BackgroundThe endoplasmic reticulum (ER) is responsible for the control of correct protein folding and protein function which is crucial for cell survival. However, under pathological conditions, such as hypoxia–ischemia (HI), there is an accumulation of unfolded proteins thereby triggering the unfolded protein response (UPR) and causing ER stress which is associated with activation of several stress sensor signaling pathways, one of them being the inositol requiring enzyme-1 alpha (IRE1α) signaling pathway. The UPR is regarded as a potential contributor to neuronal cell death and inflammation after HI. In the present study, we sought to investigate whether microRNA-17 (miR-17), a potential IRE1α ribonuclease (RNase) substrate, arbitrates downregulation of thioredoxin-interacting protein (TXNIP) and consequent NLRP3 inflammasome activation in the immature brain after HI injury and whether inhibition of IRE1α may attenuate inflammation via miR-17/TXNIP regulation.MethodsPostnatal day 10 rat pups (n = 287) were subjected to unilateral carotid artery ligation followed by 2.5 h of hypoxia (8% O2). STF-083010, an IRE1α RNase inhibitor, was intranasally delivered at 1 h post-HI or followed by an additional one administration per day for 2 days. MiR-17-5p mimic or anti-miR-17-5p inhibitor was injected intracerebroventricularly at 48 h before HI. Infarct volume and body weight were used to evaluate the short-term effects while brain weight, gross and microscopic brain tissue morphologies, and neurobehavioral tests were conducted for the long-term evaluation. Western blots, immunofluorescence staining, reverse transcription quantitative real-time polymerase chain reaction (RT-qPCR), and co-immunoprecipitation (Co-IP) were used for mechanism studies.ResultsEndogenous phosphorylated IRE1α expression was significantly increased after HI. Intranasal administration of STF-083010 alleviated brain injury and improved neurological behavior. MiR-17-5p expression was reduced after HI, and this decrease was attenuated by STF-083010 treatment. MiR-17-5p mimic administration ameliorated TXNIP expression, NLRP3 inflammasome activation, caspase-1 cleavage, and IL-1β production, as well as brain infarct volume. Conversely, anti-miR-17-5p inhibitor reversed IRE1α inhibition-induced decrease in TXNIP expression and inflammasome activation, as well as exacerbated brain injury after HI.ConclusionsIRE1a-induced UPR pathway may contribute to inflammatory activation and brain injury following neonatal HI. IRE1a activation, through decay of miR-17-5p, elevated TXNIP expression to activate NLRP3 inflammasome and aggravated brain damage.Electronic supplementary materialThe online version of this article (10.1186/s12974-018-1077-9) contains supplementary material, which is available to authorized users.

Background and purpose Lipoxin A4 (LXA4) has been reported to reduce inflammation in several neurological injury models. We studied the effects of LXA4 on neuroinflammation after subarachnoid hemorrhage (SAH) in a rat model. Methods Two hundred and thirty eight Sprague Dawley male rats, weight 280–320 g were used. Exogenous LXA4 (0.3 and 1.0 nmol) were injected intracerebroventricularly at 1.5 hours after SAH. Neurological scores, brain water content and blood-brain barrier were evaluated at 24 hours after SAH; Morris water maze and T-maze tests were examined at 21 days after SAH. The expression of endogenous LXA4 and its receptor formyl peptide receptor 2 (FPR2), as well as p38, IL-1β and IL-6 were studied either by ELISA or western blots. Neutrophil infiltration was observed by myeloperoxidase (MPO) staining. FPR2 siRNA was used to knock down LXA4 receptor. Results The expression of endogenous LXA4 decreased and the expression of FPR2 increased after SAH. Exogenous LXA4 decreased brain water content, reduced Evans blue extravasation, and improved neurological functions and improved the learning and memory ability after SAH. LXA4 reduced neutrophil infiltration and phosphorylation of p38, IL-1β and IL-6. These effects of LXA4 were abolished by FPR2 siRNA. Conclusion Exogenous LXA4 inhibited inflammation by activating FPR2 and inhibiting p38 after SAH. LXA4 may serve as an alternative treatment to relieve early brain injury after SAH.

