Expression of a P2X(7) receptor by a subpopulation of human osteoblasts

There is now conclusive evidence that extracellular nucleotides acting via tell surface P2 receptors are important local modulators of bone cell funct ion. Multiple subtypes of P2 receptors have been localized to bone, where t heir activation modulates multiple processes including osteoblast prolifera tion, osteoblast-mediated bone formation, and osteoclast formation and reso rptive capacity, Locally released nucleotides also have been shown to sensi tize surrounding cells to the action of systemic factors such as parathyroi d hormone (PTH). In nonskeletal tissue recent attention has focused on one particular P2 receptor, the P2X(7) receptor (previously termed P2Z), and it s ability to form nonselective aqueous pores in the plasma membrane on prol onged stimulation. Expression of this receptor originally was thought to be restricted to cells of hemopoietic origin, in which it has been implicated in cell fusion, apoptosis, and release of proinflammatory cytokines, Howev er, recent reports have indicated expression of this receptor in cells of s tromal origin. In this study, we investigated the expression of the P2X(7) receptor in two human osteosarcoma cell lines, as well as several populatio ns of primary human bone-derived cells (HBDCs) at the levels of messenger R NA (mRNA) and protein. We found that there is a subpopulation of osteoblast s that expresses the P2X(7) receptor and that these receptors are functiona l as assessed by monitoring ethidium bromide uptake following pore formatio n. Inhibition of delayed lactate dehydrogenase (LDH) release in response to the specific agonist 2 ' ,3 '-(4-benzoyl)-benzoyl-adenosine triphosphate ( BzATP) by the nonspecific P2X receptor antagonist PPADS confirmed a recepto r-mediated event, After treatment with BzATP SaOS-2 cells exhibited dramati c morphological changes consistent with those observed after P2X(7)-mediate d apoptosis in hemopoietic cells. Dual staining with terminal deoxynucleoti dyl transferase-mediated deoxyuridine triphosphate-biotin nick end labeling (TUNEL) and a P2X(7)-specific monoclonal antibody confirmed the induction of apoptosis in osteoblasts expressing the P2X(7) receptor, These data show for the first time the expression of functional P2X(7) receptors in a subp opulation of osteoblasts, activation of which can result in ATP-mediated ap optosis.