Epilepsy is increasingly linked to cellular senescence and the senescence-associated secretory phenotype (SASP), which propagates sterile neuroinflammation and tissue remodeling. However, the specific SASP-related molecular signatures driving epileptogenesis remain poorly characterized. Integrating human cortical transcriptomic profiles (GSE256068) with a curated SASP gene set, we identified 22 differentially expressed candidates. Machine-learning feature selection (LASSO and SVM-RFE) converged on a robust five-biomarker panel (IGFBP4, SERPINE1, CCL2, C3, and SPX), which demonstrated consistent differential expression in an independent, cross-regional hippocampal dataset (GSE134697). A diagnostic nomogram integrating these markers achieved excellent discrimination and clinical net benefit. Functional enrichment linked this panel to oxidative phosphorylation and translational control. Crucially, the up-regulation of SERPINE1 and CCL2 highlights a neurochemical mechanism involving extracellular matrix remodeling and inferred neuroimmune microenvironment shifts. In silico regulatory network analysis further predicted upstream transcription factors governing this axis. Finally, RT-qPCR validation in the peripheral blood of a kainic acid-induced epilepsy mouse model confirmed the significant dysregulation of four key biomarkers (IGFBP4, SERPINE1, CCL2, and C3). Collectively, our study defines a highly accurate SASP-associated biomarker panel and provides profound neurochemical insights into senescence-linked inflammatory networks, offering novel targets for the diagnosis and management of epilepsy.

Traumatic brain injury (TBI) is a severe neurological condition associated with complex pathological cascades. Transforming growth factor-β2 (TGF-β2) is a pleiotropic cytokine that participates in multiple cellular events in the central nervous system; however, its potential protective role and molecular mechanisms in the context of TBI-induced neuronal injury have not been fully elucidated. In this study, we observed that TGF-β2 expression was markedly increased following TBI, with peak expression at day 7 post-injury. Using an in vitro LPS-induced inflammatory model, we found that exogenous TGF-β2 treatment significantly reduced the expression of pro-inflammatory factors TNF-α, IL-1β, and IL-6, as assessed by qRT-PCR, Western blot, and ELISA. Co-culture of neurons with conditioned medium from LPS-stimulated astrocytes revealed that TGF-β2 overexpression indirectly attenuated neuronal apoptosis and enhanced neuronal viability, as demonstrated by CCK-8, TUNEL staining, and molecular analyses. Mechanistic investigations suggested that the anti-inflammatory and anti-apoptotic effects of TGF-β2 may be mediated through activation of the Smad3 signaling pathway. Consistently, in vivo experiments indicated that TGF-β2 overexpression alleviated motor dysfunction and reduced neuronal apoptosis in a TBI mouse model. Collectively, these findings provide evidence for a protective role of TGF-β2 in inflammatory neuronal injury and offer insights into potential molecular targets for TBI therapeutic development.

Cancer is a major global cause of mortality, underscoring the urgent need to decode its fundamental biology and advance targeted therapeutic strategies. The Hippo pathway, a highly conserved signaling cascade, plays indispensable roles in regulating organ size, suppressing tumorigenesis and sustaining stem cell self-renewal. Within tumors, a subpopulation of cancer stem cells (CSCs) possesses the dual capacity for self-renewal and differentiate, thereby driving tumor growth and heterogeneity. Notably, CSC-like cells can be produced by the aberrant activation of epithelial-mesenchymal transition (EMT), a de-differentiation program promoting tumor progression. Emerging evidence indicates that dysregulation of Hippo pathway signaling leads to tumor progression by enhancing CSC characteristics and facilitating EMT. The Hippo pathway also has tissue specific regulatory patterns and plays context dependent dual roles in the tumor microenvironment, either as a tumor suppressor or as an oncogenic driver. Whereas early-stage clinical trials targeting the Hippo pathway have both confirmed the therapeutic potential and revealed major challenges associated with on-target specificity and safety. This review synthesizes recent breakthroughs in our understanding of the mechanistic links between Hippo signaling, CSCs and tumor progression, offering novel perspectives on potential therapeutic vulnerabilities.

