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Augmented ?2-adrenergic signaling dampens the neuroinflammatory response following ischemic stroke and increases stroke size.

ABSTRACT: BACKGROUND:Ischemic stroke provokes a neuroinflammatory response and simultaneously promotes release of epinephrine and norepinephrine by the sympathetic nervous system. This increased sympathetic outflow can act on ?2-adrenergic receptors expressed by immune cells such as brain-resident microglia and monocyte-derived macrophages (MDMs), but the effect on post-stroke neuroinflammation is unknown. Thus, we investigated how changes in ?2-adrenergic signaling after stroke onset influence the microglia/MDM stroke response, and the specific importance of microglia/MDM ?2-adrenergic receptors to post-stroke neuroinflammation. METHODS:To investigate the effects of ?2-adrenergic receptor manipulation on post-stroke neuroinflammation, we administered the ?2-adrenergic receptor agonist clenbuterol to mice 3?h after the onset of photothrombotic stroke. We immunostained to quantify microglia/MDM numbers and proliferation and to assess morphology and activation 3?days later. We assessed stroke outcomes by measuring infarct volume and functional motor recovery and analyzed gene expression levels of neuroinflammatory molecules. Finally, we evaluated changes in cytokine expression and microglia/MDM response in brains of mice with selective knockout of the ?2-adrenergic receptor from microglia and monocyte-lineage cells. RESULTS:We report that clenbuterol treatment after stroke onset causes enlarged microglia/MDMs and impairs their proliferation, resulting in reduced numbers of these cells in the peri-infarct cortex by 1.7-fold at 3?days after stroke. These changes in microglia/MDMs were associated with increased infarct volume in clenbuterol-treated animals. In mice that had the ?2-adrenergic receptor specifically knocked out of microglia/MDMs, there was no change in morphology or numbers of these cells after stroke. However, knockdown of ?2-adrenergic receptors in microglia and MDMs resulted in increased expression of TNF? and IL-10 in peri-infarct tissue, while stimulation of ?2-adrenergic receptors with clenbuterol had the opposite effect, suppressing TNF? and IL-10 expression. CONCLUSIONS:We identified ?2-adrenergic receptor signaling as an important regulator of the neuroimmune response after ischemic stroke. Increased ?2-adrenergic signaling after stroke onset generally suppressed the microglia/MDM response, reducing upregulation of both pro- and anti-inflammatory cytokines, and increasing stroke size. In contrast, diminished ?2-adrenergic signaling in microglia/MDMs augmented both pro- and anti-inflammatory cytokine expression after stroke. The ?2-adrenergic receptor may therefore present a therapeutic target for improving the post-stroke neuroinflammatory and repair process.

SUBMITTER: Lechtenberg KJ

PROVIDER: S-EPMC6540436 | biostudies-literature | 2019 May

REPOSITORIES: biostudies-literature

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Augmented β2-adrenergic signaling dampens the neuroinflammatory response following ischemic stroke and increases stroke size.

Lechtenberg Kendra J KJ Meyer Scott T ST Doyle Janelle B JB Peterson Todd C TC Buckwalter Marion S MS

Journal of neuroinflammation 20190528 1

<h4>Background</h4>Ischemic stroke provokes a neuroinflammatory response and simultaneously promotes release of epinephrine and norepinephrine by the sympathetic nervous system. This increased sympathetic outflow can act on β2-adrenergic receptors expressed by immune cells such as brain-resident microglia and monocyte-derived macrophages (MDMs), but the effect on post-stroke neuroinflammation is unknown. Thus, we investigated how changes in β2-adrenergic signaling after stroke onset influence th ...[more]

