David J. Loane

Role of microglia in neurotrauma

SummaryMicroglia are the primary mediators of the immune defense system of the CNS and are integral to the subsequent inflammatory response. The role of microglia in the injured CNS is under scrutiny, as research has begun to fully explore how postinjury inflammation contributes to secondary damage and recovery of function. Whether microglia are good or bad is under debate, with strong support for a dual role or differential activation of microglia. Microglia release a number of factors that modulate secondary injury and recovery after injury, including pro- and anti-inflammatory cytokines, chemokines, nitric oxide, prostaglandins, growth factors, and Superoxide species. Here we review experimental work on the complex and varied responses of microglia in terms of both detrimental and beneficial effects. Addressed in addition are the effects of microglial activation in two examples of CNS injury: spinal cord and traumatic brain injury. Microglial activation is integral to the response of CNS tissue to injury. In that light, future research is needed to focus on clarifying the signals and mechanisms by which microglia can be guided to promote optimal functional recovery.

Neuroprotection for traumatic brain injury: translational challenges and emerging therapeutic strategies

Traumatic brain injury (TBI) causes secondary biochemical changes that contribute to subsequent tissue damage and associated neuronal cell death. Neuroprotective treatments that limit secondary tissue loss and/or improve behavioral outcome have been well established in multiple animal models of TBI. However, translation of such neuroprotective strategies to human injury have been disappointing, with the failure of more than thirty controlled clinical trials. Both conceptual issues and methodological differences between preclinical and clinical injury have undoubtedly contributed to these translational difficulties. More recently, changes in experimental approach, as well as altered clinical trial methodologies, have raised cautious optimism regarding the outcomes of future clinical trials. Here we critically review developing experimental neuroprotective strategies that show promise, and we propose criteria for improving the probability of successful clinical translation.

Neuroinflammation after traumatic brain injury: Opportunities for therapeutic intervention

Traumatic brain injury (TBI) remains one of the leading causes of mortality and morbidity worldwide, yet despite extensive efforts to develop neuroprotective therapies for this devastating disorder there have been no successful outcomes in human clinical trials to date. Following the primary mechanical insult TBI results in delayed secondary injury events due to neurochemical, metabolic and cellular changes that account for many of the neurological deficits observed after TBI. The development of secondary injury represents a window of opportunity for therapeutic intervention to prevent progressive tissue damage and loss of function after injury. To establish effective neuroprotective treatments for TBI it is essential to fully understand the complex cellular and molecular events that contribute to secondary injury. Neuroinflammation is well established as a key secondary injury mechanism after TBI, and it has been long considered to contribute to the damage sustained following brain injury. However, experimental and clinical research indicates that neuroinflammation after TBI can have both detrimental and beneficial effects, and these likely differ in the acute and delayed phases after injury. The key to developing future anti-inflammatory based neuroprotective treatments for TBI is to minimize the detrimental and neurotoxic effects of neuroinflammation while promoting the beneficial and neurotrophic effects, thereby creating optimal conditions for regeneration and repair after injury. This review outlines how post-traumatic neuroinflammation contributes to secondary injury after TBI, and discusses the complex and varied responses of the primary innate immune cells of the brain, microglia, to injury. In addition, emerging experimental anti-inflammatory and multipotential drug treatment strategies for TBI are discussed, as well as some of the challenges faced by the research community to translate promising neuroprotective drug treatments to the clinic.

Progressive Neurodegeneration After Experimental Brain Trauma: Association With Chronic Microglial Activation

Recent clinical studies indicate that traumatic brain injury (TBI) produces chronic and progressive neurodegenerative changes leading to late neurologic dysfunction, but little is known about the mechanisms underlying such changes. Microglial-mediated neuroinflammationis an important secondary injury mechanism after TBI. In human studies, microglial activation has been found to persist for many years after the initial brain trauma, particularly after moderate to severe TBI. In the present study, adult C57Bl/6 mice were subjected to single moderate-level controlled cortical impact and were followed up by longitudinal T2-weighted magnetic resonance imaging in combination with stereologic histologic assessment of lesion volume expansion, neuronal loss, and microglial activation for up to 1 year after TBI. Persistent microglial activation was observed in the injured cortex through 1 year after injury and was associated with progressive lesion expansion, hippocampal neurodegeneration, and loss of myelin. Notably, highly activated microglia that expressed major histocompatibility complex class II (CR3/43), CD68, and NADPH oxidase (NOX2) were detected at the margins of the expanding lesion at 1 year after injury; biochemical markers of neuroinflammation and oxidative stress were significantly elevated at this time point. These data support emerging clinical TBI findings and provide a mechanistic link between TBI-induced chronic microglial activation and progressive neurodegeneration.