PURPOSE:This study aimed to investigate the clinical characteristics and identify risk factors associated with postoperative seizure outcomes in pediatric patients with low-grade epilepsy-associated brain tumors (LEATs) who underwent gross total resection (GTR). METHODS:We retrospectively analyzed the clinical data of pediatric patients who underwent GTR of LEATs. Descriptive statistics were used to summarize the clinical features, and patients were categorized into two groups based on postoperative seizure outcomes: seizure-free and non-seizure-free. Binary logistic regression analysis was performed to identify independent risk factors for failure to achieve seizure freedom. RESULTS:A total of 75 pediatric patients with LEATs who underwent GTR were included in the study. Among them, 58 (77.3 %) achieved seizure freedom, while 17 (22.7 %) remained non-seizure-free postoperatively. Binary logistic regression analysis identified longer epilepsy duration (OR = 1.03, p = 0.034), the number of antiepileptic drugs (AEDs) (OR = 10.90, p = 0.005), and non-temporal lobe tumor location (OR = 6.02, p = 0.028) as significant independent risk factors for persistent postoperative seizures. CONCLUSIONS:Our findings suggest that even after gross total resection, a prolonged history of epilepsy, extratemporal tumor location, and polytherapy with AEDs are associated with a lower likelihood of achieving seizure freedom. Early surgical intervention and comprehensive preoperative assessment may help optimize postoperative seizure outcomes in pediatric patients with LEATs.

Focal cortical dysplasia (FCD) is a structural lesion that is the most common anatomical lesion identified in children, and the second most common in adults with drug-resistant focal-onset epilepsy. These lesions vary in size, location, and histopathological manifestations. FCDs are classified into three subtypes associated with loss-of-function mutations in PI3K/AKT, TSC1/TSC2, RHEB, and DEPDC/NPRL2/NPRL3. During the decades of research into FCD, experimental models have played an irreplaceable role in the research design of studies investigating disease pathogenesis, pathophysiology, and treatment. Further, the establishment of FCD experimental models has moved the field forward by (1) revealing the cellular processes and signaling pathways underlying FCD pathogenesis and (2) varying the methods and materials to study the function of FCD proteins. Currently, FCD experimental models are predominantly murine, with each model providing unique insights into FCD lesions. This review briefly summarizes the pathology and molecular functions of FCD, further comparing the available modeling methods and indexes, as well as the utilization of models, followed by an analysis of the similarities, advantages, and disadvantages between these models and human FCD.

Rhabdomyolysis (RM) leads to dysfunction in the core organs of kidney, lung and heart, which is an important reason for the high mortality and disability rate of this disease. However, there is a lack of systematic research on the characteristics of rhabdomyolysis-induced injury in various organs and the underlying pathogenetic mechanisms, and especially the interaction between organs. We established a rhabdomyolysis model, observed the structural and functional changes in kidney, heart, and lung. It is observed that rhabdomyolysis results in significant damage in kidney, lung and heart of rats, among which the pathological damage of kidney and lung was significant, and of heart was relatively light. Meanwhile, we analyzed the differentially expressed proteins (DEPs) in the kidney, heart and lung between the RM group and the sham group based on liquid chromatography-tandem mass spectrometry (LC-MS/MS). In our study, Serpina3n was significantly up-regulated in the kidney, heart and lung. Serpina3n is a secreted protein and specifically inhibits a variety of proteases and participates in multiple physiological processes such as complement activation, inflammatory responses, apoptosis pathways, and extracellular matrix metabolism. It is inferred that Serpina3n may play an important role in multiple organ damage caused by rhabdomyolysis and could be used as a potential biomarker. This study comprehensively describes the functional and structural changes of kidney, heart and lung in rats after rhabdomyolysis, analyzes the DEPs of kidney, heart and lung, and determines the key role of Serpina3n in multiple organ injury caused by rhabdomyolysis. SIGNIFICANCE: This study comprehensively describes the functional and structural changes of kidney, heart and lung in rats after rhabdomyolysis, analyzes the DEPs of kidney, heart and lung, and determines the key role of Serpina3n in multiple organ injury caused by rhabdomyolysis.

