A role for P2X₇ in microglial proliferation

Abstract

Microglia, glial cells with an immunocompetent role in the CNS, react to stimuli from the surrounding environment with alterations of their phenotypic response. Amongst other activating signals, the endotoxin lipopolysaccharide (LPS) is widely used as a tool to mimic bacterial infection in the CNS. LPS-activated microglia undergo dramatic changes in cell morphology/activity; in particular, they stop proliferating and differentiate from resting to effector cells. Activated microglia also show modifications of purinoreceptor signalling with a significant decrease in P2X 7 expression. In this study, we demonstrate that the down-regulation of the P2X 7 receptor in activated microglia may play an important role in the antiproliferative effect of LPS. Indeed, chronic blockade of the P2X 7 receptor by antagonists (oxidized ATP, KN62 and Brilliant Blue G), or treatment with the ATP-hydrolase apyrase, severely decreases microglial proliferation, down-regulation of P2X 7 receptor expression by small RNA interference (siRNA) decreases cell proliferation, and the proliferation of P2X 7 -deficient N9 clones and primary microglia, in which P2X 7 expression is down-regulated by siRNA, is unaffected by either LPS or P2X 7 antagonists. Furthermore, flow cytometric analysis indicates that exposure to oxidized ATP or treatment with LPS reversibly decreases cell cycle progression, without increasing the percentage of apoptotic cells. Overall, our data show that the P2X 7 receptor plays an important role in controlling microglial proliferation by supporting cell cycle progression. The P2X 7 receptor is an ATP-gated ion channel that has the unusual property of losing its ion selectivity and undergoing dilation to form a non-selective pore during prolonged ATP exposure. Because of this property, the P2X 7 receptor causes massive calcium entry, and this is often linked to an apoptotic fate of the cell (Di Virgilio 1995;Ferrari et al. 1997a). However, recent data have shown that tonic activation of this receptor mediates a proliferative rather than a death signal. The activation of transfected P2X 7 receptors by endogenously released ATP has been shown to enhance cell proliferation in human lymphoblastoid cells (Baricordi et al. 1999). The mechanism by which P2X 7 receptor stimulates cell growth has been recently linked to an increase in cellular energy stores (Adinolfi et al. 2005). The P2X 7 receptor is highly expressed in microglia, the resident cells of the brain which contribute to innate immunity. Microglial cells are normally in a resting state but, during infection or after traumatic stimuli, they differentiate into effector cells and acquire cytotoxic/phagocytic features to fight the pathogenic stimulus (Miyake 2004). Microglial activation is a complex process which proceeds through several steps and involves multiple factors, including cytokines and a wide variety of bacterial components (Norenberg et al. 1997 Abbreviations used: BAPTA-AM, acetoxymethyl-1,2-bis(2-aminophenoxy)ethane-N,N,N¢,N¢-tetraacetic acid; BzATP, 2...

Microglial Activation in Stroke: Therapeutic Targets

“…62 The P2X 7 receptors are expressed by resting microglia, and this expression is upregulated after brain injury. 63 Activation of P2X 7 receptors triggers microglia proliferation, 64,65 superoxide production, 66 release of interleukin 1␤ (IL-1␤), 67,68 and release of TNF␣. 64,69 However, microglia also exhibit reduced phagocytosis during P2X 7 receptor stimulation.…”

Physiological and pathological functions of P2X7 receptor in the spinal cord

“…For example, Morioka et al [75] have recently shown that p2x7 stimulation in spinal microglia produces downregulation of glutamate transport. Bianco et al [76] have also reported the involvement of the P2X7R in modulation of the proliferation capability of microglia adding complexity to the mechanisms by which P2X7R can be involved in the microglial response to injury.…”

P2X7: a growth-promoting receptor—implications for cancer

“…It is known that microglia, under the effect of LPS, stop proliferating and differentiate into an activated phenotype. Verderio and coworkers showed that inhibition of proliferation of mouse microglia by LPS is mediated by down-modulation of the P2X 7 receptor [8]. In the same year, Raffaghello et al [9] reported that P2X 7 receptor expression sustained growth of human neuroblastoma cells.…”

“…In 2006, Verderio and co-workers provided an important and independent support to the growth-promoting activity of P2X 7 by showing that proliferation of microglia cell clones and primary microglia was modulated by this receptor [8]. Microglia are resident cells of the brain that participate in reaction to traumas and in defence against foreign microorganisms.…”