Involvement of the purinergic P2X7 receptor in the formation of multinucleated giant cells

Multinucleated giant cells (MGC), a hallmark of chronic inflammatory reactions, remain an enigma of cell biology. There is evidence implicating the purinergic P2X(7) receptor in the fusion process leading to MGC. To investigate this, we used HEK 293 cells stably transfected with either 1) the full-length rat P2X(7) receptor (P2X(7) cells), 2) a rat P2X(7) receptor lacking the C-terminal domain (P2X(7)TC), or 3) a mock vector, and rat alveolar macrophages (MA) expressing the native receptor. P2X(7) cells cultured in serum-free medium formed increased numbers of MGC and displayed a higher fusion index compared with mock transfectants. Stimulation of P2X(7) pore-forming activity in P2X(7) cells by polymyxin B (PMB) further increased significantly the formation of MGC. Conversely, blockers of P2X-receptors including oxidized ATP, brilliant blue G, and pyridoxal phosphate-6-azophenyl-2'-4'-disulfonic acid inhibited significantly MGC formation in both unstimulated and PMB-stimulated P2X(7)-transfected cells. In contrast, cells transfected with the truncated P2X(7)TC were devoid of pore-forming activity, did not respond to PMB stimulation, and failed to form enhanced numbers of MGC, thus behaving as mock transfectants. As found for P2X(7)-transfected cells, PMB also potentiated dose-dependently the formation of multinucleated MA by rat alveolar MA. Pretreatment with oxidized ATP abrogated the PMB stimulatory effects. Together, these data demonstrate unequivocally the participation of P2X(7) receptor in the process of MGC formation. Our study also provides evidence suggesting that stimulation of the P2X(7) receptor pathway in MA may mediate increased formation of MGC during chronic inflammatory reactions.