Real-Time Noninvasive Bioluminescence, Ultrasound and Photoacoustic Imaging in NFκB-RE-Luc Transgenic Mice Reveal Glia Maturation Factor-Mediated Immediate and Sustained Spatio-Temporal Activation of NFκB Signaling Post-Traumatic Brain Injury in a Gender-Specific Manner.

Neurotrauma especially traumatic brain injury (TBI) is the leading cause of death and disability worldwide. To improve upon the early diagnosis and develop precision-targeted therapies for TBI, it is critical to understand the underlying molecular mechanisms and signaling pathways. The transcription factor, nuclear factor kappa B (NFκB), which is ubiquitously expressed, plays a crucial role in the normal cell survival, proliferation, differentiation, function, as well as in disease states like neuroinflammation and neurodegeneration. Here, we hypothesized that real-time noninvasive bioluminescence molecular imaging allows rapid and precise monitoring of TBI-induced immediate and rapid spatio-temporal activation of NFκB signaling pathway in response to Glia maturation factor (GMF) upregulation which in turn leads to neuroinflammation and neurodegeneration post-TBI. To test and validate our hypothesis and to gain novel mechanistic insights, we subjected NFκB-RE-Luc transgenic male and female mice to TBI and performed real-time noninvasive bioluminescence imaging (BLI) as well as photoacoustic and ultrasound imaging (PAI). Our BLI data revealed that TBI leads to an immediate and sustained activation of NFκB signaling. Further, our BLI data suggest that especially in male NFκB-RE-Luc transgenic mice subjected to TBI, in addition to brain, there is widespread activation of NFκB signaling in multiple organs. However, in the case of the female NFκB-RE-Luc transgenic mice, TBI induces a very specific and localized activation of NFκB signaling in the brain. Further, our microRNA data suggest that TBI induces significant upregulation of mir-9-5p, mir-21a-5p, mir-34a-5p, mir-16-3p, as well as mir-155-5p within 24 h and these microRNAs can be successfully used as TBI-specific biomarkers. To the best of our knowledge, this is one of the first and unique study of its kind to report immediate and sustained activation of NFκB signaling post-TBI in a gender-specific manner by utilizing real-time non-invasive BLI and PAI in NFκB-RE-Luc transgenic mice. Our study will prove immensely beneficial to gain novel mechanistic insights underlying TBI, unravel novel therapeutic targets, as well as enable us to monitor in real-time the response to innovative TBI-specific precision-targeted gene and stem cell-based precision medicine.

A role for glia maturation factor dependent activation of mast cells and microglia in MPTP induced dopamine loss and behavioural deficits in mice.

The molecular mechanism mediating degeneration of nigrostriatal dopaminergic neurons in Parkinson's disease (PD) is not yet fully understood. Previously, we have shown the contribution of glia maturation factor (GMF), a proinflammatory protein in dopaminergic neurodegeneration mediated by activation of mast cells (MCs). In this study, methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-induced nigrostriatal neurodegeneration and astro-glial activations were determined by western blot and immunofluorescence techniques in wild type (WT) mice, MC-deficient (MC-KO) mice and GMF-deficient (GMF-KO) mice, with or without MC reconstitution before MPTP administration. We show that GMF-KO in the MCs reduces the synergistic effects of MC and Calpain1 (calcium-activated cysteine protease enzyme)-dependent dopaminergic neuronal loss that reduces motor behavioral impairments in MPTP-treated mouse. Administration of MPTP increase in calpain-mediated proteolysis in nigral dopaminergic neurons further resulting in motor decline in mice. We found that MPTP administered WT mice exhibits oxidative stress due to significant increases in the levels of malondialdehyde, superoxide dismutase and reduction in the levels of reduced glutathione and glutathione peroxidase activity as compared with both MC-KO and GMF-KO mice. The number of TH-positive neurons in the ventral tegmental area, substantia nigra and the fibers in the striatum were significantly reduced while granulocyte macrophage colony-stimulating factor (GM-CSF), MC-Tryptase, GFAP, IBA1, Calpain1 and intracellular adhesion molecule 1 expression were significantly increased in WT mice. Similarly, tyrosine hydroxylase, dopamine transporters and vesicular monoamine transporters 2 proteins expression were significantly reduced in the SN of MPTP treated WT mice. The motor behavior as analyzed by rotarod and hang test was significantly reduced in WT mice as compared with both the MC-KO and GMF-KO mice. We conclude that GMF-dependent MC activation enhances the detrimental effect of astro-glial activation-mediated oxidative stress and neuroinflammation in the midbrain, and its inhibition may slowdown the progression of PD.

