ChemR23 signaling ameliorates brain injury via inhibiting NLRP3 inflammasome-mediated neuronal pyroptosis in ischemic stroke.

BACKGROUND: Inflammatory response has been recognized as a pivotal pathophysiological process during cerebral ischemia. ChemR23 signaling is involved in the pathophysiology of various inflammatory diseases. Nevertheless, the role of ChemR23 signaling in ischemic stroke remains largely unknown. METHODS: Permanent ischemic stroke mouse model was accomplished by middle cerebral artery occlusion (MCAO). Resolvin E1 (RvE1) or chemerin-9 (C-9), the agonists of ChemR23, were administered by intracerebroventricular (i.c.v) injection before MCAO induction. Then, analysis of neurobehavioral deficits and brain sampling were done at Day 1 after MCAO. The brain samples were further analyzed by histological staining, immunofluorescence, RNA sequencing, ELISA, transmission electron microscope, and western blots. Furthermore, oxygen-glucose deprivation (OGD) was employed in SH-SY5Y to mimic MCAO in vitro, and ChemR23 signaling pathway was further studied by overexpression of ChemR23 or administration of related agonists or antagonists. Analysis of cell death and related pathway markers were performed. RESULTS: ChemR23 expression was upregulated following MCAO. Under in vitro and in vivo ischemic conditions, ChemR23 deficiency or inhibition contributed to excessive NLRP3-mediated maturation and release of IL-1ß and IL-18, as well as enhanced cleavage of GSDMD-N and neuronal pyroptosis. These influences ultimately aggravated brain injury and neuronal damage. On the other hand, ChemR23 activation by RvE1 or C-9 mitigated the above pathophysiological abnormalities in vivo and in vitro, and overexpression of ChemR23 in SH-SY5Y cells also rescued OGD-induced neuronal pyroptosis. Blockade of NLRP3 mimics the protective effects of ChemR23 activation in vitro. CONCLUSION: Our data indicated that ChemR23 modulates NLRP3 inflammasome-mediated neuronal pyroptosis in ischemic stroke. Activation of ChemR23 may serve as a promising potential target for neuroprotection in cerebral ischemia.

ChemR23 activation attenuates cognitive impairment in chronic cerebral hypoperfusion by inhibiting NLRP3 inflammasome-induced neuronal pyroptosis.

Neuroinflammation plays critical roles in vascular dementia (VaD), the second leading cause of dementia, which can be induced by chronic cerebral hypoperfusion (CCH). NLRP3 inflammasome-induced pyroptosis, the inflammatory programmed cell death, has been reported to contribute to the development of VaD. ChemR23 is a G protein-coupled receptor that has emerging roles in regulating inflammation. However, the role of ChemR23 signalling in NLRP3 inflammasome-induced pyroptosis in CCH remains elusive. In this study, a CCH rat model was established by permanent bilateral common carotid artery occlusion (BCCAO) surgery. Eight weeks after the surgery, the rats were intraperitoneally injected with the ChemR23 agonist Resolvin E1 (RvE1) or chemerin-9 (C-9). Additionally, primary rat hippocampal neurons and SH-SY5Y cells were adopted to mimic CCH injury in vitro. Our results showed that the levels of ChemR23 expression were decreased from the 8th week after BCCAO, accompanied by significant cognitive impairment. Further analysis revealed that CCH induced neuronal damage, synaptic injury and NLRP3-related pyroptosis activation in hippocampal neurons. However, pharmacologic activation of ChemR23 with RvE1 or C-9 counteracted these changes. In vitro experiments also showed that ChemR23 activation prevented primary neuron pyroptosis induced by chronic hypoxia. In addition, manipulating ChemR23 expression markedly regulated NLRP3 inflammasome-induced neuronal pyroptosis through PI3K/AKT/Nrf2 signalling in SH-SY5Y cells under hypoglycaemic and hypoxic conditions. Collectively, our data demonstrated that ChemR23 activation inhibits NLRP3 inflammasome-induced neuronal pyroptosis and improves cognitive function via the PI3K/AKT/Nrf2 signalling pathway in CCH models. ChemR23 may serve as a potential novel therapeutic target to treat CCH-induced cognitive impairment.

Platelet-derived extracellular vesicles promote endothelial dysfunction in sepsis by enhancing neutrophil extracellular traps.