Background and Purpose Mincle (macrophage-inducible C-type lectin, CLEC4E) receptor is reported involved in neuroinflammation in cerebral ischemia and traumatic brain injury. This study was designed to investigate the role of Mincle and its downstream Syk signal pathway in early brain injury after SAH in a rat model. Methods Two hundreds and fifteen (215) male Sprague-Dawley rats (280–320g) were subjected to endovascular perforation model of SAH. SAH grade, neurological score, and brain water content were measured at 24 h after SAH. Mincle/Syk as well as CARD9 (a member of the caspase-associated recruitment domain (CARD), involved in innate immune response), interleukin-1β (IL-1β) and myeloperoxidase (MPO) expressions were analyzed by western blot at 24 h after SAH. Specific cell types that expressed Mincle were detected with double immunofluorescence staining. Mincle siRNA, the endogenous ligand of Mincle receptor SAP130, and a selective Syk phosphorylation inhibitor piceatannol were used for intervention. Results Brain water content increased and neurological functions decreased in rats after SAH. The expression of SAP130, Mincle, Syk and p-Syk increased at 12h and peaked at 24h after SAH. Mincle siRNA reduced IL-1β and infiltration of MPO positive cells, decreased brain water content, and improved neurological functions at 24h after SAH. The endogenous ligand of Mincle receptor SAP130 up-regulated the expression of p-Syk and CARD9, and increased the levels of IL-1β and MPO, even though it did not increase brain water content nor it deteriorated neurological function at 24h after SAH. Syk inhibitor piceatannol reduced brain edema at 24h after SAH. Conclusion Mincle/Syk is involved in early brain injury after SAH, and they may serve as new targets for therapeutic intervention.

Background and Purpose Milk fat globule–EGF factor-8 (MFGE8) has been reported to be neuroprotective in ischemic stroke. However the effects of MFGE8 in early brain injury after subarachnoid hemorrhage (SAH) have not been investigated. We investigated the role of MFGE8 in early brain injury and the potential mechanisms in anti-oxidation after SAH. Methods Two dosages (1 µg and 3.3 µg) of recombinant human MFGE8 (rhMFGE8) were injected intracerebroventricularly at 1.5 hours after SAH. SAH grades, neurological scores and brain water content were measured at 24 hours and 72 hours. For mechanistic study, MFGE8 siRNA, integrin β3 siRNA and heme oxygenase (HO) inhibitor SnPP IX were used for intervention. The oxidative stress and expression of MFGE8, integrin β3, HO-1, ERK and Nrf2 were measured by western blots 24 hours after SAH. Results The expression of MFGE8 and HO-1 increased and peaked 24 hours after SAH. Administration of rhMFGE8 decreased brain water content and improved neurological functions both at 24 hours and 72 hours after SAH. rhMFGE8 reduced oxidative stress and enhanced the expression of ERK, Nrf2 and HO-1; and the effects were abolished by integrin β3 siRNA and HO inhibitor SnPP IX. Conclusion rMFGE8 attenuated oxidative stress which may be mediated by integrin β3/Nrf2/HO pathway after SAH. rMFGE8 may serve as an alternative treatment to ameliorate early brain injury for SAH patients.