Mast Cell Activation, Neuroinflammation, and Tight Junction Protein Derangement in Acute Traumatic Brain Injury.

Traumatic brain injury (TBI) is one of the major health problems worldwide that causes death or permanent disability through primary and secondary damages in the brain. TBI causes primary brain damage and activates glial cells and immune and inflammatory cells, including mast cells in the brain associated with neuroinflammatory responses that cause secondary brain damage. Though the survival rate and the neurological deficiencies have shown significant improvement in many TBI patients with newer therapeutic options, the underlying pathophysiology of TBI-mediated neuroinflammation, neurodegeneration, and cognitive dysfunctions is understudied. In this study, we analyzed mast cells and neuroinflammation in weight drop-induced TBI. We analyzed mast cell activation by toluidine blue staining, serum chemokine C-C motif ligand 2 (CCL2) level by enzyme-linked immunosorbent assay (ELISA), and proteinase-activated receptor-2 (PAR-2), a mast cell and inflammation-associated protein, vascular endothelial growth factor receptor 2 (VEGFR2), and blood-brain barrier tight junction-associated claudin 5 and Zonula occludens-1 (ZO-1) protein expression in the brains of TBI mice. Mast cell activation and its numbers increased in the brains of 24 h and 72 h TBI when compared with sham control brains without TBI. Mouse brains after TBI show increased CCL2, PAR-2, and VEGFR2 expression and derangement of claudin 5 and ZO-1 expression as compared with sham control brains. TBI can cause mast cell activation, neuroinflammation, and derangement of tight junction proteins associated with increased BBB permeability. We suggest that inhibition of mast cell activation can suppress neuroimmune responses and glial cell activation-associated neuroinflammation and neurodegeneration in TBI.

Mast Cells Release Chemokine CCL2 in Response to Parkinsonian Toxin 1-Methyl-4-Phenyl-Pyridinium (MPP(+)).

Microglial activation and release of inflammatory cytokines and chemokines are crucial events in neuroinflammation. Microglial cells interact and respond to other inflammatory cells such as T cells and mast cells as well as inflammatory mediators secreted from these cells. Recent studies have shown that neuroinflammation causes and accelerates neurodegenerative disease such as Parkinson's disease (PD) pathogenesis. 1-methyl-4-phenyl-pyridinium ion (MPP(+)), the active metabolite of neurotoxin 1-methyl-4-phenyl-1,2,3,6-tetrahydro pyridine activates glial cells and mediate neurodegeneration through release of inflammatory mediators. We have shown that glia maturation factor (GMF) activates glia and induces neuroinflammation and neurodegeneration and that MPP(+) activates mast cells and release proinflammatory cytokines and chemokines. The chemokine (C-C motif) ligand 2 (CCL2) levels have been shown to be elevated and play a role in PD pathogenesis. In the present study, we analyzed if MPP(+) activates mouse and human mast cells to release chemokine CCL2. Mouse bone marrow-derived mast cells (BMMCs) and human umbilical cord blood-derived cultured mast cells (hCBMCs) were incubated with MPP(+) (10 µM) for 24 h and CCL2 levels were measured in the supernatant media by ELISA. MPP(+)-significantly induced CCL2 release from BMMCs and hCBMCs. Additionally, GMF overexpression in BMMCs obtained from wild-type mice released significantly more CCL2, while BMMCs obtained from GMF-deficient mice showed less CCL2 release. Further, we show that MPP(+)-induced CCL2 release was greater in BMMCs-astrocyte co-culture conditions. Uncoupling protein 4 (UCP4) which is implicated in neurodegenerative diseases including PD was detected in BMMCs by immunocytochemistry. Our results suggest that mast cells may play role in PD pathogenesis.

Absence of glia maturation factor protects dopaminergic neurons and improves motor behavior in mouse model of parkinsonism.

