Inflammatory episodes observed in multiple sclerosis are characterized by perivascular infiltration of central nervous system (CNS) tissue by circulating leucocytes directed by chemokines and activation of both microglia and astrocytes.... more
Inflammatory episodes observed in multiple sclerosis are characterized by perivascular infiltration of central nervous system (CNS) tissue by circulating leucocytes directed by chemokines and activation of both microglia and astrocytes. An in vitro approach, using a three-dimensional aggregate culture system derived from foetal rat brain tissue, is being taken to study the effect of inflammatory mediators on CNS cells. These aggregates model the brain in vivo in terms of glial and neuronal development and cell-cell interactions. Aggregates cultures were treated with lipopolysaccharide (LPS) and the pro-inflammatory cytokines tumour necrosis factor-alpha (TNF-α) and interferon-gamma (IFNγ). The effect of these treatments on the expression of a range of chemokine receptors (CCR1, CCR2, CCR5, CXCR3 and CX3CR1) and other markers of cell activation (MHC class I and II, CR3) was determined at the mRNA level by semiquantitative RT-PCR and at the protein level by confocal microscopy. Rat peritoneal macrophages were loaded with a fluorescent cell-tracer and their ability to infiltrate inflamed aggregates treated with IFNγ was analysed by FACS and confocal microscopy.
The COVID-19 pandemic imposed a series of behavioral changes that resulted in increased social isolation and a more sedentary life for many across all age groups, but, above all, for the elderly population who are the most vulnerable to... more
The COVID-19 pandemic imposed a series of behavioral changes that resulted in increased social isolation and a more sedentary life for many across all age groups, but, above all, for the elderly population who are the most vulnerable to infections and chronic neurodegenerative diseases. Systemic inflammatory responses are known to accelerate neurodegenerative disease progression, which leads to permanent damage, loss of brain function, and the loss of autonomy for many aged people. During the COVID-19 pandemic, a spectrum of inflammatory responses was generated in affected individuals, and it is expected that the elderly patients with chronic neurodegenerative diseases who survived SARSCoV-2 infection, it will be found, sooner or later, that there is a worsening of their neurodegenerative conditions. Using mouse prion disease as a model for chronic neurodegeneration, we review the effects of social isolation, sedentary living, and viral infection on the disease progression with a focus on sickness behavior and on the responses of microglia and astrocytes. Focusing on aging, we discuss the cellular and molecular mechanisms related to immunosenescence in chronic neurodegenerative diseases and how infections may accelerate their progression.
Intracortical neural probes enable researchers to measure electrical and chemical signals in the brain. However, penetration injury from probe insertion into living brain tissue leads to an inflammatory tissue response. In turn, microglia... more
Intracortical neural probes enable researchers to measure electrical and chemical signals in the brain. However, penetration injury from probe insertion into living brain tissue leads to an inflammatory tissue response. In turn, microglia are activated, which leads to encapsulation of the probe and release of pro-inflammatory cytokines. This inflammatory tissue response alters the electrical and chemical microen-vironment surrounding the implanted probe, which may in turn interfere with signal acquisition. Dexamethasone (Dex), a potent anti-inflammatory steroid, can be used to prevent and diminish tissue disruptions caused by probe implantation. Herein, we report retrodialysis administration of dexa-methasone while using in vivo two-photon microscopy to observe real-time microglial reaction to the implanted probe. Microdialysis probes under artificial cerebrospinal fluid (aCSF) perfusion with or without Dex were implanted into the cortex of transgenic mice that express GFP in microglia under the CX3CR1 promoter and imaged for 6 h. Acute morphological changes in microglia were evident around the microdialysis probe. The radius of microglia activation was 177.1 mm with aCSF control compared to 93.0 mm with Dex perfusion. T-stage morphology and microglia directionality indices were also used to quantify the microglial response to implanted probes as a function of distance. Dexamethasone had a profound effect on the microglia morphology and reduced the acute activation of these cells.