BACKGROUND: The role of platelet-derived extracellular vesicles (PEVs) in the development of sepsis was investigated in this study. METHODS: After collection of blood samples from sepsis patients and normal volunteers, the extracellular vesicles (EVs) were separated, followed by the isolation of PEVs from the blood of rats. Next, a sepsis rat model was constructed by cecal ligation and puncture (CLP), and rats received tail vein injection of PEVs to explore the role of PEVs in sepsis. Subsequently, nanoparticle tracking analysis (NTA) and transmission electron microscopy (TEM) were adopted to determine the diameter of EVs and observe the morphology of PEVs, respectively; flow cytometry to detect the percentage of CD41-and CD61-positive EVs in isolated EVs; and ELISA to assess neutrophil extracellular trap (NET) formation, endothelial function injury-related markers in clinical samples or rat blood and serum inflammatory factor level. RESULTS: Compared with normal volunteers, the percentage of CD41- and CD61-positive EVs and the number of EVs were significantly elevated in sepsis patients. Moreover, sepsis patients also presented notably increased histone H3, myeloperoxidase (MPO), angiopoietin-2 and endocan levels in the blood, and such increase was positively correlated with the number of EVs. Also, animal experiments demonstrated that PEVs significantly promoted NET formation, mainly manifested as up-regulation of histone H3, high mobility group protein B1 (HMGB1), and MPO; promoted endothelial dysfunction (up-regulation of angiopoietin-2, endocan, and syndecan-1); and stimulated inflammatory response (up-regulation of interleukin (IL) -1ß, IL-6, tumor necrosis factor (TNF)-α, and monocyte chemoattractant protein (MCP) -1) in the blood of sepsis rats. CONCLUSION: PEVs aggravate endothelial function injury and inflammatory response in sepsis by promoting NET formation.

Impact of high-sensitivity troponin elevation and dynamic changes on 90-day mortality in patients with acute ischemic stroke after mechanical thrombectomy: results from an observational cohort.

BACKGROUND: A study was undertaken to evaluate the impact of high-sensitivity cardiac troponin I (hs-cTnI) elevation and hs-cTnI dynamic changes on 90-day mortality in patients with acute ischemic stroke (AIS) treated with mechanical thrombectomy (MT). METHODS: Patients with AIS receiving MT were included in the study. Sixty hours after AIS onset, hs-cTnI levels were measured before and after MT to determine elevated and dynamic changes. Patients were stratified into either normal or hs-cTnI elevation groups according to the pre-MT hs-cTnI cut-off value of 0.03 ng/L. hs-cTnI dynamic changes were defined as an increase or decrease of more than 20% pre-MT and post-MT, and at least one hs-cTnI level >0.03 ng/L. Multivariate Cox regression models were used to investigate the association between hs-cTnI elevation, hs-cTnI dynamic changes, and 90-day mortality in patients with AIS after MT. RESULTS: A total of 423 patients with AIS after MT were included in our final analysis, of whom only 72 (17%) showed hs-cTnI elevation. Post-MT hs-cTnI retesting was performed in 354 patients, and 90 (25.4%) patients presented with hs-cTnI dynamic changes. 119 patients died within 90 days. After adjusting for potential confounding factors, the Cox regression model showed that patients with hs-cTnI dynamic changes, rather than hs-cTnI elevation, were associated with 90-day mortality (p<0.05). Compared with the hs-cTnI non-dynamic changes, these results showed that a statistical association was present between rising hs-cTnI dynamic changes and 90-day mortality (p>0.05). CONCLUSIONS: hs-cTnI dynamic changes, dominated by the rising pattern rather than hs-cTnI elevation, were independent factors associated with 90-day mortality in patients with AIS after MT, especially in elderly subjects.

Ubiquitin-binding domain in ABIN1 is critical for regulating cell death and inflammation during development.

ABIN1 is a polyubiquitin-binding protein known to regulate NF-κB activation and cell death signaling. Mutations in Abin1 can cause severe immune diseases in human, such as psoriasis, systemic lupus erythematosus, and systemic sclerosis. Here, we generated mice that disrupted the ubiquitin-binding domain of ABIN1 (Abin1UBD/UBD) died during later embryogenesis owing to TNFR1-mediated cell death, similar to Abin1-/- mice. Abin1UBD/UBD cells were rendered sensitive to TNF-α-induced apoptosis and necroptosis as the inhibition of ABIN1UBD and A20 recruitment to the TNF-RSC complex leads to attenuated RIPK1 deubiquitination. Accordingly, the embryonic lethality of Abin1UBD/UBD mice was rescued via crossing with RIPK1 kinase-dead mice (Ripk1K45A/K45A) or the co-deletion of Ripk3 and one allele of Fadd, but not by the loss of Ripk3 or Mlkl alone. Unexpectedly, Abin1UBD/UBD mice with the co-deletion of Ripk3 and both Fadd alleles died at E14.5. This death was caused by spontaneous RIPK1 ubiquitination-dependent multiple inflammatory cytokines over production and could be rescued by the co-deletion of Ripk1 or Tnfr1 combined with Ifnar. Collectively, these data demonstrate the importance of the ABIN1 UBD domain, which mediates the ABIN1-A20 axis, at limiting RIPK1 activation-dependent cell death during embryonic development. Furthermore, our findings reveal a previously unappreciated ubiquitin pathway that regulates RIPK1 ubiquitination by FADD/Casp8 to suppress spontaneous IKKε/TBK1 activation.