Previously, we have shown that aberrant expression of glia maturation factor (GMF), a proinflammatory protein, is associated with the neuropathological conditions underlying diseases suggesting an important role for GMF in neurodegeneration. In the present study, we demonstrate that absence of GMF suppresses dopaminergic (DA) neuron loss, glial activation, and expression of proinflammatory mediators in the substantia nigra pars compacta (SN) and striatum (STR) of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) treated mice. Dopaminergic neuron numbers in the SN and fiber densities in the STR were reduced in wild type (Wt) mice when compared with GMF-deficient (GMF-KO) mice after MPTP treatment. We compared the motor abnormalities caused by MPTP treatment in Wt and GMF-KO mice as measured by Rota rod and grip strength test. Results show that the deficits in motor coordination and decrease in dopamine and its metabolite content were protected significantly in GMF-KO mice after MPTP treatment when compared with control Wt mice under identical experimental conditions. These findings were further supported by the immunohistochemical analysis that showed reduced glial activation in the SN of MPTP-treated GMF-KO mice. Similarly, in MPTP-treated GMF-KO mice, production of inflammatory tumor necrosis factor alpha, interleukine-1 beta, granulocyte macrophage-colony stimulating factor, and the chemokine (C-C motif) ligand 2 MCP-1 was suppressed, findings consistent with a role for GMF in MPTP neurotoxicity. In conclusion, present investigation provides the first evidence that deficiency of GMF protects the DA neuron loss and reduces the inflammatory load following MPTP administration in mice. Thus depletion of endogenous GMF represents an effective and selective strategy to slow down the MPTP-induced neurodegeneration.

Glia maturation factor expression in entorhinal cortex of Alzheimer's disease brain.

Alzheimer's disease (AD) is characterized by the presence of neuropathological lesions containing amyloid plaques (APs) and hyperphosphorylated Tau containing neurofibrillary tangles (NFTs) and is associated with neuroinflammation and neurodegeneration. Entorhinal cortex (Brodmann's area 28) is involved in memory associated functions and is one of the first brain areas targeted to form the neuropathological lesions and also severely affected cortical region in AD. Glia maturation factor (GMF), a central nervous system protein and a proinflammatory molecule is known to be up-regulated in the specific areas of AD brain. Our previous immunohistochemical studies using temporal cortex showed that GMF is expressed in the vicinity of APs and NFTs in AD brains. In the present study, we have analyzed the expression of GMF and its association with APs and NFTs in the entorhinal cortex of AD brains by using immunohistochemistry combined with thioflavin-S fluorescence labeling methods. Results showed that GMF immunoreactive glial cells, glial fibrillary acidic protein labeled reactive astrocytes and ionized calcium binding adaptor molecule-1 labeled activated microglia were increased in the entorhinal cortical layers especially at the sites of 6E10 labeled APs and Tau containing NFTs. In conclusion, increased expression of GMF by the glial cells in the entorhinal cortex region, and the co-localization of GMF with APs and NFTs suggest that GMF may play important proinflammatory roles in the pathogenesis of AD.

Glia maturation factor expression in hippocampus of human Alzheimer's disease.

Alzheimer's disease (AD) is characterized by the presence of neuropathological lesions containing amyloid plaques (APs) and neurofibrillary tangles (NFTs) associated with neuroinflammation and neuronal degeneration. Hippocampus is one of the earliest and severely damaged areas in AD brain. Glia maturation factor (GMF), a known proinflammatory molecule is up-regulated in AD. Here, we have investigated the expression and distribution of GMF in relation to the distribution of APs and NFTs in the hippocampus of AD brains. Our immunohistochemical results showed GMF is expressed specifically in the vicinity of high density of APs and NFTs in the hippocampus of AD patients. Moreover, reactive astrocytes and activated microglia surrounds the APs and NFTs. We further demonstrate that GMF immunoreactive glial cells were increased at the sites of Tau containing NFTs and APs of hippocampus in AD brains. In conclusion, up-regulated expression of GMF in the hippocampus, and the co-localization of GMF and thioflavin-S stained NFTs and APs suggest that GMF may play important role in the pathogenesis of AD.

Enhanced expression of glia maturation factor correlates with glial activation in the brain of triple transgenic Alzheimer's disease mice.

