The two-hit hypothesis for neuroinflammation: role of exogenous ATP in modulating inflammation in the brain (original) (raw)
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Prostaglandins, Leukotrienes and Essential Fatty Acids, 2003
Upregulation and activation of phospholipases A 2 (PLA 2 ) and cyclooxygenases (COX) leading to prostaglandin E 2 (PGE 2 ) production have been implicated in a number of neurodegenerative diseases. In this study, we investigated PGE 2 production in primary rat astrocytes in response to agents that activate PLA 2 including pro-inflammatory cytokines (IL-1b; TNFa and IFNg), the P2 nucleotide receptor agonist ATP, and oxidants (H 2 O 2 and menadione). Exposure of astrocytes to cytokines resulted in a timedependent increase in PGE 2 production that was marked by increased expression of secretory sPLA 2 and COX-2, but not COX-1 and cytosolic cPLA 2 . Although astrocytes responded to ATP or phorbol ester (PMA) with increased cPLA 2 phosphorylation and arachidonic acid release, ATP or PMA only caused a small increase in levels of PGE 2 . However, when astrocytes were first treated with cytokines, further exposure to ATP or PMA, but not H 2 O 2 or menadione, markedly increased PGE 2 production. These results suggest that ATP release during neuronal excitation or injury can enhance the inflammatory effects of cytokines on PGE 2 production and may contribute to chronic inflammation seen in Alzheimer's disease. r
PLoS ONE, 2011
Cyclooxygenases (COX) are prostanoid synthesizing enzymes constitutively expressed in the brain that contribute to excitotoxic neuronal cell death. While the neurotoxic role of COX-2 is well established and has been linked to prostaglandin E 2 synthesis, the role of COX-1 is not clearly understood. In a model of N-Methyl-D-aspartic acid (NMDA) induced excitotoxicity in the mouse cerebral cortex we found a distinctive temporal profile of COX-1 and COX-2 activation where COX-1, located in microglia, is responsible for the early phase of prostaglandin E 2 synthesis (10 minutes after NMDA), while both COX-1 and COX-2 contribute to the second phase (3-24 hours after NMDA). Microglial COX-1 is strongly activated by ATP but not excitatory neurotransmitters or the Toll-like receptor 4 ligand bacterial lipopolysaccharide. ATP induced microglial COX-1 dependent prostaglandin E 2 synthesis is dependent on P2X7 receptors, extracellular Ca 2+ and cytoplasmic phospholipase A2. NMDA receptor activation induces ATP release from cultured neurons leading to microglial P2X7 receptor activation and COX-1 dependent prostaglandin E 2 synthesis in mixed microglial-neuronal cultures. Pharmacological inhibition of COX-1 has no effect on the cortical lesion produced by NMDA, but counteracts the neuroprotection exerted by inhibition of COX-2 or observed in mice lacking the prostaglandin E 2 receptor type 1. Similarly, the neuroprotection exerted by the prostaglandin E 2 receptor type 2 agonist butaprost is not observed after COX-1 inhibition. P2X7 receptors contribute to NMDA induced prostaglandin E 2 production in vivo and blockage of P2X7 receptors reverses the neuroprotection offered by COX-2 inhibition. These findings suggest that purinergic signaling in microglia triggered by neuronal ATP modulates excitotoxic cortical lesion by regulating COX-1 dependent prostanoid production and unveil a previously unrecognized protective role of microglial COX-1 in excitotoxic brain injury.
Journal of Neurochemistry, 2003
During inflammatory or degenerative processes microglial cells are likely to be exposed to activating agents that persist in brain parenchyma for prolonged periods. As our knowledge on microglial activation is largely based on in vitro studies in which microglial cultures are activated by a single administration of pro-inflammatory stimuli, we investigated the effects of repeated endotoxin (LPS) challenges on microglial functional state. Primary rat microglial cultures were subjected to one, two or three consecutive LPS-stimulation and the production of tumor necrosis factor-a (TNF-a), nitric oxide (NO), prostaglandin E 2 (PGE 2 ) and 15-deoxy-D 12,14 -PGJ 2 (15d-PGJ 2 ) measured. The ability of microglial cells to produce NO, TNF-a and 15d-PGJ 2 upon the first LPS challenge rapidly declined after the second and the third stimulations, whereas PGE 2 synthesis remained constantly elevated.
