Dataset Information

TRIM9-Mediated Resolution of Neuroinflammation Confers Neuroprotection upon Ischemic Stroke in Mice.

ABSTRACT: Excessive and unresolved neuroinflammation is a key component of the pathological cascade in brain injuries such as ischemic stroke. Here, we report that TRIM9, a brain-specific tripartite motif (TRIM) protein, was highly expressed in the peri-infarct areas shortly after ischemic insults in mice, but expression was decreased in aged mice, which are known to have increased neuroinflammation after stroke. Mechanistically, TRIM9 sequestered β-transducin repeat-containing protein (β-TrCP) from the Skp-Cullin-F-box ubiquitin ligase complex, blocking IκBα degradation and thereby dampening nuclear factor κB (NF-κB)-dependent proinflammatory mediator production and immune cell infiltration to limit neuroinflammation. Consequently, Trim9-deficient mice were highly vulnerable to ischemia, manifesting uncontrolled neuroinflammation and exacerbated neuropathological outcomes. Systemic administration of a recombinant TRIM9 adeno-associated virus that drove brain-wide TRIM9 expression effectively resolved neuroinflammation and alleviated neuronal death, especially in aged mice. These findings reveal that TRIM9 is essential for resolving NF-κB-dependent neuroinflammation to promote recovery and repair after brain injury and may represent an attractive therapeutic target.

SUBMITTER: Zeng J

PROVIDER: S-EPMC6485958 | biostudies-literature |

REPOSITORIES: biostudies-literature

ACCESS DATA

Similar Datasets

Project description:BackgroundNeuroinflammation, which is mainly mediated by excessive microglia activation, plays a major role in ischemic stroke. Overactivated microglia secrete numerous inflammatory cytokines, causing excessive inflammatory responses and ultimately exacerbating ischemic brain injury. Hence, compounds that attenuate neuroinflammation could become promising drug candidates for ischemic stroke. Fraxetin has an anti-inflammatory effect in many inflammatory diseases. However, whether it possesses an anti-inflammatory capacity in microglia-mediated neuroinflammation after ischemic brain injury is unknown. Our study aimed to investigate the suppression effect of fraxetin on neuroinflammation in lipopolysaccharide (LPS)-activated microglia and establish whether fraxetin could alleviate ischemic brain injury in a rodent model of ischemic stroke.MethodsFor the in vitro experiment, primary microglia were obtained from 1-day-old C57/BL6J mice. The cells were activated with LPS and treated with fraxetin at a non-cytotoxic concentration. Real-time PCR, enzyme-linked immunosorbent assays, and immunofluorescence staining were used to evaluate the anti-inflammatory effects of fraxetin. The potential molecular mechanisms were explored and verified through RNA-sequencing analysis, western blotting and real-time PCR. For the in vivo experiment, focal ischemia was induced by middle cerebral artery occlusion (MCAO) in 8-week-old male C57/BL6J mice. Fraxetin (5 mg/kg) or an equal volume of saline was injected into mice intraperitoneally after MCAO, and 2% 2,3,5-triphenyltetrazolium chloride staining was applied to measure infarct volume. Behavioral tests were conducted to measure neurological deficits in the mice. Real-time PCR, western blotting, and immunofluorescence staining were used to examine the expression of inflammatory cytokines and microglia activation in the ischemic penumbra.ResultsFraxetin effectively inhibited the expression of proinflammatory cytokines including inducible nitric oxide synthase, tumor necrosis factor-α, interleukin-1 beta, and interleukin-6 in LPS-activated microglia. Fraxetin also suppressed the PI3K/Akt/NF-κB signaling pathway in activated microglia, which contributed to its anti-inflammatory effects. Furthermore, the administration of fraxetin attenuated ischemic brain injury and behavioral deficits after stroke. Finally, fraxetin was found to attenuate the activation of microglia both in vitro and in vivo.ConclusionsOur results suggest that fraxetin has a suppression effect on microglia-mediated neuroinflammation, and this effect is associated with the PI3K/Akt/NF-κB signaling pathway. Fraxetin may therefore have potential neuroprotective properties for ischemic stroke.