Acute kidney injury (AKI) can cause distal cardiac dysfunction; however, the underlying mechanism is unknown. Oxidative stress is proved prominent in AKI-induced cardiac dysfunction, and a possible bridge role of oxidative-stress products in cardio-renal interaction has been reported. Therefore, this study aimed to investigate the critical role of circulating reactive oxygen species (ROS) in mediating cardiac dysfunction after bilateral renal ischemia-reperfusion injury (IRI). We observed the diastolic dysfunction in the mice following renal IRI, accompanied by reduced ATP levels, oxidative stress, and branched-chain amino acids (BCAA) accumulation in the heart. Notably, ROS levels showed a sequential increase in the kidneys, circulation, and heart. Treatment with tempol, an ROS scavenger, significantly restored cardiac diastolic function in the renal IRI mice, corroborating the bridge role of circulating ROS. Accumulating evidence has identified oxidative stress as upstream of Mst1/Hippo in cardiac injury, which could regulate the expression of downstream genes related to mitochondrial quality control, leading to lower ATP, higher ROS and metabolic disorder. To verify this, we examined the activation of the Mst1/Hippo pathway in the heart of renal IRI mice, which was alleviated by tempol treatment as well. In vitro, analysis revealed that Mst1-knockdown cardiomyocytes could be activated by hydrogen peroxide (H2O2). Analysis of Mst1-overexpression cardiomyocytes confirmed the critical role of the Mst1/Hippo pathway in oxidative stress and BCAA dysmetabolism. Therefore, our results indicated that circulating ROS following renal IRI activates the Mst1/Hippo pathway of myocardium, leading to cardiac oxidative stress and diastolic dysfunction. This finding provides new insights for the clinical exploration of improved treatment options for cardiorenal syndrome.

Intracerebral hemorrhage (ICH) is a severe cerebrovascular disease with a high disability rate and high mortality, and pyroptosis is a type of programmed cell death in the acute phase of ICH. Neuronal Per-Arnt-Sim domain protein 4 (Npas4) is a specific transcription factor highly expressed in the nervous system, yet the role of NPAS4 in ICH-induced pyroptosis is not fully understood. NLR family Pyrin-domain-containing 6 (NLRP6), a new member of the Nod-like receptor family, aggravates pyroptosis via activating cysteine protease-1 (Caspase-1) and Caspase-11. In this study, we found that NPAS4 was upregulated in human and mouse peri-hematoma brain tissues and peaked at approximately 24 h after ICH modeling. Additionally, NPAS4 knockdown improved neurologic dysfunction and brain damage induced by ICH in mice after 24 h. Meanwhile, inhibiting NPAS4 expression reduced the levels of myeloperoxidase (MPO)-positive cells and Caspase-1/TUNEL-double-positive cells and decreased cleaved Caspase-1, cleaved Caspase-11, and N-terminal GSDMD levels. Consistently, NPAS4 overexpression reversed the above alternations after ICH in the mice. Moreover, NPAS4 could interact with the Nlrp6 promoter region (−400–−391 bp and −33–−24 bp) and activate the transcription of Nlrp6. Altogether, our study demonstrated that NPAS4, as a transcription factor, can exacerbate pyroptosis and transcriptionally activate NLRP6 in the acute phase of intracerebral hemorrhage in mice.

Background:Epilepsy is a chronic disease that is characterized by transient brain dysfunction caused by an abrupt abnormal neuronal discharge. Recent studies have indicated that the pathways related to inflammation and innate immunity play significant roles in the pathogenesis of epilepsy, suggesting an interrelationship between immunity and inflammatory processes and epilepsy. However, the immune-related mechanisms are still not precisely understood; therefore, this study aimed to explore the immune-related mechanisms in epilepsy disorders, highlight the role of immune cells at the molecular level in epilepsy, and provide therapeutic targets for patients with epilepsy.Methods:Brain tissue samples from healthy and epileptic individuals were collected for transcriptome sequencing to identify differentially expressed genes (DEGs) and differentially expressed (DE)-long coding RNAs (lncRNAs). Based on interactions from the miRcode, starBase2.0, miRDB, miRTarBase, TargetScan, and ENCORI databases, a lncRNA-associated competitive endogenous RNA (ceRNA) network was created. Gene ontology and the Kyoto encyclopedia of genes analyses established that the genes in the ceRNA network were mainly enriched in immune-related pathways. Immune cell infiltration, screening, and protein-protein interaction analyses of the immune-related ceRNAs, and correlation analysis between immune-related core messenger RNA (mRNA) and immune cells were also performed.Results:Nine hub genes (EGFR, GRB2, KRAS, FOS, ESR1, MAPK1, MAPK14, MAPK8, and PPARG) were obtained. Also, 38 lncRNAs, one miRNA (hsa-miR-27a-3p), and one mRNA (EGFR) comprised the final core ceRNA network. Mast cells, plasmacytoid dendritic cells, and immature dendritic cells all showed positive correlations with EGFR, while Cluster of differentiation 56 dim natural killer cells (CD56dim natural killer cells) showed negative correlations. Finally, we employed an epilepsy mouse model to validate EGFR, which is consistent with disease progression.Conclusions:In conclusion, the pathophysiology of epilepsy was correlated with EGFR. Thus, EGFR could be a novel biomarker of juvenile focal epilepsies, and our findings provide promising therapeutic targets for epilepsy.