2010
Background: Brain inflammation is accompanied by brain injury. However, it is controversial whether inflammatory responses are harmful or beneficial to neurons. Because many studies have been performed using cultured microglia and neurons, it has not been possible to assess the influence of multiple cell types and diverse factors that dynamically and continuously change in vivo. Furthermore, behavior of microglia and other inflammatory cells could have been overlooked since most studies have focused on neuronal death. Therefore, it is essential to analyze the precise roles of microglia and brain inflammation in the injured brain, and determine their contribution to neuronal damage in vivo from the onset of injury. Methods and Findings: Acute neuronal damage was induced by stereotaxic injection of ATP into the substantia nigra pars compacta (SNpc) and the cortex of the rat brain. Inflammatory responses and their effects on neuronal damage were investigated by immunohistochemistry, electron microscopy, quantitative RT-PCR, and stereological counting, etc. ATP acutely caused death of microglia as well as neurons in a similar area within 3 h. We defined as the core region the area where both TH + and Iba-1 + cells acutely died, and as the penumbra the area surrounding the core where Iba-1 + cells showed activated morphology. In the penumbra region, morphologically activated microglia arranged around the injury sites. Monocytes filled the damaged core after neurons and microglia died. Interestingly, neither activated microglia nor monocytes expressed iNOS, a major neurotoxic inflammatory mediator. Monocytes rather expressed CD68, a marker of phagocytic activity. Importantly, the total number of dopaminergic neurons in the SNpc at 3 h (,80% of that in the contralateral side) did not decrease further at 7 d. Similarly, in the cortex, ATP-induced neuron-damage area detected at 3 h did not increase for up to 7 d. Conclusions: Different cellular components (microglia, astrocytes, monocytes, and neutrophils) and different factors (proinflammatory and neurotrophic) could be produced in inflammatory processes depending on the nature of the injury. The results in this study suggest that the inflammatory responses of microglia and monocytes in response to ATP-induced acute injury could not be neurotoxic.
Journal of Neuroinflammation
Background: Pharmaceutical treatment with probable anti-inflammatory substances that attack cells in various ways including receptors, ion channels, or transporter systems may slow down the progression of inflammatory conditions. Astrocytes and microglia are the most prominent target cells for inflammation in the central nervous system. Their responses upon inflammatory stimuli work through the NO/cyclic GMP/protein kinase G systems that can downregulate the ATP-induced Ca 2+ signaling, as well as G protein activities which alter Na + transporters including Na + /K +-ATPase pump activity, Toll-like receptor 4 (TLR4), glutamate-induced Ca 2+ signaling, and release of pro-inflammatory cytokines. The rationale for this project was to investigate a combination of pharmaceutical substances influencing the NO and the G i /G s activations of inflammatory reactive cells in order to make the cells return into a more physiological state. The ATP-evoked Ca 2+ signaling is important maybe due to increased ATP release and subsequent activation of purinergic receptors. A balance between intercellular Ca 2+ signaling through gap junctions and extracellular signaling mediated by extracellular ATP may be important for physiological function. Methods: Astrocytes in primary cultures were incubated with lipopolysaccharide in a physiological glucose concentration for 24 h to induce inflammatory reactivity. The probable anti-inflammatory substances sildenafil and 1α,25-Dihydroxyvitamin D3 together with endomorphin-1, naloxone, and levetiracetam, were used in the presence of high glucose concentration in the medium to restore the cells. Glutamate-, 5-HT-, and ATP-evoked intracellular Ca 2+ release, Na + /K +-ATPase expression, expression of inflammatory receptors, and release of tumor necrosis factor alpha were measured. Results: Sildenafil in ultralow concentration together with 1α,25-Dihydroxyvitamin D3 showed most prominent effects on the ATP-evoked intracellular Ca 2+ release. The μ-opioid agonist endomorphin-1, the μ-opioid antagonist naloxone in ultralow concentration, and the antiepileptic agent levetiracetam downregulated the glutamate-evoked intracellular Ca 2+ release and TLR4. The combination of the pharmaceutical substances in high glucose concentration downregulated the glutamate-and ATP-evoked Ca 2+ signaling and the TLR4 expression and upregulated the Na + /K +-ATPase pump.