Project description:BACKGROUND:Dendritic cell-associated C-type lectin-1 (Dectin-1) receptor has been reported to be involved in neuroinflammation in Alzheimer's disease and traumatic brain injury. The present study was designed to investigate the role of Dectin-1 and its downstream target spleen tyrosine kinase (Syk) in early brain injury after ischemic stroke using a focal cortex ischemic stroke model. METHODS:Adult male C57BL/6 J mice were subjected to a cerebral focal ischemia model of ischemic stroke. The neurological score, adhesive removal test, and foot-fault test were evaluated on days 1, 3, 5, and 7 after ischemic stroke. Dectin-1, Syk, phosphorylated (p)-Syk, tumor necrosis factor-α (TNF-α), and inducible nitric oxide synthase (iNOS) expression was analyzed via western blotting in ischemic brain tissue after ischemic stroke and in BV2 microglial cells subjected to oxygen-glucose deprivation/reoxygenation (OGD/R) injury in vitro. The brain infarct volume and Iba1-positive cells were evaluated using Nissl's and immunofluorescence staining, respectively. The Dectin-1 antagonist laminarin (LAM) and a selective inhibitor of Syk phosphorylation (piceatannol; PIC) were used for the intervention. RESULTS:Dectin-1, Syk, and p-Syk expression was significantly enhanced on days 3, 5, and 7 and peaked on day 3 after ischemic stroke. The Dectin-1 antagonist LAM or Syk inhibitor PIC decreased the number of Iba1-positive cells and TNF-α and iNOS expression, decreased the brain infarct volume, and improved neurological functions on day 3 after ischemic stroke. In addition, the in vitro data revealed that Dectin-1, Syk, and p-Syk expression was increased following the 3-h OGD and 0, 3, and 6 h of reperfusion in BV2 microglial cells. LAM and PIC also decreased TNF-α and iNOS expression 3 h after OGD/R induction. CONCLUSION:Dectin-1/Syk signaling plays a crucial role in inflammatory activation after ischemic stroke, and further investigation of Dectin-1/Syk signaling in stroke is warranted.

Project description:BACKGROUND:Remote ischemic preconditioning (RIPC) of a limb has been reported to protect against ischemic stroke. Our previous results demonstrated that the RIPC-mediated neuroprotection is associated with alterations in circulating immune cell populations. Here, we evaluated the effect of the spleen, the largest reservoir of immune cells, on RIPC-mediated neuroprotection against stroke. METHODS:Noninvasive RIPC was achieved by four repeated cycles of 5-min blood flow constriction in the hindlimbs using a tourniquet. The blood and spleens were collected before and 1 h and 3 days after preconditioning to analyze the effect of RIPC on the spleen and the correlation between splenic and peripheral lymphocytes. Moreover, spleen weight and splenic lymphocytes were compared in stroke rats with or without RIPC. Finally, splenectomy was made 1 day or 2 weeks before RIPC and 90-min middle cerebral artery occlusion (MCAO). The infarct areas and deficits were assessed. Blood was collected 1 h after RIPC and 3 days after MCAO to explore the impact of splenectomy on RIPC-induced neuroprotection and immune changes. The contralateral and ipsilateral hemispheres were collected 3 days after MCAO to detect the infiltration of immune cells after RIPC and splenectomy. RESULTS:Flow cytometry analysis demonstrated that the RIPC promptly increased the percentages of CD3+CD8+ cytotoxic T (Tc) cells in the spleen with a relatively delayed elevation in CD3+CD161+ natural killer T (NKT) and CD3-CD45RA+ B lymphocytes. The percentages of circulating lymphocytes are positively correlated with the percentages of splenic lymphocytes in normal rats. Interestingly, RIPC resulted in negative correlations between the percentages of splenic and circulating T lymphocytes, while the correlation between splenic and circulating B lymphocytes remained positive. For animals subjected to RIPC followed by MCAO, RIPC increased splenic volume with an expansion of splenic lymphocytes 3 days after MCAO. Furthermore, the removal of the spleen 1 day or 2 weeks before RIPC and MCAO reduced the protective effect of RIPC on ischemic brain injury and reversed the effects of RIPC on circulating immune cell composition. RIPC significantly reduced brain infiltration of Tc and NKT cells. Prior splenectomy showed no effect on immune cell infiltration after RIPC and stroke. CONCLUSION:These results reveal an immunomodulatory effect of the spleen, effecting mainly the spleen-derived lymphocytes, during RIPC-afforded neuroprotection against cerebral ischemia.

Dataset Information

TRIM9-Mediated Resolution of Neuroinflammation Confers Neuroprotection upon Ischemic Stroke in Mice.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets