Kimberly R. Byrnes

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.

Metabotropic glutamate receptors as targets for multipotential treatment of neurological disorders

SummaryGlutamate is a major excitatory neurotransmitter in the CNS that is involved in numerous cellular functions, including cell death and survival. Metabotropic glutamate receptors (mGluR) are G-protein coupled receptors that have been classified into three groups on the basis of signal transduction pathways and pharmacological profiles. Group I, II, and III mGluRs are found on cell types within and peripheral to the CNS, including neurons, microglia, astrocytes, oligodendrocytes, T- and B-cell lymphocytes, osteoblasts, hepatocytes, and endothelial cells, among others. These receptors have a number of effects on cells that can influence outcome after trauma, including reducing neuronal and oligodendroglial cell death, inflammation, and endothelial permeability. Thus, mGluRs are a promising multipotential therapeutic approach. Because the pathology of CNS trauma and neurodegeneration is multifactorial (including, for example, oxidative stress, mitochondrial breakdown, and inflammation), therapies that serve to modulate multiple pathophysiological pathways may prove more effective than those directed at a single target. This review examines the multipotential therapeutic utility of mGluR modulation in acute and chronic injury and neurodegeneration.

Role of the cell cycle in the pathobiology of central nervous system trauma

Up-regulation of cell cycle proteins occurs in both mitotic and post-mitotic neural cells after central nervous system (CNS) injury in adult animals. In mitotic cells, such as astroglia and microglia, they induce proliferation, whereas in post-mitotic cells such as neurons they initiate caspase-related apoptosis. We recently reported that early central administration of the cell cycle inhibitor flavopiridol after experimental traumatic brain injury (TBI) significantly reduced lesion volume, scar formation and neuronal cell death, while promoting near complete behavioral recovery. Here we show that in primary neuronal or astrocyte cultures structurally different cell cycle inhibitors (flavopiridol, roscovitine, and olomoucine) significantly reduce up-regulation of cell cycle proteins, attenuate neuronal cell death induced by etoposide, and decrease astrocyte proliferation. Flavopiridol, in a concentration dependent manner, also attenuates proliferation/activation of microglia. In addition, we demonstrate that central administration of flavopiridol improves functional outcome in dose-dependent manner after fluid percussion induced brain injury in rats. Moreover, delayed systemic administration of flavopiridol significantly reduces brain lesion volume and edema development after TBI. These data provide further support for the therapeutic potential of cell cycle inhibitors for the treatment of clinical CNS injury and that protective mechanisms likely include reduction of neuronal cell death, inhibition of glial proliferation and attenuation of microglial activation.

Metabotropic glutamate receptor 5 activation inhibits microglial associated inflammation and neurotoxicity

The Group I metabotropic glutamate receptor 5 (mGluR5) can modulate addiction, pain, and neuronal cell death. Expression of some mGluRs, such as Group II and III mGluRs, has been reported in microglia and may affect their activation. However, the expression and role of mGluR5 in microglia is unclear. Using immunocytochemistry and Western blot, we demonstrate that mGluR5 protein is expressed in primary microglial cultures. Activation of mGluR5 using the selective agonist (RS)‐2‐chloro‐5‐hydroxyphenylglycine (CHPG) significantly reduces microglial activation in response to lipopolysaccharide, as indicated by a reduction in nitric oxide, reactive oxygen species, and TNFα production. Microglial induced neurotoxicity is also markedly reduced by CHPG treatment. The anti‐inflammatory effects of CHPG are not observed in microglial cultures from mGluR5 knockout mice and are blocked by selective mGluR5 antagonists, suggesting that these actions are mediated by the mGluR5 receptor. Anti‐inflammatory actions of mGluR5 activation are attenuated by phospholipase C and protein kinase C inhibitors, as well as by calcium chelators, suggesting that the mGluR5 activation in microglia involves the Gαq‐protein signal transduction pathway. These data indicate that microglial mGluR5 may represent a novel target for modulating neuroinflammation, an important component of both acute and chronic neurodegenerative disorders.

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.

Roscovitine Reduces Neuronal Loss, Glial Activation, and Neurologic Deficits after Brain Trauma

Traumatic brain injury (TBI) causes both direct and delayed tissue damage. The latter is associated with secondary biochemical changes such as cell cycle activation, which leads to neuronal death, inflammation, and glial scarring. Flavopiridol—a cyclin-dependent kinase (CDK) inhibitor that is neither specific nor selective—is neuroprotective. To examine the role of more specific CDK inhibitors as potential neuroprotective agents, we studied the effects of roscovitine in TBI. Central administration of roscovitine 30 mins after injury resulted in significantly decreased lesion volume, as well as improved motor and cognitive recovery. Roscovitine attenuated neuronal death and inhibited activation of cell cycle pathways in neurons after TBI, as indicated by attenuated cyclin G1 accumulation and phosphorylation of retinoblastoma protein. Treatment also decreased microglial activation after TBI, as reflected by reductions in ED1, galectin-3, p22PHOX, and Iba-1 levels, and attenuated astrogliosis, as shown by decreased accumulation of glial fibrillary acidic protein. In primary cortical microglia and neuronal cultures, roscovitine and other selective CDK inhibitors attenuated neuronal cell death, as well as decreasing microglial activation and microglial-dependent neurotoxicity. These data support a multifactorial neuroprotective effect of cell cycle inhibition after TBI—likely related to inhibition of neuronal apoptosis, microglial-induced inflammation, and gliosis—and suggest that multiple CDKs are potentially involved in this process.