Traumatic brain injury in aged animals increases lesion size and chronically alters microglial/macrophage classical and alternative activation states

Traumatic brain injury (TBI) causes chronic microglial activation that contributes to subsequent neurodegeneration, with clinical outcomes declining as a function of aging. Microglia/macrophages (MG/Mɸ) have multiple phenotypes, including a classically activated, proinflammatory (M1) state that might contribute to neurotoxicity, and an alternatively activated (M2) state that might promote repair. In this study we used gene expression, immunohistochemical, and stereological analyses to show that TBI in aged versus young mice caused larger lesions associated with an M1/M2 balance switch and increased numbers of reactive (bushy and hypertrophic) MG/Mɸ in the cortex, hippocampus, and thalamus. Chitinase3-like 3 (Ym1), an M2 phenotype marker, displayed heterogeneous expression after TBI with amoeboid-like Ym1-positive MG/Mɸ at the contusion site and ramified Ym1-positive MG/Mɸ at distant sites; this distribution was age-related. Aged-injured mice also showed increased MG/Mɸ expression of major histocompatibility complex II and NADPH oxidase, and reduced antioxidant enzyme expression which was associated with lesion size and neurodegeneration. Thus, altered relative M1/M2 activation and an nicotinamide adenine dinucleotide phosphate oxidase (NADPH oxidase)-mediated shift in redox state might contribute to worse outcomes observed in older TBI animals by creating a more proinflammatory M1 MG/Mɸ activation state.

Eicosapentaenoic acid confers neuroprotection in the amyloid-β challenged aged hippocampus

Among the changes that occur in the hippocampus with age, is a deficit in long-term potentiation (LTP). This impairment is associated with inflammatory changes, which are typified by increased concentration of the pro-inflammatory cytokine interleukin-1beta (IL-1beta). Activated microglia are the most likely cell source of IL-1beta, but data demonstrating an age-related increase in microglial activation is equivocal. Here we demonstrate that the age-related deficit in LTP is accompanied by increased expression of cell surface markers of activated microglia (major histocompatibility complex II and CD40) and increased IL-1beta production, and that these changes may be stimulated by interferon-gamma. Treatment of aged rats with eicosapentaenoic acid (EPA) attenuates these changes and we suggest that IL-4 mediates the action of EPA. We demonstrate that aged rats exhibit an exaggerated response to intracerebroventricular injection of beta-amyloid peptide 1-40 (Abeta). Thus Abeta inhibited LTP in aged, but not young, rats and induced a further increase in hippocampal IL-1beta concentration. Of particular significance is the demonstration that EPA protects the aged brain so that the increased vulnerability to Abeta is ameliorated in EPA-treated rats.

Microglia in the TBI brain: The good, the bad, and the dysregulated.

As the major cellular component of the innate immune system in the central nervous system (CNS) and the first line of defense whenever injury or disease occurs, microglia play a critical role in neuroinflammation following a traumatic brain injury (TBI). In the injured brain microglia can produce neuroprotective factors, clear cellular debris and orchestrate neurorestorative processes that are beneficial for neurological recovery after TBI. However, microglia can also become dysregulated and can produce high levels of pro-inflammatory and cytotoxic mediators that hinder CNS repair and contribute to neuronal dysfunction and cell death. The dual role of microglial activation in promoting beneficial and detrimental effects on neurons may be accounted for by their polarization state and functional responses after injury. In this review article we discuss emerging research on microglial activation phenotypes in the context of acute brain injury, and the potential role of microglia in phenotype-specific neurorestorative processes such as neurogenesis, angiogenesis, oligodendrogenesis and regeneration. We also describe some of the known molecular mechanisms that regulate phenotype switching, and highlight new therapeutic approaches that alter microglial activation state balance to enhance long-term functional recovery after TBI. An improved understanding of the regulatory mechanisms that control microglial phenotypic shifts may advance our knowledge of post-injury recovery and repair, and provide opportunities for the development of novel therapeutic strategies for TBI.

