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Early Axonal Injury and Delayed Cytotoxic Cerebral Edema are Associated with Microglial Activation in a Mouse Model of Sepsis.

ABSTRACT: Sepsis-induced brain injury is associated with an acute deterioration of mental status resulting in cognitive impairment and acquisition of new functional limitations in sepsis survivors. However, the exact nature of brain injury in this setting is often subtle and remains to be fully characterized both in preclinical studies and at the bedside. Given the translation potential for the use of magnetic resonance imaging (MRI) to define sepsis-induced brain injury, we sought to determine and correlate the cellular changes with neuroradiographic presentations in a classic murine model of sepsis induced by cecal ligation and puncture (CLP). Sepsis was induced in 6-10-week-old male C57/BL6 mice by CLP. We used immunohistochemistry (IHC) to define neuropathology in a mouse model of sepsis along with parallel studies using MRI, focusing on cerebral edema, blood-brain barrier (BBB) disruption, and microglial activation on days 1 and 4 days after CLP. We demonstrate that septic mice had evidence of early axonal injury, inflammation, and robust microglial activation on day 1 followed by cytotoxic edema on day 4 in the cortex, thalamus, and hippocampus in the absence of BBB disruption. We note the superiority of the MRI to detect subtle brain injury and cytotoxic cerebral edema in comparison with the traditional gold standard assessment, i.e., percent brain water (wet-dry weight method). We conclude that inflammatory changes in the septic brain can be detected in real time, and further studies are needed to understand axonal injury and the impact of inhibition of microglial activation on the development of cerebral edema.

SUBMITTER: Pang D

PROVIDER: S-EPMC7062563 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:Sepsis-associated brain injury (SABI) is characterized by an acute deterioration of mental status resulting in cognitive impairment and acquisition of new and persistent functional limitations in sepsis survivors. Previously, we reported that septic mice had evidence of axonal injury, robust microglial activation, and cytotoxic edema in the cerebral cortex, thalamus, and hippocampus in the absence of blood-brain barrier disruption. A key conceptual advance in the field was identification of sulfonylurea receptor 1 (SUR1), a member of the adenosine triphosphate (ATP)-binding cassette protein superfamily, that associates with the transient receptor potential melastatin 4 (TRPM4) cation channel to play a crucial role in cerebral edema development. Therefore, we hypothesized that knockout (KO) of Abcc8 (Sur1 gene) is associated with a decrease in microglial activation, cerebral edema, and improved neurobehavioral outcomes in a murine cecal ligation and puncture (CLP) model of sepsis. Sepsis was induced in 4-6-week-old Abcc8 KO and wild-type (WT) littermate control male mice by CLP. We used immunohistochemistry to define neuropathology and microglial activation along with parallel studies using magnetic resonance imaging, focusing on cerebral edema on days 1 and 4 after CLP. Abcc8 KO mice exhibited a decrease in axonal injury and cytotoxic edema vs. WT on day 1. Abcc8 KO mice also had decreased microglial activation in the cerebral cortex vs. WT. These findings were associated with improved spatial memory on days 7-8 after CLP. Our study challenges a key concept in sepsis and suggests that brain injury may not occur merely as an extension of systemic inflammation. We advance the field further and demonstrate that deletion of the SUR1 gene ameliorates CNS pathobiology in sepsis including edema, axonal injury, neuroinflammation, and behavioral deficits. Benefits conferred by Abcc8 KO in the murine CLP model warrant studies of pharmacological Abcc8 inhibition as a new potential therapeutic strategy for SABI.

Project description:We previously found that Mertk and its ligand Gas6, astrocytic genes involved in phagocytosis, are upregulated after acute sleep deprivation. These results suggested that astrocytes may engage in phagocytic activity during extended wake, but direct evidence was lacking. Studies in humans and rodents also found that sleep loss increases peripheral markers of inflammation, but whether these changes are associated with neuroinflammation and/or activation of microglia, the brain's resident innate immune cells, was unknown. Here we used serial block-face scanning electron microscopy to obtain 3D volume measurements of synapses and surrounding astrocytic processes in mouse frontal cortex after 6-8 h of sleep, spontaneous wake, or sleep deprivation (SD) and after chronic (∼5 d) sleep restriction (CSR). Astrocytic phagocytosis, mainly of presynaptic components of large synapses, increased after both acute and chronic sleep loss relative to sleep and wake. MERTK expression and lipid peroxidation in synaptoneurosomes also increased to a similar extent after short and long sleep loss, suggesting that astrocytic phagocytosis may represent the brain's response to the increase in synaptic activity associated with prolonged wake, clearing worn components of heavily used synapses. Using confocal microscopy, we then found that CSR but not SD mice show morphological signs of microglial activation and enhanced microglial phagocytosis of synaptic elements, without obvious signs of neuroinflammation in the CSF. Because low-level sustained microglia activation can lead to abnormal responses to a secondary insult, these results suggest that chronic sleep loss, through microglia priming, may predispose the brain to further damage.SIGNIFICANCE STATEMENT We find that astrocytic phagocytosis of synaptic elements, mostly of presynaptic origin and in large synapses, is upregulated already after a few hours of sleep deprivation and shows a further significant increase after prolonged and severe sleep loss, suggesting that it may promote the housekeeping of heavily used and strong synapses in response to the increased neuronal activity of extended wake. By contrast, chronic sleep restriction but not acute sleep loss activates microglia, promotes their phagocytic activity, and does so in the absence of overt signs of neuroinflammation, suggesting that like many other stressors, extended sleep disruption may lead to a state of sustained microglia activation, perhaps increasing the brain's susceptibility to other forms of damage.

Project description:Systemic inflammation affects brain functions. In our previous study in which lipopolysaccharide (LPS) was injected intraperitoneally into mice at sublethal doses, choroid plexus macrophages produced interleukin-1β and stimulated neighboring stromal cells. Activated stromal cells stimulate choroid plexus epithelial cells, and then choroid plexus epithelium-derived cytokines enter the brain parenchyma and stimulate astrocytes. Stimulated astrocytes then produce cytokines such as CCL11, CXCL10 and G-CSF and change the brain parenchymal microenvironment. However, the effects of an altered brain microenvironment on other brain cells remain to be determined. In the present study, we hypothesized that microglia are activated in response to astrocyte-induced changes in the brain microenvironment. Using the brains of mice treated with intraperitoneal LPS injection, Luminex multiplex cytokine immunoassays revealed increased hippocampal concentrations of CCL11, CXCL10 and G-CSF at 48 h after systemic LPS challenge. The concentrations of all cytokines examined returned to control levels at 72 h after LPS injection, which indicated a resolution of the neuroinflammation. Immunohistochemistry revealed that microglia were hypertrophied in mice at 48 h after systemic LPS challenge. Following isolation of microglial cells from the brain using magnetic-activated cell sorting, gene expression assays were performed with real-time reverse transcriptase-polymerase chain reaction. Isolated microglial cells exhibited much higher gene expression of the receptors for CCL11, CXCL10 and G-CSF than other brain cells. Microglial cells isolated from the brains of mice at 48 h after systemic LPS challenge exhibited the M2-like phenotype. In conclusion, microglial hypertrophy occurs following astrocytic reactions in a mouse model of sublethal endotoxemia-induced systemic inflammation, and hypertrophic microglia are polarized toward the M2-like phenotype and involved in the resolution of neuroinflammation.

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Early Axonal Injury and Delayed Cytotoxic Cerebral Edema are Associated with Microglial Activation in a Mouse Model of Sepsis.

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