Several lines of evidence implicate microglial activation and abnormal immune response in the etiology of psychosis. Previous positron emission tomography (PET) neuroimaging studies of the translocator protein 18 kDa, TSPO, were limited... more
Several lines of evidence implicate microglial activation and abnormal immune response in the etiology of psychosis. Previous positron emission tomography (PET) neuroimaging studies of the translocator protein 18 kDa, TSPO, were limited by low affinity of the first-generation radioligand, low resolution scanners, and small sample sizes. Moreover, there is a dearth of literature on microglial activation in individuals at clinical high risk (CHR) for psychosis. We used a novel second generation TSPO radioligand, [18F]FEPPA, to examine whether microglial activation is elevated in the dorsolateral prefrontal cortex (DLPFC) and hippocampus of antipsychotic-naïve CHR. Twentyfour CHR (antipsychotic-naïve n = 22) and 23 healthy volunteers (HV) completed a high resolution[18F]FEPPA PET scan and MRI. The PET data were analyzed using the validated two-tissue compartment model with arterial plasma input function with total volume of distribution (VT) as outcome measure. All analyses were controlled for the TSPO rs6971 polymorphism. We did not observe any significant differences in microglial activation, as indexed by [18F]FEPPA VT, between CHR and HV in either the DLPFC (F(1, 44)=.41, p=.52) or the hippocampus (F(1, 44) = 2.78, p= .10).Exploratory associations show that in CHR [18F]FEPPA VT was positively correlated with apathy (DLPFC: r = .55, p= .008; Hippocampus: r = .52, p= .013) and state anxiety (DLPFC: r = .60, p=.003; Hippocampus: r = .48, p= .024). The lack of significant group differences in [18F]FEPPA VT suggests that microglial activation is not significantly elevated in the clinical high risk state thatprecedes psychosis.
Many RNA virus CNS infections cause neurological disease. Because Piry virus has a limited human pathogenicity and exercise reduces activation of microglia in aged mice, possible influences of environment and aging on microglial... more
Many RNA virus CNS infections cause neurological disease. Because Piry virus has a limited human pathogenicity and exercise reduces activation of microglia in aged mice, possible influences of environment and aging on microglial morphology and behavior in mice sublethal encephalitis were investigated. Female albino Swiss mice were raised either in standard (S) or in enriched (EE) cages from age 2 to 6 months (young-Y), or from 2 to 16 months (aged-A). After behavioral tests, mice nostrils were instilled with Piry-virus-infected or with normal brain homogenates. Brain sections were immunolabeled for virus antigens or microglia at 8 days post-infection (dpi), when behavioral changes became apparent, and at 20 and 40 dpi, after additional behavioral testing. Young infected mice from standard (SYPy) and enriched (EYPy) groups showed similar transient impairment in burrowing activity and olfactory discrimination , whereas aged infected mice from both environments (EAPy, SAPy) showed permanent reduction in both tasks. The beneficial effects of an enriched environment were smaller in aged than in young mice. Six-hundred and forty microglial cells, 80 from each group were reconstructed. An unbiased, stereological sampling approach and multivariate statistical analysis were used to search for microglial morphological families. This procedure allowed distinguishing between microglial morphology of infected and control subjects. More severe virus-associated microglial changes were observed in young than in aged mice, and EYPy seem to recover microglial homeostatic morphology earlier than SYPy. Because Piry-virus encephalitis outcomes were more severe in aged mice, it is suggested that the reduced inflammatory response in those individuals may aggravate encephalitis outcomes.