We previously demonstrated that glia maturation factor (GMF), a brain specific protein, isolated, sequenced and cloned in our laboratory, induce expression of proinflammatory cytokines and chemokines in the central nervous system. We also reported that the up-regulation of GMF in astrocytes leads to the destruction of neurons suggesting a novel pathway of GMF-mediated cytotoxicity of brain cells, and implicated its involvement in the pathogenesis of inflammatory neurodegenerative diseases. In the present study, we examined the expressions of GMF in triple-transgenic Alzheimer's disease (3xTg-AD) mice. Our results show a 13-fold up-regulation of GMF and 8-12-fold up-regulation of proinflammatory cytokines tumor necrosis factor-alpha (TNF-α), interleukin-6 (IL-6), IL-1ß, interferon gamma (IFN-γ), and chemokine (C-C motif) ligand 2 (CCL2) and C-X-C motif chemokine 10 (CXCL10/IP-10) mRNA as determined by quantitative real-time RT-PCR in the brain of 3xTg-AD mice as compared to non-transgenic (Non-Tg) mice. In conclusion, the increase in GMF and cytokine/chemokine expression was correlated with reactive glial fibrillary acidic protein positive astrocytes and ionized calcium binding adaptor molecule 1 (Iba-1)-positive microglia in 3xTg-AD mice.

Efficient down-regulation of glia maturation factor expression in mouse brain and spinal cord.

Long-lasting siRNA-based down-regulation of gene of interest can be achieved by lentiviral-based expression vectors driving the production of short hairpin RNA (shRNA). We investigated an attractive therapeutic approach to target the expression of proinflammatory GMF by using lentiviral vector encoding GMF-specific shRNA to reduce GMF levels in the spinal cord and brain of mice. To determine the effect of GMF-shRNA on GMF protein levels, we performed quantitative ELISA analysis in brain and in thoracic, cervical and lumbar regions of spinal cord from mice followed by GMF-shRNA (G-shRNA) or control shRNA (C-shRNA) treatments. Our results show a marked reduction of GMF protein levels in brain and spinal cord of mice treated with GMF-shRNA compared to control shRNA treatment. Consistent with the GMF protein analysis, the immunohistochemical examination of the spinal cord sections of EAE mice treated with GMF-shRNA showed significantly reduced GMF-immunoreactivity. Thus, the down-regulation of GMF by GMF-shRNA was efficient and wide spread in CNS as evident by the significantly reduced levels of GMF protein in the brain and spinal cord of mice.

Immune Suppression of Glia Maturation Factor Reverses Behavioral Impairment, Attenuates Amyloid Plaque Pathology and Neuroinflammation in an Alzheimer's Disease Mouse Model.

Alzheimer's disease (AD) is an irreversible progressive neurodegenerative disorder recognized by accumulation of amyloid-plaques (APs) and neurofibrillary tangles (NFTs) and eventually loss of memory. Glia maturation factor (GMF), a neuroinflammatory protein first time isolated and cloned in our laboratory plays an important role in the pathogenesis of AD. However, no studies have been reported on whether anti-GMF antibody administration could downregulate neuroinflammation and attenuate amyloid pathology in AD brain. We investigated the potential effect of single dose of (2 mg/kg b.wt/mouse) intravenously (iv) injected with anti-GMF antibodyon cognitive function, neuroprotection, neuroinflammation and Aß load in the brain of 9-month-old 5XFAD mice. Following 4 weeks of anti-GMF antibody delivery in mice, we found reduced expression of GMF, astrocytic glial fibrillary acidic protein (GFAP) and microglial ionizing calcium binding adaptor molecule 1 (Iba1) as well as improvement inneuroinflammatory response via inhibition of pro-inflammatory cytokines (TNF-α, IL-1ß and IL-6) production and amyloid pathology in the cerebral cortex and hippocampal CA1 region of 5XFAD mice. Correspondingly, blockade of GMF function with anti-GMF antibody improved spatial learning, memory, and long-term recognition memory in 5XFAD mice. The present study demonstrates that the immune checkpoint blockade of GMF function with anti-GMF antibody coordinates anti-inflammatory effects to attenuate neurodegeneration in the cortex and hippocampal CA1 region of 5XFAD mouse brain. Further, our data suggest, that pharmacological immune neutralization of GMF is a promising neuroprotective strategy totherapeutically target neuroinflammation and neurodegeneration in AD. Graphical Abstract 5XFAD mice Polyclonal anti-GMF antibody.

Acute Traumatic Brain Injury-Induced Neuroinflammatory Response and Neurovascular Disorders in the Brain.