Non-Steroidal Anti-Inflammatory Drugs and Brain Inflammation: Effects on Microglial Functions
Pharmaceuticals, 2010
The term NSAID refers to structurally diverse chemical compounds that share the ability to inhibit the activity of the prostaglandin (PG) biosynthetic enzymes, the cyclooxygenase (COX) isoforms 1 and 2. The suppression of PG synthesis at sites of inflammation has been regarded as primarily responsible for the beneficial properties of NSAIDs, but several COX-independent effects have been described in recent years. Epidemiological studies indicate that NSAIDs are neuroprotective, although the mechanisms underlying their beneficial effect remain largely unknown. Microglial cells play a major role in brain inflammation and are often viewed as major contributors to the neurodegeneration. Therefore, microglia represent a likely target for NSAIDs within the brain. In the present review, we focused on the direct effects of NSAIDs and selective COX-2 inhibitors on microglial functions and discuss the potential efficacy in controlling brain inflammation.
Distinct migratory and cytokine responses of human microglia and macrophages to ATP
Brain, Behavior, and Immunity, 2010
Microglia and hematogenous myeloid cells are prominent components of inflammatory central nervous system (CNS) lesions associated with tissue injury. To help define the basis for recruitment of such cells into lesions and their contribution to the disease process, we characterized the migratory and cytokine responses of human adult and fetal microglia in the presence of extracellular ATP comparing them to monocytes and macrophages. Adult microglia showed increased migration in response to low ATP concentrations (1-10 lM) whereas fetal microglia also migrated in response to higher ATP dosages (100-300 lM). The enhanced migration of microglia was reproduced with 2-MeSADP, a P2Y1/12/13 agonist. In contrast, the chemokine CCL2 did not promote migration of microglia, but promoted the migration of monocytes. Monocyte migration was also enhanced with low concentrations of ATP, whereas higher concentrations of ATP mediated an inhibitory effect. ATP had only an inhibitory effect on macrophages, which was not reproduced with hydrolysis products ADP or adenosine. ATP led to a decrease in LPS-induced pro-inflammatory cytokine release (TNFa, IL-6) in both microglia and macrophages without suppression of an anti-inflammatory response (IL-10). These in vitro based results suggest that ATP can selectively favor the recruitment of microglia rather than hematogenous myeloid cells while promoting an anti-inflammatory state in both hematogenous and resident myeloid cells of the CNS. Our results highlight the importance of environmental signals in shaping the properties of the innate immune response to injury in the CNS.