Delayed mGluR5 activation limits neuroinflammation and neurodegeneration after traumatic brain injury

BackgroundTraumatic brain injury initiates biochemical processes that lead to secondary neurodegeneration. Imaging studies suggest that tissue loss may continue for months or years after traumatic brain injury in association with chronic microglial activation. Recently we found that metabotropic glutamate receptor 5 (mGluR5) activation by (RS)-2-chloro-5-hydroxyphenylglycine (CHPG) decreases microglial activation and release of associated pro-inflammatory factors in vitro, which is mediated in part through inhibition of reduced nicotinamide adenine dinucleotide phosphate (NADPH) oxidase. Here we examined whether delayed CHPG administration reduces chronic neuroinflammation and associated neurodegeneration after experimental traumatic brain injury in mice.MethodsOne month after controlled cortical impact traumatic brain injury, C57Bl/6 mice were randomly assigned to treatment with single dose intracerebroventricular CHPG, vehicle or CHPG plus a selective mGluR5 antagonist, 3-((2-Methyl-4-thiazolyl)ethynyl)pyridine. Lesion volume, white matter tract integrity and neurological recovery were assessed over the following three months.ResultsTraumatic brain injury resulted in mGluR5 expression in reactive microglia of the cortex and hippocampus at one month post-injury. Delayed CHPG treatment reduced expression of reactive microglia expressing NADPH oxidase subunits; decreased hippocampal neuronal loss; limited lesion progression, as measured by repeated T2-weighted magnetic resonance imaging (at one, two and three months) and white matter loss, as measured by high field ex vivo diffusion tensor imaging at four months; and significantly improved motor and cognitive recovery in comparison to the other treatment groups.ConclusionMarkedly delayed, single dose treatment with CHPG significantly improves functional recovery and limits lesion progression after experimental traumatic brain injury, likely in part through actions at mGluR5 receptors that modulate neuroinflammation.

Activation of Metabotropic Glutamate Receptor 5 Modulates Microglial Reactivity and Neurotoxicity by Inhibiting NADPH Oxidase

Microglial-related factors have been implicated in the signaling cascades that contribute to neuronal cell death in various neurodegenerative disorders. Thus, strategies that reduce microglial activation and associated neurotoxicity may have therapeutic benefit. Group II and III metabotropic glutamate receptors (mGluRs) are expressed in microglia and can modulate microglial activity in primary cell cultures. We demonstrate that the group I receptor member mGluR5 is highly expressed in primary microglial cultures and the BV2 microglial cell line. Activation of mGluR5 using the selective agonist (RS)-2-chloro-5-hydroxyphenylglycine (CHPG) significantly attenuates microglial activation in response to lipopolysaccharide and interferon-γ, as indicated by a reduction in the expression of inducible nitric-oxide synthase, production of nitric oxide and tumor necrosis factor-α, and intracellular generation of reactive oxygen species. In addition, microglial-induced neurotoxicity is also markedly reduced by CHPG treatment. The anti-inflammatory effects of CHPG are mediated by the mGluR5 receptor, because either a selective mGluR5 antagonist or small interference RNA knockdown attenuated the actions of this drug. CHPG blocked the lipopolysaccharide-induced increase in expression and enzymatic activity of NADPH oxidase. Moreover, the protective effects of CHPG were significantly reduced when the NADPH oxidase subunits p22phox or gp91phox were knocked down by small interference RNA. These data suggest that mGluR5 represents a novel target for modulating microglial-dependent neuroinflammation, and may have therapeutic relevance for neurological disorders that exhibit microglial-mediated neurodegeneration.

Interleukin-4 mediates the neuroprotective effects of rosiglitazone in the aged brain

Increased expression of proinflammatory cytokines, like interleukin-1 beta (IL-1 beta), is a feature of the aged brain and it is generally accepted that the primary cell source of these cytokines is activated microglia. In hippocampus of aged rats, the increase in IL-1 beta is accompanied by microglial activation and impaired long-term potentiation (LTP). Peroxisome proliferator-activated receptors (PPARs) possess anti-inflammatory properties that target microglia. In this study the PPAR gamma agonist, rosiglitazone, was orally administered to young and aged rats, and we report that the age-related increases in NO and IL-1 beta production were attenuated in hippocampus of rosiglitazone-treated aged rats and that this was associated with a restoration of LTP. In addition, treatment with rosiglitazone increased interleukin-4 (IL-4) mRNA and reversed the age-related decrease in hippocampal IL-4 concentration. Significantly, while rosiglitazone attenuated the LPS-induced increase in MHCII and IL-1 beta concentration in glia prepared from wildtype mice, it failed to exert an effect in glia prepared from IL-4(-/-) mice, thereby suggesting that the anti-inflammatory actions of rosiglitazone are mediated by its ability to increase IL-4 expression.