title: P2X7-induced zeiosis promotes osteogenic differentiation and mineralization of postmenopausal bone marrow-derived mesenchymal stem cells.
abstract: Polymorphisms of the P2X7 receptor have been associated with increased risk of fractures in postmenopausal women. Although both osteoblasts and osteoclasts express P2X7 receptors, their function in osteogenesis remains controversial. Here, we investigated the role of the P2X7 receptor on osteogenic differentiation and mineralization of bone marrow mesenchymal stem cell (BMSC) cultures from postmenopausal women (age 71±3 yr, n=18). We focused on the mechanisms related to intracellular [Ca(2+)]i oscillations and plasma membrane-dynamics. ATP, and the P2X7 agonist BzATP (100 μM), increased [Ca(2+)]i in parallel to the formation of membrane pores permeable to TO-PRO-3 dye uptake. ATP and BzATP elicited reversible membrane blebs (zeiosis) in 38 ± 1 and 70 ± 1% of the cells, respectively. P2X7-induced zeiosis was Ca(2+) independent, but involved phospholipase C, protein kinase C, and Rho-kinase activation. BzATP (100 μM) progressively increased the expression of Runx-2 and Osterix transcription factors by 452 and 226% (at d 21), respectively, alkaline phosphatase activity by 88% (at d 28), and mineralization by 329% (at d 43) of BMSC cultures in a Rho-kinase-dependent manner. In summary, reversible plasma membrane zeiosis involving cytoskeleton rearrangements due to activation of the P2X7-Rho-kinase axis promotes osteogenic differentiation and mineralization of BMSCs, thus providing new therapeutic targets for postmenopausal bone loss.
authors.affiliation: Laboratório de Farmacologia e Neurobiologia and Center for Drug Discovery and Innovative Medicines, Instituto de Ciências Biomédicas Abel Salazar-Universidade do Porto (ICBAS-UP), Porto, Portugal; and.
authors.forename: José Bernardo
authors.initials: JB
authors.lastname: Noronha-Matos
authors.affiliation: Laboratório de Farmacologia e Neurobiologia and Center for Drug Discovery and Innovative Medicines, Instituto de Ciências Biomédicas Abel Salazar-Universidade do Porto (ICBAS-UP), Porto, Portugal; and.
authors.forename: João
authors.initials: J
authors.lastname: Coimbra
authors.affiliation: Laboratório de Farmacologia e Neurobiologia and Center for Drug Discovery and Innovative Medicines, Instituto de Ciências Biomédicas Abel Salazar-Universidade do Porto (ICBAS-UP), Porto, Portugal; and.
authors.forename: Ana
authors.initials: A
authors.lastname: Sá-e-Sousa
authors.affiliation: Serviço de Ortopedia e Traumatologia, Centro Hospitalar de Vila Nova de Gaia/Espinho, Vila Nova de Gaia, Portugal.
authors.forename: Rui
authors.initials: R
authors.lastname: Rocha
authors.affiliation: Serviço de Ortopedia e Traumatologia, Centro Hospitalar de Vila Nova de Gaia/Espinho, Vila Nova de Gaia, Portugal.
authors.forename: José
authors.initials: J
authors.lastname: Marinhas
authors.affiliation: Serviço de Ortopedia e Traumatologia, Centro Hospitalar de Vila Nova de Gaia/Espinho, Vila Nova de Gaia, Portugal.
authors.forename: Rolando
authors.initials: R
authors.lastname: Freitas
authors.affiliation: Laboratório de Farmacologia e Neurobiologia and Center for Drug Discovery and Innovative Medicines, Instituto de Ciências Biomédicas Abel Salazar-Universidade do Porto (ICBAS-UP), Porto, Portugal; and.
authors.forename: Sónia
authors.initials: S
authors.lastname: Guerra-Gomes
authors.affiliation: Laboratório de Farmacologia e Neurobiologia and Center for Drug Discovery and Innovative Medicines, Instituto de Ciências Biomédicas Abel Salazar-Universidade do Porto (ICBAS-UP), Porto, Portugal; and.
authors.forename: Fátima
authors.initials: F
authors.lastname: Ferreirinha
authors.affiliation: Laboratório de Farmacologia e Neurobiologia and Departamento de Química, Unit for Multidisciplinary Research in Biomedicine (UMIB), and Center for Drug Discovery and Innovative Medicines, Instituto de Ciências Biomédicas Abel Salazar-Universidade do Porto (ICBAS-UP), Porto, Portugal; and.
authors.forename: Maria Adelina
authors.initials: MA
authors.lastname: Costa
authors.affiliation: Laboratório de Farmacologia e Neurobiologia and Center for Drug Discovery and Innovative Medicines, Instituto de Ciências Biomédicas Abel Salazar-Universidade do Porto (ICBAS-UP), Porto, Portugal; and farmacol@icbas.up.pt.
authors.forename: Paulo
authors.initials: P
authors.lastname: Correia-de-Sá
mesh_terms: D000368:Aged; D001842:Bone and Bones / Q000166:cytology*; D002113:Calcification, Physiologic / Q000502:physiology*; D002118:Calcium / Q000378:metabolism; D002454:Cell Differentiation / Q000502:physiology*; D004789:Enzyme Activation; D005260:Female; D006801:Humans; D059630:Mesenchymal Stem Cells / Q000166:cytology* / Q000201:enzymology; D017698:Postmenopause; D011493:Protein Kinase C / Q000378:metabolism; D058486:Receptors, Purinergic P2X7 / Q000502:physiology*; D010738:Type C Phospholipases / Q000378:metabolism; D054460:rho-Associated Kinases / Q000378:metabolism
chemical_list: D058486:Receptors, Purinergic P2X7; D054460:rho-Associated Kinases; D011493:Protein Kinase C; D010738:Type C Phospholipases; D002118:Calcium
keywords: P2 purinoceptors; Rho-kinase; bone-nodule formation; membrane blebbing; osteogenesis