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Lactoferrin Promotes Osteogenesis through TGF-beta Receptor II Binding in Osteoblasts and Activation of Canonical TGF-beta Signaling in MC3T3-E1 Cells and C57BL/6J Mice

TitleLactoferrin Promotes Osteogenesis through TGF-beta Receptor II Binding in Osteoblasts and Activation of Canonical TGF-beta Signaling in MC3T3-E1 Cells and C57BL/6J Mice
Publication TypeJournal Article
Year of Publication2018
AuthorsLi, Y, Wang, J, Ren, F, Zhang, W, Zhang, H, Zhao, L, Zhang, M, Cui, W, Wang, X, Guo, H
JournalJ Nutr
Volume148
Pagination1285-1292
Date PublishedAug 1
ISBN Number0022-3166
Accession Number29931165
Abstract

Background: Lactoferrin (LF), as a major functional protein in dairy products, is known to modulate bone anabolic effects. However, the underlying molecular mechanisms remain unclear; the receptor of LF in osteoblast differentiation has not been identified. Objective: The aims of the study were to 1) illuminate whether the receptor of LF in osteoblast differentiation is transforming growth factor-beta (TGF-beta) receptor (TbetaR) II and 2) determine whether the TGF-beta signaling pathway is activated by LF in promoting osteogenesis in vitro and in vivo, in addition to P38 and extracellular signal-regulated kinase (ERK) pathways. Methods: We utilized co-immunoprecipitation to detect any binding of LF to TbetaRII. Subsequently, the role of the TGF-beta signaling pathway involved in LF-induced osteoblast proliferation and differentiation was determined by inhibition of TbetaRI activity by inhibition and knockout of TbetaRII expression by small guide RNA (sgRNAs) in MC3T3-E1 cells. In addition, 4-wk-old male C57BL/6J mice were orally administered 100 mg LF/kg body weight for 16 wk, after which any activation of the TGF-beta signaling pathway in vivo was measured by Western blots. Results: LF was shown to directly interact with the TbetaRII protein and to activate the TGF-beta signaling pathway in MC3T3-E1 cells. Inhibition of TbetaRI activity and knockout TbetaRII expression both attenuated the stimulation of LF in osteoblast proliferation and differentiation by 30-50%. LF-induced activation of TGF-beta canonical signaling resulted in upregulation of osteogenic factors. Moreover, the expression of phosphorylated-drosophila mothers against decapentaplegic protein 2 (SMAD2) was increased by 1-fold after LF treatment in the femoral tissue of mice. Conclusions: This study provides evidence identifying TbetaRII as an LF receptor in LF-induced osteoblast differentiation. In addition, the TbetaRII-dependent TGF-beta canonical signaling pathways were proven to play an important role in mediating LF-induced osteogenesis both in MC3T3-E1 cells and in C57BL/6J mice.