Background and purpose Energy depletion is a critical factor leading to cell death and brain dysfunction after ischemic stroke. In this study we investigated whether energy depletion is involved in hyperglycemia-induced hemorrhagic transformation (HT) after ischemic stroke and determined the pathway underlying the beneficial effects of hyperbaric oxygen (HBO). Methods After 2 hours MCAO, hyperglycemia was induced by injecting of 50% dextrose (6 ml/kg) intraperitoneally at the onset of reperfusion. Immediately after it rats were exposed to HBO at 2 atmospheres absolutes (ATA) for 1 hour. ATP synthase inhibitor Oligomycin A (Olig A), NAMPT inhibitor FK866 or Sirt1 siRNA was administrated for interventions. Infarct volume, hemorrhagic volume, neurobehavioral deficits were recorded; the level of blood glucose, ATP and NAD+ and the activity of NAMPT were monitored; the expression of Sirt1, acetylated P53, acetylated NF-κB and cleaved caspase 3 were detected by western blots; the activity of MMP-9 was assayed by zymography. Results Hyperglycemia deteriorated energy metabolism and reduced the level of ATP and NAD+, exaggerated hemorrhagic transformation, blood-brain barrier disruption and neurological deficits after MCAO. HBO treatment increased the levels of the ATP and NAD+ and consequently increased Sirt1, resulting in attenuation of hemorrhagic transformation, brain infarction as well as improvement of neurological function in hyperglycemic MCAO rats. Conclusion HBO induced activation of ATP/NAD+/Sirt1 pathway and protected blood-brain barrier in hyperglycemic MCAO rats. HBO might be promising approach for treatment of acute ischemic stroke patients, especially patients with diabetes or treated with rtPA.

Apolipoprotein E (ApoE), encoded by the ApoE gene (APOE), influences the outcomes of traumatic brain injury (TBI), but the mechanism remains unclear. The present study aimed to investigate the effects of different ApoEs on the outcome of TBI and to explore the possible mechanisms. Controlled cortical impact (CCI) was performed on APOEε3 (E3) and APOEε4 (E4) transgenic mice, APOE-KO (KO) mice, and wild type (WT) mice to construct an in vivo TBI model. Neurological deficits, blood brain barrier (BBB) permeability and brain edema were detected at days 1, 3, and 7 after TBI. The results revealed no significant differences among the four groups at day 1 or day 3 after injury, but more severe deficits were found in E4 and KO mice than in E3 and WT mice. Furthermore, a significant loss of tight junction proteins was observed in E4 and KO mice compared with E3 and WT mice at day 7. Additionally, more expression and activation of NF-κB and MMP-9 were found in E4 mice compared with E3 mice. Different ApoEs had distinct effects on neuro-function and BBB integrity after TBI. ApoE3, but not E4, might inhibit the NF-κB/MMP-9 pathway to alleviate BBB disruption and improve TBI outcomes.

MMP-9 may be involved in early brain injury through degradation of laminin and neuronal death, and inhibition of MMP-9 may be a potential direction for brain protection after SAH.

Background Milk fat globule-epidermal growth factor-factor 8(MFGE8)/Integrin β3 pathway was reported to be involved in reducing oxidative stress and early brain injury after Subarachnoid Hemorrhage (SAH). In the present study, the potential effects of MFGE8 and its receptor Integrin β3 in the inhibition of apoptosis and neuroinflammation in early brain injury after SAH were investigated. Methods Ninety-five (95) male Sprague-Dawley rats were used. The SAH model was induced by endovascular perforation. Recombinant human MFGE8 (rhMFGE8), MFGE8 small interfering RNA (siRNA) and Integrin β3 siRNA were injected intracerebroventricularly. SAH grade, neurologic scores, Western blots and immunofluorescence were employed to study the mechanisms of MFGE8 and its receptor Integrin β3, as well as neurological outcome. Results SAH induced significant neuronal apoptosis and inflammation and exhibited neurological dysfunction in rats. Knockdown endogenous MFGE8 with siRNA significantly increased the protein levels of cleaved caspase 3 and IL-1β, accompanied with more neurological deficits. rhMFGE8 significantly reduced neural cell death in cortex, decreased cleaved caspase 3 and IL-1β expressions, and improved neurological functions 24 hours after SAH. The anti-apoptosis and anti-inflammation effects of rhMFGE8 were abolished by integrin-β3 siRNA. Conclusion MFGE8 could alleviate neurologic damage in early brain injury after SAH via anti-inflammation and anti-apoptosis effects. MFGE8 may serve as a promising therapeutic target for future management of SAH patients.