Betaine promotes cell differentiation of human osteoblasts in primary culture

Isabella Villa, Pamela Senesi, Anna Montesano, Anita Ferraretto, Anita Ferraretto, Fernanda Vacante, Alice Spinello, Michela Bottani, Simona Bolamperti, Alessandro Rubinacci, Livio Luzi, Livio Luzi, Ileana Terruzzi

Research output: Contribution to journalArticle

8 Citations (Scopus)

Abstract

© 2017 The Author(s). Background: Betaine (BET), a component of many foods, is an essential osmolyte and a source of methyl groups; it also shows an antioxidant activity. Moreover, BET stimulates muscle differentiation via insulin like growth factor I (IGF-I). The processes of myogenesis and osteogenesis involve common mechanisms with skeletal muscle cells and osteoblasts sharing the same precursor. Therefore, we have hypothesized that BET might be effective on osteoblast cell differentiation. Methods: The effect of BET was tested in human osteoblasts (hObs) derived from trabecular bone samples obtained from waste material of orthopedic surgery. Cells were treated with 10 mM BET at 5, 15, 60 min and 3, 6 and 24 h. The possible effects of BET on hObs differentiation were evaluated by real time PCR, western blot and immunofluorescence analysis. Calcium imaging was used to monitor intracellular calcium changes. Results: Real time PCR results showed that BET stimulated significantly the expression of RUNX2, osterix, bone sialoprotein and osteopontin. Western blot and immunofluorescence confirmed BET stimulation of osteopontin protein synthesis. BET stimulated ERK signaling, key pathway involved in osteoblastogenesis and calcium signaling. BET induced a rise of intracellular calcium by means of the calcium ions influx from the extracellular milieu through the L-type calcium channels and CaMKII signaling activation. A significant rise in IGF-I mRNA at 3 and 6 h and a significant increase of IGF-I protein at 6 and 24 h after BET stimulus was detected. Furthermore, BET was able to increase significantly both SOD2 gene expression and protein content. Conclusions: Our study showed that three signaling pathways, i.e. cytosolic calcium influx, ERK activation and IGF-I production, are enhanced by BET in human osteoblasts. These pathways could have synergistic effects on osteogenic gene expression and protein synthesis, thus potentially leading to enhanced bone formation. Taken together, these results suggest that BET could be a promising nutraceutical therapeutic agent in the strategy to counteract the concomitant and interacting impact of sarcopenia and osteoporosis, i.e. the major determinants of senile frailty and related mortality.
Original languageEnglish
JournalJournal of Translational Medicine
Volume15
Issue number1
DOIs
Publication statusPublished - Jun 7 2017

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Betaine
Osteoblasts
Cell culture
Cell Differentiation
Calcium
Insulin-Like Growth Factor I
Osteopontin
Osteogenesis
Gene expression
Fluorescent Antibody Technique
Muscle
Real-Time Polymerase Chain Reaction
Bone
Proteins
Western Blotting
Chemical activation
Integrin-Binding Sialoprotein
Sarcopenia
Gene Expression
Calcium-Calmodulin-Dependent Protein Kinase Type 2

Keywords

  • Aging
  • Bone
  • Calcium signaling
  • IGF-I
  • Nutraceutical

Cite this

Betaine promotes cell differentiation of human osteoblasts in primary culture. / Villa, Isabella; Senesi, Pamela; Montesano, Anna; Ferraretto, Anita; Ferraretto, Anita; Vacante, Fernanda; Spinello, Alice; Bottani, Michela; Bolamperti, Simona; Rubinacci, Alessandro; Luzi, Livio; Luzi, Livio; Terruzzi, Ileana.

In: Journal of Translational Medicine, Vol. 15, No. 1, 07.06.2017.