Role of Cell Cycle Proteins in CNS Injury

Following trauma or ischemia to the central nervous system (CNS), there is a marked increase in the expression of cell cycle-related proteins. This up-regulation is associated with apoptosis of post-mitotic cells, including neurons and oligodendrocytes, both in vitro and in vivo. Cell cycle activation also induces proliferation of astrocytes and microglia, contributing to the glial scar and microglial activation with release of inflammatory factors. Treatment with cell cycle inhibitors in CNS injury models inhibits glial scar formation and neuronal cell death, resulting in substantially decreased lesion volumes and improved behavioral recovery. Here we critically review the role of cell cycle pathways in the pathophysiology of experimental stroke, traumatic brain injury and spinal cord injury, and discuss the potential of cell cycle inhibitors as neuroprotective agents.

Selective CDK inhibitor limits neuroinflammation and progressive neurodegeneration after brain trauma

Traumatic brain injury (TBI) induces secondary injury mechanisms, including cell-cycle activation (CCA), which lead to neuronal cell death, microglial activation, and neurologic dysfunction. Here, we show progressive neurodegeneration associated with microglial activation after TBI induced by controlled cortical impact (CCI), and also show that delayed treatment with the selective cyclin-dependent kinase inhibitor roscovitine attenuates posttraumatic neurodegeneration and neuroinflammation. CCI resulted in increased cyclin A and D1 expressions and fodrin cleavage in the injured cortex at 6 hours after injury and significant neurodegeneration by 24 hours after injury. Progressive neuronal loss occurred in the injured hippocampus through 21 days after injury and correlated with a decline in cognitive function. Microglial activation associated with a reactive microglial phenotype peaked at 7 days after injury with sustained increases at 21 days. Central administration of roscovitine at 3 hours after CCI reduced subsequent cyclin A and D1 expressions and fodrin cleavage, improved functional recovery, decreased lesion volume, and attenuated hippocampal and cortical neuronal cell loss and cortical microglial activation. Furthermore, delayed systemic administration of roscovitine improved motor recovery and attenuated microglial activation after CCI. These findings suggest that CCA contributes to progressive neurodegeneration and related neurologic dysfunction after TBI, likely in part related to its induction of microglial activation.

Expression of two temporally distinct microglia-related gene clusters after spinal cord injury

The dual role of microglia in cytotoxicity and neuroprotection is believed to depend on the specific, temporal expression of microglial‐related genes. To better clarify this issue, we used high‐density oligonucleotide microarrays to examine microglial gene expression after spinal cord injury (SCI) in rats. We compared expression changes at the lesion site, as well as in rostral and caudal regions after mild, moderate, or severe SCI. Using microglial‐associated anchor genes, we identified two clusters with different temporal profiles. The first, induced by 4 h postinjury to peak between 4 and 24 h, included interleukin‐1β, interleukin‐6, osteopontin, and calgranulin, among others. The second was induced 24 h after SCI, and peaked between 72 h and 7 days; it included C1qB, Galectin‐3, and p22phox. These two clusters showed similar expression profiles regardless of injury severity, albeit with slight decreases in expression in mild or severe injury vs. moderate injury. Expression was also decreased rostral and caudal to the lesion site. We validated the expression of selected cluster members at the mRNA and protein levels. In addition, we demonstrated that stimulation of purified microglia in culture induces expression of C1qB, Galectin‐3, and p22phox. Finally, inhibition of p22phox activity within microglial cultures significantly suppressed proliferation in response to stimulation, confirming that this gene is involved in microglial activation. Because microglial‐related factors have been implicated both in secondary injury and recovery, identification of temporally distinct clusters of genes related to microglial activation may suggest distinct roles for these groups of factors.

FDG-PET imaging in mild traumatic brain injury: a critical review

Traumatic brain injury (TBI) affects an estimated 1.7 million people in the United States and is a contributing factor to one third of all injury related deaths annually. According to the CDC, approximately 75% of all reported TBIs are concussions or considered mild in form, although the number of unreported mild TBIs (mTBI) and patients not seeking medical attention is unknown. Currently, classification of mTBI or concussion is a clinical assessment since diagnostic imaging is typically inconclusive due to subtle, obscure, or absent changes in anatomical or physiological parameters measured using standard magnetic resonance (MR) or computed tomography (CT) imaging protocols. Molecular imaging techniques that examine functional processes within the brain, such as measurement of glucose uptake and metabolism using [18F]fluorodeoxyglucose and positron emission tomography (FDG-PET), have the ability to detect changes after mTBI. Recent technological improvements in the resolution of PET systems, the integration of PET with magnetic resonance imaging (MRI), and the availability of normal healthy human databases and commercial image analysis software contribute to the growing use of molecular imaging in basic science research and advances in clinical imaging. This review will discuss the technological considerations and limitations of FDG-PET, including differentiation between glucose uptake and glucose metabolism and the significance of these measurements. In addition, the current state of FDG-PET imaging in assessing mTBI in clinical and preclinical research will be considered. Finally, this review will provide insight into potential critical data elements and recommended standardization to improve the application of FDG-PET to mTBI research and clinical practice.