Accordingly, we concluded that glioma cells with abundant production of IL-33 grow rapidly; moreover, the interactions of multiple cytokines/chemokines induced by glioma cells may develop a microenvironment that facilitates microglia/macrophage infiltration and fosters glioma growth in the brain.

Chemokines can be produced by gliomas, which mediate the infiltration of microglia, a characteristic feature of glioma-associated neuropathogenesis. ATP that is released at a high level from glioma has been reported to play a regulatory role in chemokine production in cultured glioma cells. The objective of this study was to define the potential role of extracellular ATP in the regulation of macrophage inflammatory protein-1α (MIP-1α) and monocyte chemoattractant protein-1(MCP-1) expression in glioma-associated microglia/macrophages. The results showed that Iba1(+) and ED1(+) microglia existed in the tumor at 3 and 7 day after injection of C6 glioma cells into the rat cerebral cortex (dpi). ED1(+) microglia/macrophages or Iba1(+) microglia in the glioma were also colocalized to MIP-1α- and MCP-1-expressing cells. In vitro study indicated that treatment with ATP and BzATP (an agonist for ATP ionotropic receptor P2X₇R) caused an increase in the intracellular levels of microglial MIP-1α and MCP-1. By using an extracellular Ca(2+) chelator (EGTA) and P2X₇R antagonists, oxidized ATP (oxATP) and brilliant blue G (BBG), we demonstrated that BzATP-induced production of MIP-1α and MCP-1 levels was due to P2X₇R activation and Ca(2+) -dependent regulation. Coadministration of C6 glioma cells and oxATP into the rat cerebral cortex resulted in a reduction of MIP-1α- and MCP-1-expressing microglia/macrophages. We suggest, based on the results from in vivo and in vitro studies, that a massive amount of ATP molecules released in the glioma tumor site may act as the regulator with P2X₇R signaling that increases MIP-1α and MCP-1 expression in tumor-infiltrating microglia/macrophages.

Microglia, the CNS resident macrophages responsible for the clearance of degenerating cellular fragments, are essential to tissue remodeling and repair after CNS injury. ATP can be released in large amounts after CNS injury and may mediate microglial activity through the ionotropic P2X and the metabotropic P2Y receptors. This study indicates that exposure to a high concentration of ATP for 30 min rapidly induces changes of the microglial cytoskeleton, and significantly attenuates microglial phagocytosis. A pharmacological approach showed that ATP-induced inhibition of microglial phagocytotic activity was due to P2X 7 R activation, rather than that of P2YR. Activation of P2X 7 R by its agonist, 2¢-3¢-O-(4-benzoyl)benzoyl-ATP (BzATP), produced a Ca 2+ -independent reduction in microglial phagocytotic activity. In addition, the knockdown of P2X 7 R expression by lentiviral-mediated shRNA interference or the blockade of P2X 7 R activation by the specific antagonists, oxidized ATP (oxATP) and brilliant blue G, has efficiently restored the phagocytotic activity of ATP and BzATPtreated microglia. Our results reveal that P2X 7 R activation may induce the formation of a Ca 2+ -independent signaling complex, which results in the reduction of microglial phagocytosis. This suggests that exposure to ATP for a short-term period may cause insufficient clearance of tissue debris by microglia through P2X 7 R activation after CNS injury, and that blockade of this receptor may preserve the phagocytosis of microglia and facilitate CNS tissue repair. Keywords: ATP, BzATP, microglia, oxATP, P2X 7 R, phagocytosis J. Neurochem. and Burnstock 1998; Fields and Burnstock 2006). The activation of metabotropic P2YR subfamily containing P2Y 1,2,4,6,11,12,13,14 subtypes exerts a slower response to ATP via the induction of Ca 2+ release from IP 3 -sensitive stores and activation of intracellular signal cascades (Ralevic and Burnstock 1998;Fields and Burnstock 2006).Microglia express P2X 1,4,7 and P2Y 1,4,6,12 receptors (Inoue 2002(Inoue , 2006Fields and Burnstock 2006). ATP can stimulate microglia to produce iNOS and proinflammatory cytokines through its P2XR and P2YR (Ohtani et al. 2000;Inoue 2002Inoue , 2006Di Virgilio et al. 2009). High concentrations of ATP also induce glia to release plasminogen, TNF-a, and IL-1b via P2X 7 R by complex mechanisms that are dependent on Ca 2+ signaling and extracellular signal-regulated kinase (ERK)/p38 mitogen-activated protein kinase (MAPK) cascade (Hide et al. 2000;Sanz and Di Virgilio 2000;Inoue 2002). Microglia isolated from P2Y 12 R-deficient mice are unable to produce directional branch extension toward the area of cortical damage, indicating that P2Y 12 R is critical for microglial chemotaxis to the CNS lesion site (Haynes et al. 2006). In vivo studies have indicated that P2X 4 R expression in microglia was strikingly increased in the spinal cord after nerve injury, which contributed to neuropathic pain and pathogenesis (Tsuda et al. 2003;Inoue 2006). These findings point to the intricate ...

Astrocytes, the most abundant glial cell population in the central nervous system (CNS), play physiological roles in neuronal activities. Oxidative insult induced by the injury to the CNS causes neural cell death through extrinsic and intrinsic pathways. This study reports that reactive oxygen species (ROS) generated by exposure to the strong oxidizing agent, hexavalent chromium (Cr(VI)) as a chemical-induced oxidative stress model, caused astrocytes to undergo an apoptosis-like cell death through a caspase-3-independent mechanism. Although activating protein-1 (AP-1) and NF-kappaB were activated in Cr(VI)-primed astrocytes, the inhibition of their activity failed to increase astrocytic cell survival. The results further indicated that the reduction in mitochondrial membrane potential (MMP) was accompanied by an increase in the levels of ROS in Cr(VI)-primed astrocytes. Moreover, pretreatment of astrocytes with N-acetylcysteine (NAC), the potent ROS scavenger, attenuated ROS production and MMP loss in Cr(VI)-primed astrocytes, and significantly increased the survival of astrocytes, implying that the elevated ROS disrupted the mitochondrial function to result in the reduction of astrocytic cell viability. In addition, the nuclear expression of apoptosis-inducing factor (AIF) and endonuclease G (EndoG) was observed in Cr(VI)-primed astrocytes. Taken together, evidence shows that astrocytic cell death occurs by ROS-induced oxidative insult through a caspase-3-independent apoptotic mechanism involving the loss of MMP and an increase in the nuclear levels of mitochondrial pro-apoptosis proteins (AIF/EndoG). This mitochondria-mediated but caspase-3-independent apoptotic pathway may be involved in oxidative stress-induced astrocytic cell death in the injured CNS.