Ceramide induces macrophage migration inhibitory factor -mediated parthanatos in mouse neurons by increasing ROS levels.

Ceramides, the key component of sphingolipid metabolism and second messengers, have been associated with neurodegenerative diseases progression and pathology, and can induce neuronal apoptosis and necrosis, but the effect of ceramide on parthanatos has not been fully elucidated. In this study, we investigated the ceramide-mediated parthanatos pathway and the role of macrophage inhibitory factor (MIF) in parthanatos. We found that ceramide significantly diminished the viability and induced the death of primary cortical neurons. These effects were not prevented by treatment with the pan-caspase inhibitor Z-VAD-FMK treatment; in contrast, treatment with the poly (ADP ribosyl) polymerase-1 (PARP-1) inhibitor ABT-888 prevented these ceramide-mediated effects. Specifically, ceramide induced PARP-1 overactivation, increased PAR polymer levels, facilitated apoptosis-inducing factor (AIF) and MIF nuclear translocation and induced DNA damage. Knockdown of MIF with an adenovirus carrying a MIF short hairpin RNA (shRNA) inhibited ceramide-induced DNA damage and neuronal death, but nuclear translocation of AIF was unaffected. Furthermore, ceramide increased reactive oxygen species (ROS) levels, and N-acetyl cysteine (NAC) significantly inhibited PAR production and neuronal death. These findings suggested that ceramide induced neuronal parthanatos by increasing ROS levels and that MIF might be downstream of AIF in the ceramide-mediated parthanatos pathway. In conclusion, our results suggest that knocking down MIF expression may be a potential therapeutic strategy for nervous system diseases.

Phloretin enhances autophagy by impairing AKT activation and inducing JNK-Beclin-1 pathway activation.

Phloretin is a type of dihydrochalcone that is primarily found in apples and has been reported to possess various potent biological activities, such as anticancer, antioxidant and anti-inflammatory effects. Our previous study has shown that phloretin induces apoptosis in human glioblastoma. In this study, we found that phloretin induced autophagy in SH-SY5Y cells by decreasing p-AKT and p-mTOR levels in the AKT/mTOR pathway and increasing the activation of JNK, the phosphorylation of c-Jun and the expression of Beclin-1. Moreover, the upregulation of Beclin-1 was decreased by SP600125 or a siRNA against c-Jun. Furthermore, SP600125 and siRNAs against c-Jun and Beclin-1 inhibited phloretin-induced autophagy. In addition, inhibition of phloretin-induced autophagy by cotreatment with phloretin and 3-MA decreased phloretin-induced cytotoxicity to SH-SY5Y cells. In conclusion, our results suggest that the AKT/mTOR pathway and JNK-mediated Beclin-1 expression are involved in phloretin-induced autophagy. Phloretin can be used to protect neurons during phloretin treatment of glioblastoma.

MicroRNA-130a regulates neurological deficit and angiogenesis in rats with ischaemic stroke by targeting XIAP.

MicroRNAs (miRNAs) have already been proposed to be implicated in the development of ischaemic stroke. We aim to investigate the role of miR-130a in the neurological deficit and angiogenesis in rats with ischaemic stroke by regulating X-linked inhibitor of apoptosis protein (XIAP). Middle cerebral artery occlusion (MCAO) models were established by suture-occluded method, and MCAO rats were then treated with miR-130a mimics/inhibitors or/and altered XIAP for detection of changes of rats' neurological function, nerve damage and angiogenesis in MCAO rats. The oxygen-glucose deprivation (OGD) cellular models were established and respectively treated to determine the roles of miR-130a and XIAP in neuronal viability and apoptosis. The expression levels of miR-130a and XIAP in brain tissues of MCAO rats and OGD-treated neurons were detected. The binding site between miR-130a and XIAP was verified by luciferase activity assay. MiR-130a was overexpressed while XIAP was down-regulated in MCAO rats and OGD-treated neurons. In animal models, suppressed miR-130a improved neurological function, alleviated nerve damage and increased new vessels in brain tissues of rats with MCAO. In cellular models, miR-130a inhibition promoted neuronal viability and suppressed apoptosis. Inhibited XIAP reversed the effect of inhibited miR-130a in both MCAO rats and OGD-treated neurons. XIAP was identified as a target of miR-130a. Our study reveals that miR-130a regulates neurological deficit and angiogenesis in rats with MCAO by targeting XIAP.

Long noncoding MIAT acting as a ceRNA to sponge microRNA-204-5p to participate in cerebral microvascular endothelial cell injury after cerebral ischemia through regulating HMGB1.