Implantable neural electrodes must drastically improve chronic recording stability before they can be translated into long-term human clinical prosthetics. Previous studies suggest that sub-cellular sized and mechanically compliant probes... more
Implantable neural electrodes must drastically improve chronic recording stability before they can be translated into long-term human clinical prosthetics. Previous studies suggest that sub-cellular sized and mechanically compliant probes may result in improved tissue integration and recording longevity. However, currently these design features are restricted by the opposing mechanical requirements needed for minimally damaging insertions. We designed a non-cytotoxic, carboxymethylcellulose (CMC) based dissolvable delivery vehicle (shuttle) to provide the mechanical support for insertion of ultra-small, ultra-compliant microfabricated neural probes. Stiff CMC-based shuttles rapidly soften immediately after being placed ∼1 mm above an open craniotomy as they absorb vapors from the brain. To address this, we developed a sophisticated targeting, high speed insertion (∼80 mm/s), and release system to implant these shuttles. After implantation, the goal is for the shuttle to dissolve away leaving only the electrodes behind. Here we show the histology of chronically implanted shuttles of large (300 μm × 125 μm) and small (100 μm × 125 μm) size at discrete time points over 12 weeks. Early time points show the CMC shuttle expanded after insertion as it absorbed moisture from the brain and slowly dissolved. At later time points neuronal cell bodies populate regions within the original shuttle tract. The large CMC shuttles show that the CMC expansion can cause extended secondary damage. On the other hand, the smaller CMC shuttles show limited secondary damage, wound closure by 4 weeks, absence of activated microglia at 12 weeks, as well as evidence suggesting neural regeneration at the implant site. This shuttle, therefore, shows great promise facilitating the implantation of nontraditional ultra-small, and ultra-compliant probes.
Penetrating traumatic brain injury (PTBI) is a significant cause of death and disability in the United States. Inflammasomes are one of the key regulators of the interleukin (IL)–1b mediated inflammatory responses after traumatic brain... more
Penetrating traumatic brain injury (PTBI) is a significant cause of death and disability in the United States. Inflammasomes are one of the key regulators of the interleukin (IL)–1b mediated inflammatory responses after traumatic brain injury. However, the contribution of inflammasome signaling after PTBI has not been determined. In this study, adult male Sprague-Dawley rats were subjected to sham procedures or penetrating ballistic-like brain injury (PBBI) and sacrificed at various time-points. Tissues were assessed by immunoblot analysis for expression of IL-1b, IL-18, and components of the inflammasome: apoptosis-associated speck-like protein containing a caspase-activation and recruitment domain (ASC), caspase-1, X-linked inhibitor of apoptosis protein (XIAP), nucleotide-binding oligomerization domain (NOD)-like receptor protein 3 (NLRP3), and gasdermin-D (GSDMD). Specific cell types expressing inflammasome proteins also were evaluated immunohistochemically and assessed quantitatively. After PBBI, expression of IL-1b, IL-18, caspase-1, ASC, XIAP, and NLRP3 peaked around 48 h. Brain protein lysates from PTBI animals showed pyroptosome formation evidenced by ASC laddering, and also contained increased expression of GSDMD at 48 h after injury. ASC-positive immunoreactive neurons within the perilesional cortex were observed at 24 h. At 48 h, ASC expression was concentrated in morphologically activated cortical microglia. This expression of ASC in activated microglia persisted until 12 weeks following PBBI. This is the first report of inflammasome activation after PBBI. Our results demonstrate cell-specific patterns of inflammasome activation and pyroptosis predominantly in microglia, suggesting a sustained pro-inflammatory state following PBBI, thus offering a therapeutic target for this type of brain injury.
Traumatic brain injury (TBI) is still the worldwide leading cause of mortality and morbidity in young adults. Improved safety measures and advances in critical care have improved chances of surviving a TBI, however, numerous secondary... more
Traumatic brain injury (TBI) is still the worldwide leading cause of mortality and morbidity in young adults. Improved safety measures and advances in critical care have improved chances of surviving a TBI, however, numerous secondary mechanisms contribute to the injury in the weeks and months that follow TBI. The past 4 decades of research have addressed many of the metabolic impairments sufficient to mitigate mortality, however, an enduring secondary mechanism, i.e. neuroinflammation, has been intractable to current therapy. Neuroinflammation is particularly difficult to target with pharmacological agents due to lack of specificity, the blood brain barrier, and an incomplete understanding of the protective and pathologic influences of inflammation in TBI. Recent insights into TBI pathophysiology have established microglial activation as a hallmark of all types of TBI. The inflammatory response to injury is necessary and beneficial while the death of activated microglial is not. This review presents new insights on the therapeutic and maladaptive features of the immune response after TBI with emphasis on microglial polarization, followed by a discussion of potential targets for pharmacologic and non-pharmacologic treatments. In aggregate, this review presents a rationale for guiding TBI inflammation towards neural repair and regeneration rather than secondary injury and degeneration, which we posit could improve outcomes and reduce lifelong disease burden in TBI survivors.