Acute traumatic brain injury (TBI) leads to neuroinflammation, neurodegeneration, cognitive decline, psychological disorders, increased blood-brain barrier (BBB) permeability, and microvascular damage in the brain. Inflammatory mediators secreted from activated glial cells, neurons, and mast cells are implicated in the pathogenesis of TBI through secondary brain damage. Abnormalities or damage to the neurovascular unit is the indication of secondary injuries in the brain after TBI. However, the precise mechanisms of molecular and ultrastructural neurovascular alterations involved in the pathogenesis of acute TBI are not yet clearly understood. Moreover, currently, there are no precision-targeted effective treatment options to prevent the sequelae of TBI. In this study, mice were subjected to closed head weight-drop-induced acute TBI and evaluated neuroinflammatory and neurovascular alterations in the brain by immunofluorescence staining or quantitation by enzyme-linked immunosorbent assay (ELISA) procedure. Mast cell stabilizer drug cromolyn was administered to inhibit the neuroinflammatory response of TBI. Results indicate decreased level of pericyte marker platelet-derived growth factor receptor-beta (PDGFR-ß) and BBB-associated tight junction proteins junctional adhesion molecule-A (JAM-A) and zonula occludens-1 (ZO-1) in the brains 7 days after weight-drop-induced acute TBI as compared with the brains from sham control mice indicating acute TBI-associated BBB/tight junction protein disruption. Further, the administration of cromolyn drug significantly inhibited acute TBI-associated decrease of PDGFR-ß, JAM-A, and ZO-1 in the brain. These findings suggest that acute TBI causes BBB/tight junction damage and that cromolyn administration could protect this acute TBI-induced brain damage as well as its long-time consequences.

Cytokines, brain proteins, and growth factors in acute stroke patients: A pilot study.

BACKGROUND: Immunomodulation and cell signaling involve several cytokines, proteins, and other mediators released in response to the trauma, inflammation, or other insults to the central nervous system. This pilot study is part of the registry designed to evaluate the temporal trends among these molecules after an acute ischemic stroke (AIS) in patients. METHODS: Twelve AIS patients were enrolled within 24 hours of the symptoms onset. Two sets of plasma samples were collected: First at admission and second at 24 hours after admission. Cytokines/chemokines and other inflammatory molecules were measured using multiplex assay kit. RESULTS: An increased trend in IL-6 (22 vs. 34 pg/ml), IL-8/CXCL8 (87 vs. 98 pg/ml), MMP-9 (16225 vs. 18450 pg/ml), and GMF-ß (999 vs. 3739 pg/ml) levels was observed overtime after an AIS. Patients ≤60 years had lower levels of plasma MCP-1/CCL2 (50-647 vs. 150-1159 pg/ml), IL-6 (9-25 vs. 20-68 pg/ml), and IL-8 (30- 143 vs. 72-630 pg/ml), when compared with patients >60 years old. CONCLUSION: Cytokines/chemokines and other inflammatory mediators play an important role in the pathogenesis of stroke in addition to mediating poststroke inflammation. Further research is needed to evaluate and characterize the cumulative trends of these mediators for the clinical prognosis or as surrogate biomarkers.

Neuroprotective effects of flavone luteolin in neuroinflammation and neurotrauma.

Neuroinflammation leads to neurodegeneration, cognitive defects, and neurodegenerative disorders. Neurotrauma/traumatic brain injury (TBI) can cause activation of glial cells, neurons, and neuroimmune cells in the brain to release neuroinflammatory mediators. Neurotrauma leads to immediate primary brain damage (direct damage), neuroinflammatory responses, neuroinflammation, and late secondary brain damage (indirect) through neuroinflammatory mechanism. Secondary brain damage leads to chronic inflammation and the onset and progression of neurodegenerative diseases. Currently, there are no effective and specific therapeutic options to treat these brain damages or neurodegenerative diseases. Flavone luteolin is an important natural polyphenol present in several plants that show anti-inflammatory, antioxidant, anticancer, cytoprotective, and macrophage polarization effects. In this short review article, we have reviewed the neuroprotective effects of luteolin in neurotrauma and neurodegenerative disorders and pathways involved in this mechanism. We have collected data for this study from publications in the PubMed using the keywords luteolin and mast cells, neuroinflammation, neurodegenerative diseases, and TBI. Recent reports suggest that luteolin suppresses systemic and neuroinflammatory responses in Coronavirus disease 2019 (COVID-19). Studies have shown that luteolin exhibits neuroprotective effects through various mechanisms, including suppressing immune cell activation, such as mast cells, and inflammatory mediators released from these cells. In addition, luteolin can suppress neuroinflammatory response, activation of microglia and astrocytes, oxidative stress, neuroinflammation, and the severity of neuroinflammatory diseases such as Alzheimer's disease, Parkinson's disease, multiple sclerosis, and TBI pathogenesis. In conclusion, luteolin can improve cognitive decline and enhance neuroprotection in neurodegenerative diseases, TBI, and stroke.