Network pharmacology-based strategy to investigate pharmacological mechanisms of Andrographolide for treatment of vascular cognitive impairment.

Vascular cognitive impairment (VCI) is the second most common form of dementia. Andrographolide (Andro) shows potential effects in anti-inflammation, anti-oxidative stress, and anti-apoptosis. We have obtained 48 potential genes related to the effect of Andro on VCI through network pharmacology analysis. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis were used to reveal significant enriched pathway of potential genes, and the mitogen-activated protein kinase (MAPK) pathway was screened out. To verify the results of network pharmacology, we tested the effects of Andro in VCI model induced by bilateral common carotid artery occlusion (BCCAO) surgery. The results showed that Andro treatment ameliorated the cognitive impairment induced by BCCAO. Immunohistochemistry study revealed that Andro could reduce neuronal damage and activation of microglia in the cortex and hippocampus in BCCAO rats. To test the MAPK pathway changes, we analyzed the expression of JNK, p38 and ERK and found that Andro reduced the levels of phosphorylated-ERK (p-ERK) and phosphorylated-p38 (p-p38) in BCCAO rats. In conclusion, Andro could improve neuronal survival, reduce neuroinflammation and ameliorate cognitive impairment in VCI. The underlying mechanisms of Andro treatment may be through the inhibition of MAPK pathway.

Caspase-8 auto-cleavage regulates programmed cell death and collaborates with RIPK3/MLKL to prevent lymphopenia.

Caspase-8 is an initiator of death receptor-induced apoptosis and an inhibitor of RIPK3-MLKL-dependent necroptosis. In addition, caspase-8 has been implicated in diseases such as lymphoproliferation, immunodeficiency, and autoimmunity in humans. Although auto-cleavage is indispensable for caspase-8 activation, its physiological functions remain poorly understood. Here, we generated a caspase-8 mutant lacking E385 in auto-cleavage site knock-in mouse (Casp8ΔE385/ΔE385). Casp8ΔE385/ΔE385 cells were expectedly resistant to Fas-induced apoptosis, however, Casp8ΔE385/ΔE385 cells could switch TNF-α-induced apoptosis to necroptosis by attenuating RIPK1 cleavage. More importantly, CASP8(ΔE385) sensitized cells to RIPK3-MLKL-dependent necroptosis through promoting complex II formation and RIPK1-RIPK3 activation. Notably, Casp8ΔE385/ΔE385Ripk3-/- mice partially rescued the perinatal death of Ripk1-/- mice by blocking apoptosis and necroptosis. In contrast to the Casp8-/-Ripk3-/- and Casp8-/-Mlkl-/- mice appearing autoimmune lymphoproliferative syndrome (ALPS), both Casp8ΔE385/ΔE385Ripk3-/- and Casp8ΔE385/ΔE385Mlkl-/- mice developed transplantable lymphopenia that could be significantly reversed by RIPK1 heterozygosity, but not by RIPK1 kinase dead mutation. Collectively, these results demonstrate previously unappreciated roles for caspase-8 auto-cleavage in regulating necroptosis and maintaining lymphocytes homeostasis.

Rehabilitation of Sepsis Patients with Acute Kidney Injury Based on Intelligent Medical Big Data.