Several studies have shown a link between immunity, inflammatory processes, and epilepsy. Active neuroinflammation and marked immune cell infiltration occur in epilepsy of diverse etiologies. Microglia, as the first line of defense in the central nervous system, are the main effectors of neuroinflammatory processes. Discovery of new biomarkers associated with microglia activation after epileptogenesis indicates that targeting specific molecules may help control seizures. In this research, we used a combination of several bioinformatics approaches, including RNA sequencing, to explore differentially expressed genes (DEGs) in epileptic lesions and control samples, and to construct a protein-protein interaction (PPI) network for DEGs, which was examined utilizing plug-ins in Cytoscape software. Finally, we aimed to identify 10 hub genes in immune and inflammation-related sub-networks, which were subsequently validated in real-time quantitative polymerase chain reaction analysis in a mouse model of kainic acid-induced epilepsy. The expression patterns of nine genes were consistent with sequencing outcomes. Meanwhile, several genes, including CX3CR1, CX3CL1, GPR183, FPR1, P2RY13, P2RY12 and LPAR5, were associated with microglial activation and migration, providing novel candidate targets for immunotherapy in epilepsy and laying the foundation for further research.

Glioblastoma multiforme (GBM) is a lethal primary brain tumor with poor survival lifespan and dismal outcome. However, the effects and mechanisms of epigenetic factors on the development of GBM were still not well illustrated. We found that expression of enhancer of zeste homolog 2 (EZH2), which can catalyze histone H3K27me3 to modulate gene expression, was increased in GBM cells. Knockdown of EZH2 can suppress proliferation and migration, while increase temozolomide (TMZ) sensitivity, of GBM cells. Further, knockdown of EZH2 or its specific inhibitor GSK126 can decrease expression of Twist, while over expression of Twist can reverse si-EZH2-suppressed malignancy of GBM cells. Mechanistically, EZH2 can positively regulate mRNA stability of Twist1 mRNA. Further, miR-206, which can bind with 3 ' UTR of Twist1 mRNA, was involved in EZH2regulated mRNA stability of Twist1. Collectively, our data suggest that EZH2 might be a potential target for GBM treatment. Further, miR-206/Twist axis is involved in EZH2-regulated malignancy of GBM cells.

The NLRC4 inflammasome, a member of the nucleotide-binding and oligomerization domain-like receptor (NLR) family, amplifies inflammation by facilitating the processing of caspase-1, interleukin (IL)–1β, and IL-18. We explored whether NLRC4 knockdown alleviated inflammatory injury following intracerebral hemorrhage (ICH). Furthermore, we investigated whether NLRC4 inflammasome activation can be adjusted by the regulator of G protein signaling 2/leucine-rich repeat kinase-2 pathway. Fifty microliters of arterial blood was drawn and injected into the basal ganglion to simulate the ICH model. NLRC4 small interfering RNAs (siRNAs) were utilized to knockdown NLRC4. An LRRK2 inhibitor (GNE7915) was injected into the abdominal cavity. Short hairpin (sh) RNA lentiviruses and lentiviruses containing RGS2 were designed and applied to knockdown and promote RGS2 expression. Neurological functions, brain edema, Western blot, enzyme-linked immunosorbent, hematoxylin and eosin staining, Nissl staining, immunoprecipitation, immunofluorescence assay and Evans blue dye extravasation and autofluorescence assay were evaluated. It was shown that the NLRC4 inflammasome was activated following ICH injury. NLRC4 knockdown extenuated neuronal death, damage to the blood-brain barrier, brain edema and neurological deficiency 3 days after ICH. NLRC4 knockdown reduced myeloperoxidase (MPO) cells as well as tumor necrosis factor (TNF)-α, interleukin (IL)-6, IL-1β and IL-18 following ICH. GNE7915 reduced pNLRC4 and NLRC4 inflammasome activation. RGS2 suppressed the interaction of LRRK2 and NLRC4 and NLRC4 inflammasome activation by regulating pLRRK2. Our study demonstrated that the NLRC4 inflammasome may aggravate the inflammatory injury induced by ICH and that RGS2/LRRK2 may relieve inflammatory injury by restraining NLRC4 inflammasome activation.