This study is applied to the investigation of the long noncoding RNA myocardial infarction associated transcript's (MIAT's) role in regulating the expression of high-mobility group box 1 (HMGB1) in cerebral microvascular endothelial cell (CMEC) injury after cerebral ischemia by serving as a competitive endogenous RNA (ceRNA) to sponge microRNA-204-5p (miR-204-5p). The cerebral ischemia model of middle cerebral artery occlusion (MCAO) in rats was established by the suture method, in which rats were injected with empty plasmids and MIAT siRNA plasmids. The cerebral ischemia injury model in vitro was established through oxygen glucose deprivation (OGD) in primary cultured CMECs in rats. The cells were transfected with empty plasmids and MIAT siRNA plasmids. The MIAT/miR-204-5p/HMGB1 axis' function in damage and angiogenesis of CMECs were explored. The binding site between MIAT and miR-204-5p along with that between miR-204-5p and HMGB1 was determined. MIAT was overexpressed in MCAO rats' brain tissue and inhibited MIAT attenuated the injury of brain tissue in MCAO rats. Inhibition of MIAT promoted angiogenesis, promoted miR-204-5p expression and inhibited HMGB1 expression in brain tissue of MCAO rats. Inhibition of MIAT reduced CMEC damage, induced angiogenesis of CMECs, increased the number of surviving neurons, promoted miR-204-5p expression and inhibited HMGB1 expression in CMECs treated with OGD. MIAT promoted HMGB1 expression by competitive binding to miR-204-5p to regulate the injury of CMECs after cerebral ischemia. Our study showed that MIAT promoted HMGB1 expression by competitively binding to miR-204-5p to regulate the injury of CMECs after cerebral ischemia.

Role of XIAP gene overexpressed bone marrow mesenchymal stem cells in the treatment of cerebral injury in rats with cerebral palsy.

BACKGROUND: This study is performed to investigate the effects of adenovirus-mediated X-linked inhibitor of apoptosis protein (XIAP) overexpressed bone marrow mesenchymal stem cells (BMSCs) on brain injury in rats with cerebral palsy (CP). METHODS: Rat's BMSCs were cultured and identified. The XIAP gene of BMSCs was modified by adenovirus expression vector Ad-XIAP-GFP. The rat model of CP with ischemia and anoxia was established by ligating the left common carotid artery and anoxia for 2 h, and BMSCs were intracerebroventricularly injected to the modeled rats. The mRNA and protein expression of XIAP in brain tissue of rats in each group was detected by RT-qPCR and western blot analysis. The neurobehavioral situation, content of acetylcholine (Ach), activity of acetylcholinesterase (AchE), brain pathological injury, apoptosis of brain nerve cells and the activation of astrocytes in CP rats were determined via a series of assays. RESULTS: Rats with CP exhibited obvious abnormalities, increased Ach content, decreased AchE activity, obvious pathological damage, increased brain nerve cell apoptosis, as well as elevated activation of astrocyte. XIAP overexpressed BMSCs improved the neurobehavioral situation, decreased Ach content and increased AchE activity, attenuated brain pathological injury, inhibited apoptosis of brain nerve cells and the activation of astrocytes in CP rats. CONCLUSION: Our study demonstrates that XIAP overexpressed BMSCs can inhibit the apoptosis of brain nerve cells and the activation of astrocytes, increase AchE activity, and inhibit Ach content, so as to lower the CP caused by cerebral ischemia and hypoxia in rats.

In vivo evaluation of urokinase-loaded hollow nanogels for sonothrombolysis on suture embolization-induced acute ischemic stroke rat model.

The urokinase-type plasminogen activator (uPA) loaded hollow nanogels (nUK) were synthesized by a one-step reaction of glycol chitosan and aldehyde capped poly (ethylene oxide). The resultant formulation is sensitive to diagnostic ultrasound (US) of 2 MHz. Herein, we evaluated the in vivo sonothrombolysis performance of the nUK on acute ischemic stroke rat model which was established by suture embolization of middle cerebral artery (MCA). Via intravenous (i.v.) administration, the experimental data prove a controlled release of the therapeutic protein around the clots under ultrasound stimulation, leading to enhanced thrombolysis efficiency of the nUK, evidenced from smaller infarct volume and better clinical scores when compared to the i.v. dose of free uPA no matter with or without US intervention. Meanwhile, the preservation ability of the nanogels not only prolonged the circulation duration of the protein, but also resulted in the better blood-brain barrier protection of the nUK formulation, showing no increased risk on the hemorrhagic transformation than the controls. This work suggests that the nUK is a safe sonothrombolytic formulation for the treatment of acute ischemic stroke.