p38 MAPK has been implicated in the regulation of prom-flammatory cytokines and apoptosis in vitro. To understand its role in neurodegeneration, we determined the time course and localization of the dually phosphorylated active form of... more
p38 MAPK has been implicated in the regulation of prom-flammatory cytokines and apoptosis in vitro. To understand its role in neurodegeneration, we determined the time course and localization of the dually phosphorylated active form of p38 MAPK in hippocampus after global forebrain ischemia. Phosphorylated p38 MAPK and mitogen-activated protein kinase-activated protein 2 activity increased over 4 days after ischemia. Phosphorylated p38 MAPK immunoreactivity was observed in microglia in regions adjacent to, but not in, the dying CAl neurons. In contrast, neither c-Jun N-terminal kinase 1 nor p42/p44M~activity was altered after ischemia. These results provide the first evidence for localization of activated p38 MAPK in the CNS and support a role for p38 MAPK in the microglial response to stress.
Although the interleukin-1 converting enzyme (ICE)/CED-3 family of proteases has been implicated recently in neuronal cell death in vitro and in ovo, the role of specific genes belonging to this family in cell death in the nervous system... more
Although the interleukin-1 converting enzyme (ICE)/CED-3 family of proteases has been implicated recently in neuronal cell death in vitro and in ovo, the role of specific genes belonging to this family in cell death in the nervous system remains unknown. To address this question, we examined the in vivo expression of one of these genes, Ice, after global forebrain ischemia in gerbils. Using RT-PCR and Western immunoblot techniques, we detected an increase in the mRNA and protein expression of ICE in hippocampus during a period of 4 d after ischemia. Chromatin condensation was observed in CA1 neu-rons within 2 d after ischemia. Internucleosomal DNA fragmentation and apoptotic bodies were observed between 3 and 4 d after ischemia, a period during which CA1 neuronal death is maximal. In nonischemic brains, ICE-like immunoreactivity was relatively low in CA1 pyramidal neurons but high in scattered hippocampal interneurons. After ischemia, ICE-like immunore-activity was not altered in these neurons. ICE-like immunore-activity, however, was observed in microglial cells in the regions adjacent to the CA1 layer as early as 2 d after ischemic insult. The increase in ICE-like immunoreactivity was robust at 4 d after ischemia, a period that correlates with the DNA fragmentation observed in hippocampal homogenates of isch-emic brains. These results provide the first evidence for the localization and induction of ICE expression in vivo after isch-emia and suggest an indirect role for ICE in ischemic damage through mediation of an inflammatory response.
It is well established that microglial form and function are inextricably linked. In recent years, the traditional view that microglial form ranges between "ramified... more
It is well established that microglial form and function are inextricably linked. In recent years, the traditional view that microglial form ranges between "ramified resting" and "activated amoeboid" has been emphasized through advancing imaging techniques that point to microglial form being highly dynamic even within the currently accepted morphological categories. Moreover, microglia adopt meaningful intermediate forms between categories, with considerable crossover in function and varying morphologies as they cycle, migrate, wave, phagocytose, and extend and retract fine and gross processes. From a quantitative perspective, it is problematic to measure such variability using traditional methods, but one way of quantitating such detail is through fractal analysis. The techniques of fractal analysis have been used for quantitating microglial morphology, to categorize gross differences but also to differentiate subtle differences (e.g., amongst ramified cells). Multifractal analysis in particular is one technique of fractal analysis that may be useful for identifying intermediate forms. Here we review current trends and methods of fractal analysis, focusing on box counting analysis, including lacunarity and multifractal analysis, as applied to microglial morphology.