PMID: 31138227

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Project description:BackgroundHeat-shock protein B1 (HSPB1) is among the most well-known and versatile member of the evolutionarily conserved family of small heat-shock proteins. It has been implicated to serve a neuroprotective role against various neurological disorders via its modulatory activity on inflammation, yet its exact role in neuroinflammation is poorly understood. In order to shed light on the exact mechanism of inflammation modulation by HSPB1, we investigated the effect of HSPB1 on neuroinflammatory processes in an in vivo and in vitro model of acute brain injury.MethodsIn this study, we used a transgenic mouse strain overexpressing the human HSPB1 protein. In the in vivo experiments, 7-day-old transgenic and wild-type mice were treated with ethanol. Apoptotic cells were detected using TUNEL assay. The mRNA and protein levels of cytokines and glial cell markers were examined using RT-PCR and immunohistochemistry in the brain. We also established primary neuronal, astrocyte, and microglial cultures which were subjected to cytokine and ethanol treatments. TNFα and hHSPB1 levels were measured from the supernates by ELISA, and intracellular hHSPB1 expression was analyzed using fluorescent immunohistochemistry.ResultsFollowing ethanol treatment, the brains of hHSPB1-overexpressing mice showed a significantly higher mRNA level of pro-inflammatory cytokines (Tnf, Il1b), microglia (Cd68, Arg1), and astrocyte (Gfap) markers compared to wild-type brains. Microglial activation, and 1 week later, reactive astrogliosis was higher in certain brain areas of ethanol-treated transgenic mice compared to those of wild-types. Despite the remarkably high expression of pro-apoptotic Tnf, hHSPB1-overexpressing mice did not exhibit higher level of apoptosis. Our data suggest that intracellular hHSPB1, showing the highest level in primary astrocytes, was responsible for the inflammation-regulating effects. Microglia cells were the main source of TNFα in our model. Microglia isolated from hHSPB1-overexpressing mice showed a significantly higher release of TNFα compared to wild-type cells under inflammatory conditions.ConclusionsOur work provides novel in vivo evidence that hHSPB1 overexpression has a regulating effect on acute neuroinflammation by intensifying the expression of pro-inflammatory cytokines and enhancing glial cell activation, but not increasing neuronal apoptosis. These results suggest that hHSPB1 may play a complex role in the modulation of the ethanol-induced neuroinflammatory response.

Project description:Background. Women account for 60% of all stroke deaths and are more often permanently disabled than men, despite their higher observed stroke incidence. Considering the clinical population affected by stroke, an obvious drawback is that many pre-clinical and clinical studies only investigate young males. To improve therapeutic translation from bench to bedside, we believe that it is advantageous to include both sexes in experimental models of stroke. The aims of this study were to identify early cerebral vascular responses to ischemic stroke in females, compare the differential gene expression patterns with those seen in males, and identify potential new therapeutic targets. Results. Transient middle cerebral artery occlusion (tMCAO) was used to induce stroke in both female and male rats, the middle cerebral arteries (MCAs) were isolated 3 hours post reperfusion and RNA was extracted. Affymetrix whole transcriptome expression profiling was performed on female (n = 12) MCAs to reveal differentially expressed genes. In total, 1076 genes had an increased expression and 879 genes a decreased expression in the occluded MCAs as compared with the control MCAs from female rats. An enrichment of genes related to apoptosis, regulation of transcription, protein autophosphorylation, inflammation, oxidative stress, and tissue repair and recovery were seen in the occluded MCA. The high expression genes chosen for qPCR verification (Adamts4, Olr1, JunB, Fosl1, Serpine1, S1pr3, Ccl2 and Socs3) were all shown to be upregulated in the same manner in both females and males after tMCAO (p < 0.05; n = 23). When comparing the differentially expressed genes in female MCAs (occluded and non-occluded) with our previous findings in males after tMCAO, a total of 297 genes overlapped (all groups had 32 genes in common). Conclusions. The cascades of processes initiated in the vasculature following reperfusion are complex. Dynamic gene expression alterations were observed in the occluded MCAs, and to a less pronounced degree in the non-occluded MCAs. Dysregulation of inflammation and blood-brain barrier breakdown are possible pharmacological targets. The sample of genes (<1% of differentially expressed genes) validated for this microarray did not reveal any sex differences.

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Augmented ?2-adrenergic signaling dampens the neuroinflammatory response following ischemic stroke and increases stroke size.

Publications

Augmented β2-adrenergic signaling dampens the neuroinflammatory response following ischemic stroke and increases stroke size.

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