Overexpression of glia maturation factor reinstates susceptibility to myelin oligodendrocyte glycoprotein-induced experimental autoimmune encephalomyelitis in glia maturation factor deficient mice.

Glia maturation factor (GMF), a primarily CNS localized protein was discovered and characterized in our laboratory. We previously demonstrated that GMF is the upstream regulator for excessive production and release of proinflammatory cytokines/chemokines in brain cells leading to the destruction of oligodendrocytes, the myelin forming cells, and neurons. We also reported that mice lacking endogenous GMF (GMF-deficient, GMF-KO) were resistant to myelin oligodendrocyte glycoprotein peptide 35-55 (MOG(35-55)) induced EAE, since immunization induced only delayed EAE with diminished severity. In the present study we show that a replication-defective adenovirus-GMF construct caused expression of GMF in CNS of GMF-KO mice and reinstated MOG(35-55) induced early and severe EAE. Our results show that MOG(35-55) immunization caused only a muted EAE and inflammation/demyelination in mice lacking endogenous GMF. The diminished incidence of EAE in GMF-KO mice was consistent with the significantly reduced expressions of cytokines/chemokines. The muted severity of EAE in GMF-KO mice was restored to full blown levels upon reintroduction of GMF using an adeno-GMF-virus (Adv-GMF) vector. Consistent with the clinical findings, histological examination of the CNS of mice with EAE revealed profound differences between wild type (Wt), GMF-KO, and GMF-KO mice with re-introduced GMF (GMF-KO+Adv-GMF). Spinal cord sections from mice with EAE were analyzed for the infiltration of mononuclear cells (inflammation) and myelin loss (demyelination). In Wt mice, 40% of spinal cord quadrants were positive for demyelination and 45% of spinal cord quadrants were positive for inflammation at the peak of EAE. Drastically reduced infiltrates (15%) and demyelination (10%) were found in GMF-KO mice that developed reduced severity of EAE. Upon GMF reintroduction in GMF-KO mice, MOG(35-55) immunization caused extensive monocytes infiltration (48%) and demyelination (46%), similar to that observed in the immunized Wt mice. The levels of cytokine/chemokine in the spinal cords of mice at three time points, corresponding to the onset, peak severity and recovery period of EAE, show a distinct pattern of very large increases in IFN-γ, TNF-α, GM-CSF and MCP-1 in Wt and GMF-KO+Adv-GMF mice compared to GMF-KO and GMF-KO+Adv-LacZ mice.

Current Trends in Biomarkers for Traumatic Brain Injury.

Neurotrauma, especially Traumatic Brain Injury (TBI) is a major health concern not only for the civilian population but also for the military personnel. Currently there are no precision and regenerative therapies available for the successful treatment of TBI patients. Hence, early detection and treatment options may prevent the severity and untoward harmful effects of TBI. However, currently there are no effective biomarkers available for the rapid and robust diagnosis as well as prognosis of TBI. Several biomarkers in blood, cerebrospinal fluid (CSF), saliva and urine have been explored to assess the onset, progression, severity and prognosis of TBI recently. Present knowledge on the blood biomarkers including cytokines and chemokines and in vivo imaging modalities are useful to some extent to detect and treat TBI patients. Here, we review S100B, Glial Fibrillary Acidic Protein (GFAP), Neuron Specific Enolase (NSE), Myelin Basic Protein (MBP), Ubiquitin C-terminal Hydrolase L1 (UCHL1), tau protein, and alpha spectrin II break down products regarding their usefulness as a set of reliable biomarkers for the robust diagnosis of TBI. We suggest that these biomarkers may prove very useful for the diagnosis and prognosis of TBI.

Glia Maturation Factor (GMF) Regulates Microglial Expression Phenotypes and the Associated Neurological Deficits in a Mouse Model of Traumatic Brain Injury.