Pyrin domain-containing 3 inflammasome (NLRP3), a member of the NOD-like receptor family, has a crucial role in the inflammatory process that occurs during intracerebral hemorrhage (ICH)-induced injury. Histone deacetylase 10 (HDAC10) is a newly identified class II histone deacetylase involved in immune responses. However, how HDAC10 affects the inflammatory response after ICH remains unknown. In this study, we investigated whether HDAC10 relieves ICH injury by suppressing NLRP3 inflammasome activation through the protein tyrosine phosphatase, nonreceptor type 22 (PTPN22) pathway. We induced ICH in Sprague-Dawley rats (healthy, male adult) with a single infusion of autologous blood. To knockdown HDAC10, we injected siRNA into the rats. To further explore the mechanisms underlying the role of HDAC10 in ICH injury, PTPN22 was silenced. HDAC10 levels were upregulated after ICH in humans and rats, and reached peak levels 24 h after ICH induction in rats. HDAC10 silencing aggravated ICH injury, as demonstrated by increased modified neurological severity scores, brain water content, Evans blue extravasation, and number of myeloperoxidase (MPO) cells, and the results of Nissl and H&E staining. Furthermore, HDAC10 knockdown increased the expression of PTPN22 and accentuated inflammatory responses mediated by the NLRP3 inflammasome. HDAC10 silencing increased NLRP3 inflammasome activation, and this was effectively reversed by PTPN22 knockdown using siRNA. Furthermore, HDAC10 silencing also promoted the interaction of PTPN22 and NLRP3. Our study demonstrated that HDAC10 silencing aggravated NLRP3-mediated inflammatory responses after ICH in rats via the PTPN22 pathway. These results suggest that regulating the NLRP3 inflammasome may be a novel method to ameliorate ICH injury.

The NOD-like receptor family Pyrin domain-containing 3 (NLRP3) inflammasome has a crucial role in the inflammatory process that occurs during intracerebral hemorrhage (ICH)-induced injury. Histone deacetylase 10 (HDAC10) is a newly identified class II histone deacetylase involved in immune responses. However, how HDAC10 affects the inflammatory response after ICH remains unknown. In this study, we investigated whether HDAC10 relieves ICH injury by suppressing NLRP3 inflammasome activation through the protein tyrosine phosphatase, nonreceptor type 22 (PTPN22) pathway. We induced ICH in Sprague-Dawley rats (healthy, male adult) with a single infusion of autologous blood. To knockdown HDAC10, we injected siRNA into the rats. To further explore the mechanisms underlying the role of HDAC10 in ICH injury, PTPN22 was silenced. HDAC10 levels were upregulated after ICH in humans and rats, and reached peak levels 24 h after ICH induction in rats. HDAC10 silencing aggravated ICH injury, as demonstrated by increased modified neurological severity scores, brain water content, Evans blue extravasation, and number of myeloperoxidase (MPO) cells, and the results of Nissl and H&E staining. Furthermore, HDAC10 knockdown increased the expression of PTPN22 and accentuated inflammatory responses mediated by the NLRP3 inflammasome. HDAC10 silencing increased NLRP3 inflammasome activation, and this was effectively reversed by PTPN22 knockdown using siRNA. Furthermore, HDAC10 silencing also promoted the interaction of PTPN22 and NLRP3. Our study demonstrated that HDAC10 silencing aggravated NLRP3-mediated inflammatory responses after ICH in rats via the PTPN22 pathway. These results suggest that regulating the NLRP3 inflammasome may be a novel method to ameliorate ICH injury. (C) 2020 The Author(s). Published by Elsevier Ltd on behalf of IBRO.

