title: Effects of the combined Herba Epimedii and Fructus Ligustri Lucidi on bone turnover and TGF-β1/Smads pathway in GIOP rats.
abstract: BACKGROUND
  Kidney deficiency is the main pathogenesis of osteoporosis based on the theory of "kidney governing bones" in traditional Chinese medicine (TCM). Combined Herba Epimedii and Fructus Ligustri Lucidi, based on traditional Chinese formula Er-Zhi pills, were frequently used in TCM formulas that were prescribed for kidney tonifying and bone strengthening. However, it is unclear whether the combination of the two herbs may have a protective influence on glucocorticoid-induced osteoporosis (GIOP). The objective of this study was to evaluate the therapeutic effects and the underlying molecular mechanism of the decoction and the active fractions of the combined herbs in GIOP rats.
  
  
  METHODS
  Male Sprague-Dawley rats were divided into seven groups, including the normal control (NC), GIOP model (MO), active fractions low (100mg/kg, LAF), active fractions high (200mg/kg, HAF), decoction low (3.5g/kg, LD), decoction high (7g/kg, HD) and Calcium with Vitamin D3 (0.2773g/kg, CaD)-treated group. The GIOP model was established by intramuscular injection of dexamethasone (1mg/kg) twice a week for 8 weeks. Different kinds of indicators were measured, including bone mineral density (BMD), bone biomechanical properties, serum bone alkaline phosphatase (b-ALP), serum bone γ-carboxyglutamic acid-containing protein (BGP), serum bone morphogenetic protein-2 (BMP-2), serum tartrate-resistant acid phosphatase (TRACP) and serum carboxy terminal cross linked telopeptide of typeⅠcollagen (ICTP), bone mineral content (BMC) and bone structured histomorphometry. The protein and mRNA expression of TGF-β1, Smad2, Smad3, Smad4 and Smad7 were detected by Western blotting (WB) and quantitative real time polymerase chain reaction (qRT-PCR), respectively.
  
  
  RESULTS
  Administration of combined Herba Epimedii and Fructus Ligustri Lucidi decoction and combined active fractions could significantly prevent GC-induced bone loss by increasing the contents of serum b-ALP, BGP and BMP-2 as the markers of bone formation, reducing the serum TRACP and ICTP contents to inhibit bone resorption and enhancing BMC. They could also attenuate biomechanical properties and BMD reduction, deterioration of trabecular architecture in MO rats. The mRNA and protein expressions of TGF-β1, smad2, smad3 and smad4 were up-regulated, and the mRNA and protein expression of Smad7 was down-regulated following combined Herba Epimedii and Fructus Ligustri Lucidi treatment.
  
  
  CONCLUSIONS
  Combination of Herba Epimedii and Fructus Ligustri Lucidi exhibited protective effects on promoting bone formation and precluding bone resorption. The underlying mechanism may be attributed to its regulations on TGF-β1/Smads pathway. The substance bases of the combined herbs on anti-osteoporosis were total flavonoids of Herba Epimedii, total iridoids and flavonoids of Fructus Ligustri Lucidi.
authors.affiliation: Beijing Key Lab of TCM Collateral Disease Theory Research, School of Traditional Chinese Medicine, Capital Medical University, No.10 Xitoutiao, Youanmenwai, Fengtai District, Beijing 100069, China.
authors.forename: Yan
authors.initials: Y
authors.lastname: Yang
authors.affiliation: Beijing Key Lab of TCM Collateral Disease Theory Research, School of Traditional Chinese Medicine, Capital Medical University, No.10 Xitoutiao, Youanmenwai, Fengtai District, Beijing 100069, China.
authors.forename: Honglei
authors.initials: H
authors.lastname: Nian
authors.affiliation: Beijing Key Lab of TCM Collateral Disease Theory Research, School of Traditional Chinese Medicine, Capital Medical University, No.10 Xitoutiao, Youanmenwai, Fengtai District, Beijing 100069, China.
authors.forename: Xiufeng
authors.initials: X
authors.lastname: Tang
authors.affiliation: Beijing Key Lab of TCM Collateral Disease Theory Research, School of Traditional Chinese Medicine, Capital Medical University, No.10 Xitoutiao, Youanmenwai, Fengtai District, Beijing 100069, China. Electronic address: wxj0517@sina.com.
authors.forename: Xiujuan
authors.initials: X
authors.lastname: Wang
authors.affiliation: Beijing Key Lab of TCM Collateral Disease Theory Research, School of Traditional Chinese Medicine, Capital Medical University, No.10 Xitoutiao, Youanmenwai, Fengtai District, Beijing 100069, China. Electronic address: gzblrh45@ccmu.edu.cn.
authors.forename: Renhui
authors.initials: R
authors.lastname: Liu
mesh_terms: D000818:Animals; D015519:Bone Density / Q000187:drug effects; D016723:Bone Remodeling / Q000187:drug effects; D001842:Bone and Bones / Q000187:drug effects / Q000378:metabolism; D004365:Drugs, Chinese Herbal / Q000494:pharmacology* / Q000627:therapeutic use*; D034181:Epimedium; D005938:Glucocorticoids; D031659:Ligustrum; D008297:Male; D010024:Osteoporosis / Q000139:chemically induced / Q000188:drug therapy* / Q000378:metabolism; D008517:Phytotherapy; D012333:RNA, Messenger / Q000378:metabolism; D017207:Rats, Sprague-Dawley; D051785:Smad Proteins / Q000235:genetics / Q000378:metabolism; D053773:Transforming Growth Factor beta1 / Q000235:genetics / Q000378:metabolism
chemical_list: D004365:Drugs, Chinese Herbal; D005938:Glucocorticoids; D012333:RNA, Messenger; D051785:Smad Proteins; D053773:Transforming Growth Factor beta1
keywords: Active fractions; Bone formation; Bone resorption; Glucocorticoid-induced osteoporosis; Herba Epimedii/Fructus Ligustri Lucidi; Icariin (PubChem CID: 5318997); Oleanolic Acid (PubChem CID: 10494); Rutin (PubChem CID: 5280805); TGF-β1/Smads