Traumatic brain injury (TBI) induces inflammatory responses through microglial activation and polarization towards a more inflammatory state that contributes to the deleterious secondary brain injury. Glia maturation factor (GMF) is a pro-inflammatory protein that is responsible for neuroinflammation following insult to the brain, such as in TBI. We hypothesized that the absence of GMF in GMF-knockout (GMF-KO) mice would regulate microglial activation state and the M1/M2 phenotypes following TBI. We used the weight drop model of TBI in C57BL/6 mice wild-type (WT) and GMF-KO mice. Immunofluorescence staining, Western blot, and ELISA assays were performed to confirm TBI-induced histopathological and neuroinflammatory changes. Behavioral analysis was done to check motor coordination ability and cognitive function. We demonstrated that the deletion of GMF in GMF-KO mice significantly limited lesion volume, attenuated neuronal loss, inhibited gliosis, and activated microglia adopted predominantly anti-inflammatory (M2) phenotypes. Using an ELISA method, we found a gradual decrease in pro-inflammatory cytokines (TNF-α and IL-6) and upregulation of anti-inflammatory cytokines (IL-4 and IL-10) in GMF-KO mice compared with WT mice, thus, promoting the transition of microglia towards a more predominantly anti-inflammatory (M2) phenotype. GMF-KO mice showed significant improvement in motor ability, memory, and cognition. Overall, our results demonstrate that GMF deficiency regulates microglial polarization, which ameliorates neuronal injury and behavioral impairments following TBI in mice and concludes that GMF is a regulator of neuroinflammation and an ideal therapeutic target for the treatment of TBI.

NLRP3 inflammasome and glia maturation factor coordinately regulate neuroinflammation and neuronal loss in MPTP mouse model of Parkinson's disease.

Neuroinflammation plays an active role in the pathogenesis of several neurodegenerative diseases, including Parkinson's disease (PD). Earlier studies from this laboratory showed that glia maturation factor (GMF), a proinflammatory mediator; is up-regulated in the brain in neurodegenerative diseases and that deficiency of GMF showed decreased production of IL-1ß and improved behavioral abnormalities in mouse model of PD. However, the mechanisms linking GMF and dopaminergic neuronal death have not been completely explored. In the present study, we have investigated the expression of NLRP3 inflammasome and caspase-1 in the substantia nigra (SN) of human PD and non-PD brains by immunohistochemistry. Wild-type (WT) and GMF-/- (GMF knock-out) mice were treated with 1-methyl-4-phenyl-1,2,3,6-tetrahydro pyridine (MPTP) and the brains were isolated for neurochemical and morphological examinations. NLRP3 and caspase-1 positive cells were found significantly increased in PD when compared to non-PD control brains. Moreover, GMF co-localized with α-Synuclein within reactive astrocytes in the midbrain of PD. Mice treated with MPTP exhibit glial activation-induced inflammation, and nigrostriatal dopaminergic neurodegeneration. Interestingly, increased expression of the inflammasome components in astrocytes and microglia observed in the SN of MPTP-treated WT mice were significantly reduced in GMF-/- mice. Additionally, we show that NLRP3 activation in microglia leads to translocation of GMF and NLRP3 to the mitochondria. We conclude that downregulation of GMF may have beneficial effects in prevention of PD by modulating the cytotoxic functions of microglia and astrocytes through reduced activation of the NLRP3 inflammasome; a major contributor of neuroinflammation in the CNS.

COVID-19, Mast Cells, Cytokine Storm, Psychological Stress, and Neuroinflammation.

Coronavirus disease 2019 (COVID-19) caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is a new pandemic infectious disease that originated in China. COVID-19 is a global public health emergency of international concern. COVID-19 causes mild to severe illness with high morbidity and mortality, especially in preexisting risk groups. Therapeutic options are now limited to COVID-19. The hallmark of COVID-19 pathogenesis is the cytokine storm with elevated levels of interleukin-6 (IL-6), IL-1ß, tumor necrosis factor-alpha (TNF-α), chemokine (C-C-motif) ligand 2 (CCL2), and granulocyte-macrophage colony-stimulating factor (GM-CSF). COVID-19 can cause severe pneumonia, and neurological disorders, including stroke, the damage to the neurovascular unit, blood-brain barrier disruption, high intracranial proinflammatory cytokines, and endothelial cell damage in the brain. Mast cells are innate immune cells and also implicated in adaptive immune response, systemic inflammatory diseases, neuroinflammatory diseases, traumatic brain injury and stroke, and stress disorders. SARS-CoV-2 can activate monocytes/macrophages, dendritic cells, T cells, mast cells, neutrophils, and induce cytokine storm in the lung. COVID-19 can activate mast cells, neurons, glial cells, and endothelial cells. SARS-CoV-2 infection can cause psychological stress and neuroinflammation. In conclusion, COVID-19 can induce mast cell activation, psychological stress, cytokine storm, and neuroinflammation.