目的:探讨缺氧诱导因子2α(hypoxia-inducible factor 2α,HIF-2α)在脑出血损伤后(intracerebral hemorrhage,ICH)的作用,并明确其是否参与调控脑出血后的炎症反应.方法:健康雄性SD大鼠95只,其中35只随机分配用于以下时间点分析:Sham、12 h、24h、48 h、3d、5d和7d,每组5只;另外60只随机分为Sham组(假手术组,注射等量生理盐水)、ICH组(脑出血模型组组,胶原酶诱导的脑出血模型)、Vehicle组(空载体组组,建模前注射空载慢病毒载体)和Oe-HIF-2α组(HIF-2α过表达组,建模前注射HIF-2α过表达慢病毒载体)用于脑含水量检测、神经功能评分、免疫印记分析和免疫荧光检测.通过脑含水量和Garcia神经功能评分评估脑损伤的严重程度;免疫印迹检测HIF-2α及肿瘤坏死因子-α(tumor necrosis factor α,TNF-α)、白细胞介素-18(interleukin 18,IL-18)和白细胞介素-1β(interleukin 1β,IL-1β)的表达水平,免疫荧光检测病灶周边髓过氧化物酶(myeloperoxidase,MPO)的表达.结果:HIF-2α表达在ICH后24 h(0.555 4±0.070 2,P=0.000)开始上调并于3 d(2.368 4±0.346 6,P=0.000)达到峰值,随后降低;与ICH组相比较,Oe-HIF-2α组脑3d时含水量明显降低(0.793 5±0.002 5,P=0.000),神经功能明显改善(14.7000±0.674 9,P=0.000);与ICH组相比,Oe-HIF-2α组TNF-α(1.3504±0.191 5,P=0.000)、IL-1β(I.158 4±0.070 8,P=0.000)、IL-18(0.784 2±0.073 9,P=-0.000)等炎性介质在3d时表达明显降低;Oe-HIF-2α组(3.500 0±0.534 5)病灶周边区域MPO表达明显低于ICH组(5.125 0±0.991 0,P=0.002).结论:脑出血后,HIF-2α表达上调并且通过抑制TNF-α、IL-1β、IL-18等炎症因子的表达,抑制炎症反应减轻大鼠脑出血损伤.

Angiogenesis after intracerebral hemorrhage (ICH) injury can effectively alleviate brain damage and improve neurological function. Hypoxia-inducible factor 2α (HIF-2α) is an important angiogenic regulator and exhibits protective effects in several neurological diseases; however, its role in ICH has not yet been reported. Hence, in the present study, we explored whether HIF-2α reduces ICH injury by promoting angiogenesis. In addition, we explored the role of the vascular endothelial growth factor (VEGF)/Notch pathway in HIF-2α-mediated angiogenesis. We injected 50 μL of autologous blood taken from the femoral artery into the right striatum of healthy male adult Sprague-Dawley rats to create an autologous-blood-induced rat model of ICH. Lentiviral vectors were injected to both overexpress and knock down HIF-2α expression. VEGF receptor 2 (VEGFR2) and Notch-specific inhibitors were injected intraperitoneally to block VEGFR2- and Notch-mediated signaling after lentiviral injections. Our data showed that HIF-2α overexpression reduced neurological-damage scores and brain-water content, suggesting it had a protective effect on ICH injury. In addition, overexpression of HIF-2α promoted angiogenesis, increased focal cerebral blood flow (CBF), and reduced neuronal damage, whereas HIF-2α knockdown resulted in the opposite effects. Furthermore, we found that HIF-2α-mediated angiogenesis was blocked by a Notch-specific inhibitor. Likewise, the HIF-2α-mediated increase in phospho-VEGFR-2, cleaved-Notch1 and Notch1 expression was reversed via a VEGFR2-specific inhibitor. Taken together, our results indicate that HIF-2α promotes angiogenesis via the VEGF/Notch pathway to attenuate ICH injury. Moreover, our findings may contribute to the development of a novel strategy for alleviating ICH injury via HIF-2α-mediated upregulation of angiogenesis.