Research output: Contribution to journalArticle

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abstract = "{\circledC} 2017 The Author(s). Background: Betaine (BET), a component of many foods, is an essential osmolyte and a source of methyl groups; it also shows an antioxidant activity. Moreover, BET stimulates muscle differentiation via insulin like growth factor I (IGF-I). The processes of myogenesis and osteogenesis involve common mechanisms with skeletal muscle cells and osteoblasts sharing the same precursor. Therefore, we have hypothesized that BET might be effective on osteoblast cell differentiation. Methods: The effect of BET was tested in human osteoblasts (hObs) derived from trabecular bone samples obtained from waste material of orthopedic surgery. Cells were treated with 10 mM BET at 5, 15, 60 min and 3, 6 and 24 h. The possible effects of BET on hObs differentiation were evaluated by real time PCR, western blot and immunofluorescence analysis. Calcium imaging was used to monitor intracellular calcium changes. Results: Real time PCR results showed that BET stimulated significantly the expression of RUNX2, osterix, bone sialoprotein and osteopontin. Western blot and immunofluorescence confirmed BET stimulation of osteopontin protein synthesis. BET stimulated ERK signaling, key pathway involved in osteoblastogenesis and calcium signaling. BET induced a rise of intracellular calcium by means of the calcium ions influx from the extracellular milieu through the L-type calcium channels and CaMKII signaling activation. A significant rise in IGF-I mRNA at 3 and 6 h and a significant increase of IGF-I protein at 6 and 24 h after BET stimulus was detected. Furthermore, BET was able to increase significantly both SOD2 gene expression and protein content. Conclusions: Our study showed that three signaling pathways, i.e. cytosolic calcium influx, ERK activation and IGF-I production, are enhanced by BET in human osteoblasts. These pathways could have synergistic effects on osteogenic gene expression and protein synthesis, thus potentially leading to enhanced bone formation. Taken together, these results suggest that BET could be a promising nutraceutical therapeutic agent in the strategy to counteract the concomitant and interacting impact of sarcopenia and osteoporosis, i.e. the major determinants of senile frailty and related mortality.",
keywords = "Aging, Bone, Calcium signaling, IGF-I, Nutraceutical",
author = "Isabella Villa and Pamela Senesi and Anna Montesano and Anita Ferraretto and Anita Ferraretto and Fernanda Vacante and Alice Spinello and Michela Bottani and Simona Bolamperti and Alessandro Rubinacci and Livio Luzi and Livio Luzi and Ileana Terruzzi",
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T1 - Betaine promotes cell differentiation of human osteoblasts in primary culture

AU - Villa, Isabella

AU - Senesi, Pamela

AU - Montesano, Anna

AU - Ferraretto, Anita

AU - Ferraretto, Anita

AU - Vacante, Fernanda

AU - Spinello, Alice

AU - Bottani, Michela

AU - Bolamperti, Simona

AU - Rubinacci, Alessandro

AU - Luzi, Livio

AU - Luzi, Livio

AU - Terruzzi, Ileana

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AB - © 2017 The Author(s). Background: Betaine (BET), a component of many foods, is an essential osmolyte and a source of methyl groups; it also shows an antioxidant activity. Moreover, BET stimulates muscle differentiation via insulin like growth factor I (IGF-I). The processes of myogenesis and osteogenesis involve common mechanisms with skeletal muscle cells and osteoblasts sharing the same precursor. Therefore, we have hypothesized that BET might be effective on osteoblast cell differentiation. Methods: The effect of BET was tested in human osteoblasts (hObs) derived from trabecular bone samples obtained from waste material of orthopedic surgery. Cells were treated with 10 mM BET at 5, 15, 60 min and 3, 6 and 24 h. The possible effects of BET on hObs differentiation were evaluated by real time PCR, western blot and immunofluorescence analysis. Calcium imaging was used to monitor intracellular calcium changes. Results: Real time PCR results showed that BET stimulated significantly the expression of RUNX2, osterix, bone sialoprotein and osteopontin. Western blot and immunofluorescence confirmed BET stimulation of osteopontin protein synthesis. BET stimulated ERK signaling, key pathway involved in osteoblastogenesis and calcium signaling. BET induced a rise of intracellular calcium by means of the calcium ions influx from the extracellular milieu through the L-type calcium channels and CaMKII signaling activation. A significant rise in IGF-I mRNA at 3 and 6 h and a significant increase of IGF-I protein at 6 and 24 h after BET stimulus was detected. Furthermore, BET was able to increase significantly both SOD2 gene expression and protein content. Conclusions: Our study showed that three signaling pathways, i.e. cytosolic calcium influx, ERK activation and IGF-I production, are enhanced by BET in human osteoblasts. These pathways could have synergistic effects on osteogenic gene expression and protein synthesis, thus potentially leading to enhanced bone formation. Taken together, these results suggest that BET could be a promising nutraceutical therapeutic agent in the strategy to counteract the concomitant and interacting impact of sarcopenia and osteoporosis, i.e. the major determinants of senile frailty and related mortality.

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