, The Second Affiliated Hospital of NanChang University, NanChang, JiangXi, China.
authors.forename: Si Jian
authors.initials: SJ
authors.lastname: Lin
authors.affiliation: Orthopaedic Department, The Second Affiliated Hospital of NanChang University, NanChang, JiangXi, China.|Orthopaedics Research Institute of Jiangxi, NanChuang University, NanChang, JiangXi, China.
authors.forename: Wen Bing
authors.initials: WB
authors.lastname: Wan
authors.affiliation: Rehabilitation Department, The Second Affiliated Hospital of NanChang University, NanChang, JiangXi, China.
authors.forename: Hui Ling
authors.initials: HL
authors.lastname: Zuo
authors.affiliation: Rehabilitation Department, The Second Affiliated Hospital of NanChang University, NanChang, JiangXi, China.
authors.forename: Fen Fen
authors.initials: FF
authors.lastname: Yao
authors.affiliation: Orthopaedic Department, The Second Affiliated Hospital of NanChang University, NanChang, JiangXi, China.|Orthopaedics Research Institute of Jiangxi, NanChuang University, NanChang, JiangXi, China.
authors.forename: Hui Bing
authors.initials: HB
authors.lastname: Ruan
authors.affiliation: Orthopaedic Department, The Second Affiliated Hospital of NanChang University, NanChang, JiangXi, China.|Orthopaedics Research Institute of Jiangxi, NanChuang University, NanChang, JiangXi, China.
authors.forename: Jin
authors.initials: J
authors.lastname: Xu
authors.affiliation: Orthopaedic Department, The Second Affiliated Hospital of NanChang University, NanChang, JiangXi, China.|Orthopaedics Research Institute of Jiangxi, NanChuang University, NanChang, JiangXi, China.
authors.forename: Wei
authors.initials: W
authors.lastname: Song
authors.affiliation: Orthopaedic Department, The Second Affiliated Hospital of NanChang University, NanChang, JiangXi, China.|Orthopaedics Research Institute of Jiangxi, NanChuang University, NanChang, JiangXi, China.
authors.forename: Yi Cheng
authors.initials: YC
authors.lastname: Zhou
authors.affiliation: Orthopaedic Department, The Second Affiliated Hospital of NanChang University, NanChang, JiangXi, China.|Orthopaedics Research Institute of Jiangxi, NanChuang University, NanChang, JiangXi, China.
authors.forename: Shi Yao
authors.initials: SY
authors.lastname: Wen
authors.affiliation: Regeneration and Rehabilitation Engineering Research Institute on Bone and Nerve of JiangXi, NanChang, JiangXi, China.|Rehabilitation Department, The Second Affiliated Hospital of NanChang University, NanChang, JiangXi, China.
authors.forename: Jiang Hua
authors.initials: JH
authors.lastname: Dai
authors.affiliation: Regeneration and Rehabilitation Engineering Research Institute on Bone and Nerve of JiangXi, NanChang, JiangXi, China.|Rehabilitation Department, The Second Affiliated Hospital of NanChang University, NanChang, JiangXi, China.
authors.forename: Mei Lan
authors.initials: ML
authors.lastname: Zhu
authors.affiliation: Regeneration and Rehabilitation Engineering Research Institute on Bone and Nerve of JiangXi, NanChang, JiangXi, China.|Orthopaedics Research Institute of Jiangxi, NanChuang University, NanChang, JiangXi, China.|Rehabilitation Department, The Second Affiliated Hospital of NanChang University, NanChang, JiangXi, China.
authors.forename: Jun
authors.initials: J
authors.lastname: Luo
mesh_terms: D000818:Animals; D015519:Bone Density / Q000187:drug effects*; D002454:Cell Differentiation / Q000187:drug effects; D049109:Cell Proliferation / Q000187:drug effects; D002478:Cells, Cultured; D002726:Chlorogenic Acid / Q000494:pharmacology*; D002867:Chromones / Q000494:pharmacology; D019938:Cyclin D1 / Q000378:metabolism; D005260:Female; D006801:Humans; D059630:Mesenchymal Stem Cells / Q000166:cytology; D009025:Morpholines / Q000494:pharmacology; D010006:Osteoblasts / Q000166:cytology* / Q000378:metabolism; D010012:Osteogenesis / Q000187:drug effects*; D015663:Osteoporosis, Postmenopausal / Q000517:prevention & control*; D010052:Ovariectomy; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism; D000081082:Phosphoinositide-3 Kinase Inhibitors; D054592:Protein Tyrosine Phosphatase, Non-Receptor Type 11 / Q000235:genetics / Q000378:metabolism; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism; D034622:RNA Interference; D034741:RNA, Small Interfering / Q000235:genetics; D051381:Rats; D017207:Rats, Sprague-Dawley
chemical_list: D002726:Chlorogenic Acid; D002867:Chromones; D019938:Cyclin D1; D009025:Morpholines; D019869:Phosphatidylinositol 3-Kinases; D000081082:Phosphoinositide-3 Kinase Inhibitors; D054592:Protein Tyrosine Phosphatase, Non-Receptor Type 11; D051057:Proto-Oncogene Proteins c-akt; D034741:RNA, Small Interfering; C530106:Ccnd1 protein, rat; C085911:2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one; C516727:Ptpn11 protein, rat