The objective of this study was to explore rehabilitation of patients with acute kidney injury (AKI) treated with Xuebijing injection by using intelligent medical big data analysis system. Based on Hadoop distributed processing technology, this study designed a medical big data analysis system and tested its performance. Then, this analysis system was used to systematically analyze rehabilitation of sepsis patients with AKI treated with Xuebijing injection. It is found that the computing time of this system does not increase obviously with the increase of cases. The results of systematic analysis showed that the glomerular filtration rate (59.31 ± 3.87% vs 44.53 ± 3.53%) in the experimental group was obviously superior than that in the controls after one week of treatment. The levels of urea nitrogen (9.32 ± 2.21 mmol/L vs. 14.32 ± 0.98 mmol/L), cystatin C (1.65 ± 0.22 mg/L vs. 2.02 ± 0.13 mg/L), renal function recovery time (6.12 ± 1.66 days vs. 8.66 ± 1.17 days), acute physiology and chronic health evaluation system score (8.98 ± 2.12 points vs. 12.45 ± 2.56 points), sequential organ failure score (7.22 ± 0.86 points vs. 8.61 ± 0.97 points), traditional Chinese medicine (TCM) syndrome score (6.89 ± 1.11 points vs. 11.33 ± 1.23 points), and ICU time (16.43 ± 2.37 days vs. 12.15 ± 2.56 days) in the experimental group were obviously lower than those in the controls, and the distinctions had statistical significance (P < 0.05). The significant efficiency (37.19% vs. 25.31%) and total effective rate (89.06% vs. 79.06%) in the experimental group were obviously superior than those in the controls, and distinction had statistical significance (P < 0.05). In summary, the medical big data analysis system constructed in this study has high efficiency. Xuebijing injection can improve the renal function of sepsis patients with kidney injury, and its therapeutic effect is obviously better than that of Western medicine, and it has clinical application and promotion value.

Predicting the Risk of Acute Kidney Injury in Patients After Percutaneous Coronary Intervention (PCI) or Cardiopulmonary Bypass (CPB) Surgery: Development and Assessment of a Nomogram Prediction Model.

BACKGROUND We sought to create a model that incorporated ultrasound examinations to predict the risk of acute kidney injury (AKI) after percutaneous coronary intervention (PCI) or cardiopulmonary bypass (CPB) surgery. MATERIAL AND METHODS A total of 292 patients with AKI after PCI or CPB surgery were enrolled for the study. Afterwards, treatment-related information, including data pertaining to ultrasound examination, was collected. A random forest model and multivariate logistic regression analysis were then used to establish a predictive model for the risk of AKI. Finally, the predictive quality and clinical utility of the model were assessed using calibration plots, receiver-operating characteristic curve, C-index, and decision curve analysis. RESULTS Predictive factors were screened and the model was established with a C-index of 0.955 in the overall sample set. Additionally, an area under the curve of 0.967 was obtained in the training group. Moreover, decision curve analysis also revealed that the prediction model had good clinical applicability. CONCLUSIONS The prediction model was efficient in predicting the risk of AKI by incorporating ultrasound examinations and a number of factors. Such included operation methods, age, congestive heart failure, body mass index, heart rate, white blood cell count, platelet count, hemoglobin, uric acid, and peak intensity (kidney cortex as well as kidney medulla).

PTEN inhibition enhances angiogenesis in an in vitro model of ischemic injury by promoting Akt phosphorylation and subsequent hypoxia inducible factor-1α upregulation.

Angiogenesis is an important pathophysiological response to cerebral ischemia. PTEN is a lipid phosphatase whose loss activates PI3K/Akt signaling, which is related to HIF-1α upregulation and enhanced angiogenesis in human cancer cells. However, the specific roles of PTEN in endothelial cell functions and angiogenesis after cerebral ischemia remain unknown. Therefore, we sought to examine the potential effects of PTEN inhibition on post-ischemic angiogenesis in human blood vessel cells and to determine the underlying mechanism. In this present study, human umbilical vein endothelial cells (HUVECs) were exposed to oxygen-glucose deprivation (OGD), cell proliferation, migration and apoptosis, in vitro tube formation and expression of PTEN/Akt pathway and angiogenic factors were examined in HUVECs after treatment with PTEN inhibitor bisperoxovanadium (bpV) at different doses. The results showed that bpV significantly increased the cell proliferation and reduced cell apoptosis indicating that the drug exerts a cytoprotective effect on HUVECs with OGD exposure. bpV also enhanced cell migration and tube formation in HUVECs following OGD, and upregulated HIF-1α and VEGF expressions, but attenuated endostatin expression. Additionally, western blotting analysis demonstrated that Akt phosphorylation in HUVECs was significantly increased after bpV treatment. These findings suggest that PTEN inhibition promotes post-ischemic angiogenesis in HUVECs after exposure to OGD and this enhancing effect might be achieved through activation of the Akt signal cascade.

Aquaporin-4 deletion ameliorates hypoglycemia-induced BBB permeability by inhibiting inflammatory responses.