Glia, 2005
Microglia are the major cell type involved in neuroinflammatory events in brain diseases such as encephalitis, stroke, and neurodegenerative disorders, and contribute significantly to the release of prostaglandins (PGs) during neuronal insults. In this report, we studied the immediateearly intracellular signalling pathways in microglia, following bacterial lipopolysaccharide (LPS) stimulation, leading to the synthesis and release of PGE 2 . Here we show that LPS induces cyclooxygenase (COX) 2 by activating sphingomyelinases leading to the release of ceramides, which in turn, activate the p38 mitogen-activated protein kinases (MAPK), but not the p42/44 MAPK. We further show that exogenously added ceramide analogue (C 2 -ceramide) also induce PGE 2 synthesis through a p38 MAPK-dependent pathway. This potential nature of ceramides in activating microglia suggests that endogenously produced ceramides during neuronal apoptosis in ischemia or neurodegenerative diseases could also contribute to the amplification of neuroinflammatory events. In contrast to protein kinase C (PKC) and phosphocholine-specific phospholipase C (PC-PLC), which transcriptionally regulate LPS-induced COX-2 synthesis, inhibition of phospholipase A 2 (PLA 2 ) has no effect on COX-2 transcription, although it inhibits the release of PGE 2 . Transcriptional regulation of LPS-induced COX-2 by PKC is further proved by the ability of the PKC inhibitor, G€ o 6976, to inhibit LPS-induced 8-isoprostane synthesis, but not affecting LPS-induced COX-2 activity. Our data with 8-isoprostane also indicates that COX-2 plays a major role in ROS production in LPS-activated microglia. This detailed view of the intracellular signaling pathway in microglial activation and COX-2 expression opens a new therapeutic window in the search for new and more effective central anti-inflammatory agents. in Wiley InterScience (www.interscience.
Inflammation in acute CNS injury: a focus on the role of substance P
British journal of pharmacology, 2015
Recently, a number of reports have shown that neurogenic inflammation may play a role in the secondary injury response following acute injury to the central nervous system (CNS), including traumatic brain injury (TBI) and stroke. In particular substance P (SP) release appears to be critically involved. Specifically, expression of the neuropeptide SP is increased in acute CNS injury, with the magnitude of SP release being related to both the frequency and magnitude of the insult. SP release is associated with an increase in blood-brain barrier permeability and the development of vasogenic oedema as well as neuronal injury and worsened functional outcome. Moreover, inhibiting the actions of SP through use of a NK1 antagonist is highly beneficial in both focal and diffuse models of TBI, as well as in ischaemic stroke, with a therapeutic window of up to 12h. We propose that NK1 antagonists represent a novel therapeutic option for treatment of neurogenic inflammation following acute CNS ...
Journal of Neuroimmunology, 2004
To test whether extracellular ATP can play a role in the neuroimmunopathology of Alzheimer's disease (AD), we evaluated the capacity of the ATP-binding purinoreceptor, P2X7, to modulate cytokine secretion on cultured human macrophages and microglia pre-activated 24 h with the 42 amino acid h-amyloid peptide (Ah(1 -42)) or lipopolysaccharide. Thirty minutes of exposure to the selective P2X7 agonist 2V -3V -O-(4-benzoylbenzoyl)adenosine 5V -triphosphate (BzATP) resulted in the secretion of IL-1h after either Ah(1 -42) or LPS stimulation of human macrophages that was dependent on the concentration of the stimulus used to pre-activate the cells. Further tests on human microglia treated with BzATP (300 AM) resulted in a 1.5-and 3.5-fold enhancement of IL-1a and IL-1h secretion, respectively, from cells pre-activated by 10 AM Ah(1 -42) and a 1.6-and 3.9-fold enhancement of IL-1a and IL-1h secretion, respectively, from cells pre-activated by 1 Ag/ml LPS. BzATP induction of IL-1a and IL-1h secretion from microglia was completely reversed by pre-incubation of the cells with the P2X7 antagonist, adenosine 5V -triphosphate 2V ,3V -acyclic dialcohol (oxidized ATP). In contrast to its effects on IL-1a and IL-1h secretion, BzATP induced TNF-a after LPS stimulation, but not after stimulation with Ah(1 -42), induced IL-18 secretion regardless of whether microglia were pre-activated and attenuated IL-6 secretion after either LPS or Ah(1 -42) pre-activation. These results demonstrate that extracellular ATP can modulate Ah-induced cytokine secretion from human macrophages and microglia and thus may play a role in the neuroimmunopathology of AD. D