Objective To investigate the effects of naringenin (NGN) on inflammatory injury after intracerebral hemorrhage (ICH) in rats and its possible molecular mechanism. Methods SD rats were randomly divided into sham group, ICH group, ICH+DMSO+PEG400 group, ICH+low-dose NGN group (10 mg/kg), ICH+middle-dose NGN group (20 mg/kg), ICH+high-dose NGN group (40 mg/kg), ICH+DMSO group, ICH+phorbol 12-myristate 13-acetate (PMA, PKC-δ activator) group, and ICH+PMA+NGN group. Autologous blood ICH model of SD rat was established, and the rats from the corresponding groups were intraperitoneally injected with NGN. In 24 h after ICH, the rats were evaluated for neurological function score, and then the rats were sacrificed to measure brain water content and perform HE staining for pathological changes. Western blotting was used to detect protein kinase C (PKC)-δ phosphorylation and expression of pro-inflammatory cytokines in sham group, ICH group, ICH+DMSO+PEG400 group, and ICH+ middle-dose NGN group. Intraventricular injection of PMA was carried out to induce PKC-δ phosphorylation in rats, and then ICH model was established, the expression of pro-inflammatory cytokines in rat brain tissues was detected by Western blotting. Results Compared with the ICH group and the solvent control group, NGN improved the neurological score (P < 0.05), reduced brain water content (P < 0.05), attenuated neuronal degeneration and cell death, and decreased PKC-δ phosphorylation and the expression levels of NF-κB, interleukin (IL)-1β, IL-6 after ICH (P < 0.01). PMA treatment resulted in the enhanced phosphorylation level of PKC-δ and the up-regulation of NF-κB, IL-1β, and IL-6 (P < 0.01) and reversed the effect of NGN on the decreased expression of NF-κB, IL -1β and IL-6 in rats after ICH (P < 0.01). Conclusion NGN attenuates the inflammatory injury after ICH in rats, which may be related to its inhibitory effect on PKC-δ phosphorylation.

Background: Brain-derived neurotrophic factor (BDNF) is a neurotrophin that plays a pivotal role in neuronal development by modulating synaptic activity and regeneration. BDNF has two forms, the mature BDNF (mBDNF) and the precursor BDNF (proBDNF), which seem to act in opposite functional ways. The involvement of mBDNF/proBDNF ratio and its role after cerebral intracerebral hemorrhage (ICH) are not fully characterized yet. Methods: To address these issues, adenosine 5′-triphosphate disodium salt hydrate (ATP), a P2 purine agonist that evokes the release of mBDNF, was administrated to regulate mBDNF/proBDNF ratio. Data was evidenced by hematoma volume, neurological scores, edema or blood-brain barrier disruption, HE, Nissl staining and immunofluorescence. To investigate the regulation mechanism of mBDNF/proBDNF ratio by ATP, we used P2X4r LEN, P38-mitogen activated protein kinase (MAPK), Ca2+, and soluble N-ethylmaleimide-sensitive factor attachment receptors (SNAREs) inhibitor. Findings: ATP promoted mBDNF/proBDNF ratio and protected against ICH injury including a reduction in hematoma volume, an increase in neurological scores, alleviation of edema or blood-brain barrier disruption, and an improvement in outcomes of HE, Nissl staining, and immunofluorescence. In addition, the ratio of mBDNF/proBDNF was mediated by P2X4r primarily, and then regulated by ATP-induced activation of P38-MAPK, dependent upon the presence of extracellular and endoplasmic reticulum store Ca2+ and related to SNARE-mediated exocytosis. Interpretation: Our study demonstrates that the mBDNF/proBDNF ratio is able to alleviate ICH injury by ATP administration through P2X4 receptor/P38-MAPK/Ca2+/SNAREs regulation. These results may provide a new strategy to alleviate ICH injury by promoting the mBDNF/proBDNF ratio in neurotrophy.Funding Statement: This work was funded by the National Natural Science Foundation of China (Nos. 81671158 and 81771261), and the Natural Science Youth Foundation of China (No. 81701165), and the Natural Science Foundation of Chongqing Science and Technology Committee, China (No. cstc2015jcyjA10048). Declaration of Interests: The authors declare that they have no competing interests.Ethical Approval Statement: All animals used in this experiment were cared for in strict accordance with the Guide for the Care and Use of Laboratory Animals (NIH Publication No. 85-23, revised 1996). All animal experimental procedures were approved by the Ethics Committee of Experimental Animals of Chongqing Medical University.

