Hesperetin stimulates differentiation of primary rat osteoblasts involving the BMP signalling pathway

J Nutr Biochem. 2010 May;21(5):424-31. doi: 10.1016/j.jnutbio.2009.01.017. Epub 2009 May 7.


Hesperidin found in citrus fruits has been reported to be a promising bioactive compound for maintaining an optimal bone status in ovariectomized rodent models. In this study, we examined the capacity of hesperetin (Hp) to affect the proliferation, differentiation and mineralization of rodent primary osteoblasts. Then, the impact of Hp on signalling pathways known to be implicated in bone formation was explored. We exposed osteoblasts to physiological concentrations of 1 microM Hp (Hp1) and 10 microM Hp (Hp10). Neither proliferation nor mineralization was affected by Hp at either dose during 19 days of exposure. Hp at both doses enhanced differentiation by significantly increasing alkaline phosphatase (ALP) activity from Day 14 of exposure (Day 19: Hp1: +9%, Hp10: +14.8% vs. control; P<.05). However, Hp did not induce an obvious formation of calcium nodules. The effect of Hp10 on ALP was inhibited by addition of noggin protein, suggesting a possible action of this flavanone through the bone morphogenetic protein (BMP) pathway. Indeed, Hp10 significantly induced (1.2- to 1.4-fold) mRNA expression of genes involved in this signalling pathway (i.e., BMP2, BMP4, Runx2 and Osterix) after 48 h of exposure. This was strengthened by enhanced phosphorylation of the complex Smad1/5/8. Osteocalcin mRNA level was up-regulated by Hp only at 10 microM (2.2 fold vs. control). The same dose of Hp significantly decreased osteopontin (OPN) protein level (50% vs. control) after 14 days of culture. Our findings suggest that Hp may regulate osteoblast differentiation through BMP signalling and may influence the mineralization process by modulating OPN expression.

MeSH terms

  • Alkaline Phosphatase / metabolism
  • Animals
  • Bone Morphogenetic Proteins / genetics
  • Bone Morphogenetic Proteins / metabolism*
  • Carrier Proteins / metabolism
  • Cell Differentiation / drug effects*
  • Cell Proliferation / drug effects
  • Cells, Cultured
  • Core Binding Factor Alpha 1 Subunit / genetics
  • Core Binding Factor Alpha 1 Subunit / metabolism
  • Dose-Response Relationship, Drug
  • Gene Expression Regulation / drug effects
  • Hesperidin / antagonists & inhibitors
  • Hesperidin / pharmacokinetics
  • Hesperidin / pharmacology*
  • Minerals / metabolism
  • Osteoblasts / cytology
  • Osteoblasts / drug effects*
  • Osteocalcin / genetics
  • Osteocalcin / metabolism
  • Osteopontin / genetics
  • Osteopontin / metabolism
  • Phosphorylation / drug effects
  • RNA, Messenger / metabolism
  • Rats
  • Rats, Wistar
  • Signal Transduction / drug effects*
  • Smad Proteins / genetics
  • Smad Proteins / metabolism
  • Time Factors
  • Transcription Factors / genetics
  • Transcription Factors / metabolism


  • Bone Morphogenetic Proteins
  • Carrier Proteins
  • Core Binding Factor Alpha 1 Subunit
  • Minerals
  • RNA, Messenger
  • Runx2 protein, rat
  • Smad Proteins
  • Sp7 protein, rat
  • Spp1 protein, rat
  • Transcription Factors
  • Osteocalcin
  • Osteopontin
  • noggin protein
  • Hesperidin
  • Alkaline Phosphatase
  • hesperetin