Brain Injury-Mediated Neuroinflammatory Response and Alzheimer's Disease.

Traumatic brain injury (TBI) is a major health problem in the United States, which affects about 1.7 million people each year. Glial cells, T-cells, and mast cells perform specific protective functions in different regions of the brain for the recovery of cognitive and motor functions after central nervous system (CNS) injuries including TBI. Chronic neuroinflammatory responses resulting in neuronal death and the accompanying stress following brain injury predisposes or accelerates the onset and progression of Alzheimer's disease (AD) in high-risk individuals. About 5.7 million Americans are currently living with AD. Immediately following brain injury, mast cells respond by releasing prestored and preactivated mediators and recruit immune cells to the CNS. Blood-brain barrier (BBB), tight junction and adherens junction proteins, neurovascular and gliovascular microstructural rearrangements, and dysfunction associated with increased trafficking of inflammatory mediators and inflammatory cells from the periphery across the BBB leads to increase in the chronic neuroinflammatory reactions following brain injury. In this review, we advance the hypothesis that neuroinflammatory responses resulting from mast cell activation along with the accompanying risk factors such as age, gender, food habits, emotional status, stress, allergic tendency, chronic inflammatory diseases, and certain drugs can accelerate brain injury-associated neuroinflammation, neurodegeneration, and AD pathogenesis.

Psychological Stress-Induced Immune Response and Risk of Alzheimer's Disease in Veterans from Operation Enduring Freedom and Operation Iraqi Freedom.

PURPOSE: Psychological stress is a significant health problem in veterans and their family members. Traumatic brain injury (TBI) and stress lead to the onset, progression, and worsening of several inflammatory and neurodegenerative diseases in veterans and civilians. Alzheimer's disease (AD) is a progressive, irreversible neuroinflammatory disease that causes problems with memory, thinking, and behavior. TBIs and chronic psychological stress cause and accelerate the pathology of neuroinflammatory diseases such as AD. However, the precise molecular and cellular mechanisms governing neuroinflammation and neurodegeneration are currently unknown, especially in veterans. The purpose of this review article was to advance the hypothesis that stress and TBI-mediated immune response substantially contribute and accelerate the pathogenesis of AD in veterans and their close family members and civilians. METHODS: The information in this article was collected and interpreted from published articles in PubMed between 1985 and 2020 using the key words stress, psychological stress, Afghanistan war, Operation Enduring Freedom (OEF), Iraq War, Operation Iraqi Freedom (OIF), Operation New Dawn (OND), traumatic brain injury, mast cell and stress, stress and neuroimmune response, stress and Alzheimer's disease, traumatic brain injury, and Alzheimer's disease. FINDINGS: Chronic psychological stress and brain injury induce the generation and accumulation of beta-amyloid peptide, amyloid plaques, neurofibrillary tangles, and phosphorylation of tau in the brain, thereby contributing to AD pathogenesis. Active military personnel and veterans are under enormous psychological stress due to various war-related activities, including TBIs, disabilities, fear, new environmental conditions, lack of normal life activities, insufficient communications, explosions, military-related noise, and health hazards. Brain injury, stress, mast cell, and other immune cell activation can induce headache, migraine, dementia, and upregulate neuroinflammation and neurodegeneration in veterans of Operation Enduring Freedom, Operation Iraqi Freedom, and Operation New Dawn. TBIs, posttraumatic stress disorder, psychological stress, pain, glial activation, and dementia in active military personnel, veterans, or their family members can cause AD several years later in their lives. We suggest that there are increasing numbers of veterans with TBIs and stress and that these veterans may develop AD late in life if no appropriate therapeutic intervention is available. IMPLICATIONS: Per these published reports, the fact that TBIs and psychological stress can accelerate the pathogenesis of AD should be recognized. Active military personnel, veterans, and their close family members should be evaluated regularly for stress symptoms to prevent the pathogenesis of neurodegenerative diseases, including AD.