Objective To investigate the effect of tyrosine kinase Fyn on oxidative stress after intracerebral hemorrhage (ICH) in rats and its potential mechanism. Methods A total of 96 male SD rats were randomly assigned to 4 groups, that is, Sham group, ICH group, negative control group (ICH+Si-NC group) and Fyn knockdown group (ICH+Si-Fyn) group. ICH model was induced via injecting 50 μL autologous blood into the basal ganglia. Small interfering fragments were injected into the lateral ventricle to inhibit the expression of Fyn. Mortality, Nissl staining, neurological function score, blood-brain barrier permeability test were detected at 24 h after ICH, and ELASA was used to detect the contents of SOD, GSH, GSH-PX, MDA, and H2O2 in the brain tissue of ICH rats. The expression of Fyn, nuclear factor erythroid 2-related factor 2 (Nrf2) and Nrf2 downstream proteins, heme oxygenase-1 (HO-1) and quinine oxidoreductase 1 (NQO1) were detected by Western blotting. Results Compared with the Sham group, the ICH group had increased expression of Fyn in brain tissue (P < 0.05), elevated mortality, increased neuronal degeneration and necrosis, decreased modified Garcia score (P < 0.05), elevated score of balance beam test (P < 0.05), decreased blood-brain barrier permeability, inhibited activities of brain tissue SOD, GSH and GSH-PX (P < 0.05), increased contents of MDA and H2O2 (P < 0.05), and enhanced expression levels of Nrf2, HO-1 and NQO1 (P < 0.05). There were no significant differences in above indicators between the ICH group and the ICH+Si-NC group. Compared with the ICH+Si-NC group, Si-Fyn transfection effectively inhibited the expression of Fyn, decreased the mortality of the ICH+Si-Fyn group, attenuated neuronal degeneration and necrosis, improved neurological function score (P < 0.05), and decreased blood-brain barrier permeability. The activities of SOD, GSH and GSH-PX of the brain tissue around hematoma were enhanced (P < 0.05), while the contents of MDA and H2O2 were decreased (P < 0.05) and the expression of Nrf2, HO-1 and NQO1 were up-regulated (P < 0.05). Conclusion Inhibition of Fyn attenuates oxidative stress injury after cerebral hemorrhage in rats, and its mechanism may be related to activation of Nrf2 signaling pathway.

Objective To investigate the role of protein tyrosine phosphatase non-receptor type 22 (PTPN22) in inflammatory response following intracerebral hemorrhage (ICH) in rats and explore the mechanism that mediates its effect. Methods Rat models of ICH were established by injecting autologous blood into the brain of 30 SD rats, and Western blotting was performed at 3, 6, 12, 24, and 48 h after ICH (6 rats at each time point) and in 6 sham-operated rats to determine the time window of PTPN22 expression after ICH. Another 72 rats were randomized equally into sham operation group, ICH group, ICH+negative control (ICH+NC) group, and ICH+PTPN22 interference group, and the neurological function score (mNSS) and brain water content were assessed at 24 h after ICH or the sham operation. Western blotting was used to detect the expression of cleaved interleukin-18 (cleaved IL-18), cleaved-IL-1β and cleaved caspase-1 around the hematoma in the rats. Histomorphological changes of the brain tissues of the rats were observed using HE and Nissl staining, and the expression of myeloperoxidase (MPO) around the hematoma was detected using immunofluorescence staining. Results The expression of PTPN22 began to increase significantly at 12 h after ICH in the rats, peaked at 24 h and began to decrease at 48 h (P < 0.05). Compared with those in ICH+NC group, the rats in ICH+PTPN22 interference group had significantly lowered neurological scores and reduced brain water content (P < 0.05) with obviously alleviated brain tissue damage as shown by HE and Nissl staining. The expression of NLRP3, cleaved IL-1β, cleaved IL-18 and cleaved caspase-1 and the number of MPO-positive cells around the hematoma were all significantly lowered in PTPN22 interference group compared with ICH+NC group (P < 0.05). Conclusion Small interfering RNA-mediated interference of PTPN22 attenuates inflammatory response after ICH in rats by inhibiting the activation of NLRP3.