BACKGROUND: Severe hypoglycemia induces brain edema by upregulating aquaporin-4 (AQP4) expression and by degrading tight junctions. Acute severe hypoglycemia induces a proinflammatory environment that may contribute to a disruption in the epithelial barrier by decreasing tight junction protein expression. Interestingly, the altered AQP4 expression has been considered to play a critical role in neuroinflammation during acute brain injury. It has been shown that AQP4 deletion reduces brain inflammation in AQP4-null mice after intracerebral LPS injection. However, the effect of AQP4 deletion regarding protection against hypoglycemia-induced blood-brain barrier (BBB) breakdown is unknown. METHODS: An acute severe hypoglycemic stress model was established via injection of 4 unit/kg body weight of insulin. Evans blue (EB) staining and water measurement were used to assess BBB permeability. Western blot, reverse transcription polymerase chain reaction, and immunofluorescence were used to detect the expression of related proteins. The production of cytokines was assessed via enzyme-linked immunosorbent assay. RESULTS: Hypoglycemia-induced brain edema and BBB leakage were reduced in AQP4-/- mice. AQP4 deletion upregulated PPAR-γ and inhibited proinflammatory responses. Moreover, knockdown of aquaporin-4 by small interfering RNA in astrocytes co-cultured with endothelial cells effectively reduced transendothelial permeability and degradation of tight junctions. Treatment with PPAR-γ inhibitors showed that upregulation of PPAR-γ was responsible for the protective effect of AQP4 deletion under hypoglycemic conditions. CONCLUSIONS: Our data suggest that AQP4 deletion protects BBB integrity by reducing inflammatory responses due to the upregulation of PPAR-γ expression and attenuation of proinflammatory cytokine release. Reduction in AQP4 may be protective in acute severe hypoglycemia.

Pro-Resolving Lipid Mediators Improve Neuronal Survival and Increase Aß42 Phagocytosis.

Inflammation in the brain is a prominent feature in Alzheimer's disease (AD). Recent studies suggest that chronic inflammation can be a consequence of failure to resolve the inflammation. Resolution of inflammation is mediated by a family of lipid mediators (LMs), and the levels of these specialized pro-resolving mediators (SPMs) are reduced in the hippocampus of those with AD. In the present study, we combined analysis of LMs in the entorhinal cortex (ENT) from AD patients with in vitro analysis of their direct effects on neurons and microglia. We probed ENT, an area affected early in AD pathogenesis, by liquid chromatography-tandem mass spectrometry (LC-MS-MS), and found that the levels of the SPMs maresin 1 (MaR1), protectin D1 (PD1), and resolvin (Rv) D5, were lower in ENT of AD patients as compared to age-matched controls, while levels of the pro-inflammatory prostaglandin D2 (PGD2) were higher in AD. In vitro studies showed that lipoxin A4 (LXA4), MaR1, resolvin D1 (RvD1), and protectin DX (PDX) exerted neuroprotective activity, and that MaR1 and RvD1 down-regulated ß-amyloid (Aß)42-induced inflammation in human microglia. MaR1 exerted a stimulatory effect on microglial uptake of Aß42. Our findings give further evidence for a disturbance of the resolution pathway in AD, and indicate that stimulating this pathway is a promising treatment strategy for AD.

Chronic airway-induced allergy in mice modifies gene expression in the brain toward insulin resistance and inflammatory responses.

BACKGROUND: Chronic systemic inflammation affects brain functionality and may negatively influence the progression of neurodegenerative disorders. Allergy is a chronic inflammatory disease affecting more than 20% of the Western population. Little is known regarding the influence of allergy on brain functions. The aim of the present study was to obtain a global overview of the genes that drive the effects of peripheral inflammation associated with chronic airway-induced allergy in the brain. METHODS: Airway allergy was induced in C57B/6J mice using ovalbumin as the allergen. Microarray analysis was performed in the hippocampus and frontal cortex in association with Affymetrix. For the data analysis, principal component analysis and orthogonal to latent structures discriminant analysis followed by pathway analysis were used. Quantitative polymerase chain reaction (qPCR) and protein analysis by Western blotting were performed for the validation of microarray results. RESULTS: Microarray analysis showed low-grade changes in gene expression in the brain induced by airway-associated allergy. Changes in expression were observed for genes involved in antigen processing and presentation, cytokine-cytokine interaction, Toll-like receptor and mitogen-activated protein kinase signaling, as determined by pathway analysis. We confirmed a reduction of insulin-degrading enzyme at the protein level and a decrease in insulin receptor phosphorylation in the brains of allergic mice. Other allergy-induced gene expression changes were confirmed by qPCR, including increased levels of tumor necrosis factor receptor superfamily member 23 and lipopolysaccharide-binding protein. CONCLUSION: Airway-associated allergy induces changes in brain gene expression toward induction of insulin resistance and inflammatory responses with potential implications for neurodegenerative disorders.