Apolipoprotein C-III in the high-density lipoprotein proteome of cerebral lacunar infarction patients impairs its anti-inflammatory function.

High-density lipoprotein (HDL) proteomic study has identified substantial changes associated with various disease states. In the current study, the HDL proteomes in patients with cerebral lacunar infarction (LACI) and control subjects were investigated. A total of 12 LACI patients without evident large vessel occlusions and 12 controls were enrolled in the study. The HDL fraction from each sample was isolated from the plasma by ultracentrifugation. The protemics of the HDL were investigated using nano liquid chromatography coupled to tandem mass spectrometry. There were 55 proteins identified as differentially expressed in the LACI and control groups. Among the 55 proteins, 33 were upregulated and 22 were downregulated in the patients with LACI. The identified proteins were associated with numerous molecular functions, including lipid and cholesterol transport, lipid metabolism, inflammatory response, the complement and coagulation pathway, metal ion metabolism, hemostasis and endopeptidase inhibitory activity. Serum amyloid A, apolipoprotein C (apoC-III) and apolipoprotein A-II (apoA-II) were selected to confirm the proteomics results via western blotting. HDL from the LACI patients exhibited an impaired ability to inhibit the binding of THP-1 cells to endothelial cells compared with the controls (P<0.01). ApoC-III-rich HDL also had a significantly reduced ability to inhibit the binding of THP-1 cells to endothelial cells (P<0.01). The expression of vascular cell adhesion molecule-1 protein by the endothelial cells exhibited a similar pattern of response to the different HDL samples. In conclusion, the present study demonstrates major modifications of the HDL proteome in patients with LACI. The ApoC-III enrichment of the HDL of patients with LACI may cause a reduction in the anti-inflammatory ability of HDL, which may contribute to the progression of the disease.

Discrepant relationships between admission blood pressure and mortality in different stroke subtypes.

The relationship between blood pressure(BP) and clinical outcome in patients with acute stroke is still controversial. The present study aimed to elucidate the impact of admission blood pressure on mortality in patients with acute stroke of different subtypes. Data were from ChinaQUEST (QUality Evaluation of Stroke Care and Treatment), a multicenter, prospective hospital registry study in 37 cities across China. A total of 6427 patients were admitted within 24h of onset and after following up for 12months, 5501 were included in the final analysis. Multivariate Cox regression model were used in data analysis. A "U-curve shaped" relationship was observed between admission systolic or diastolic BP and mortality at 12months in the overall study population. Compared to first quartile, the Hazard ratio (HR) for the systolic BP of top quartile was 1.444 (95%CI 1.854-1.636), while the HR was 0.692 (95%CI 0.802-0.930) for the second quartile. Similar associations were observed when we applied admission diastolic BP. In subgroup analysis, the U-shaped effect was remained only in patients with intracranial hemorrhage (ICH). The HR for the systolic BP of top quartile was 2.274 (95%CI 1.878-2.755), while the HR was 0.751 (95%CI 0.571-0.986) for the second quartile. Moreover, admission diastolic BP of top quartile was significantly associated with elevated risk of death for patients with ischemic stroke caused by small vessel diseases (LACI)(HR 1.470; CI 1.040-2.078). In addition, we found a heterogeneity of the admission BP distribution among different subtypes, which may explain the "U-curve" effect.

Prevalence and risk factors of intracranial and extracranial artery stenosis in asymptomatic rural residents of 13 villages in China.

BACKGROUND: The present study aimed to investigate the prevalence and risk factors for extracranial carotid artery stenosis (ECAS) and intracranial carotid artery stenosis (ICAS) simultaneously in asymptomatic Chinese pure rural population. METHODS: We analyzed 2589 asymptomatic subjects aged over 30 yr. by ultrasonography and transcranial Doppler simultaneously in 13 isolated villages by door-to-door investigation. Both ECAS and ICAS were defined as more than 50% stenosis. Demographics, medical history documentation, and investigation of biochemical results were performed for each subject. Univariate and multivariate logistic regression analyses were employed to assess the risk factors associated with ECAS and ICAS, respectively. RESULTS: One hundred twenty-two (4.7%) residents with ICAS and 56 (2.2%) with ECAS were found in 2589 subjects. Three factors emerged as independent risk factors for ICAS: age (95% confidence interval [CI] = 1.01-1.04, odds ratio [OR] = 1.07), hypertension (95% CI = 1.98-4.37, OR = 2.94), and diabetes mellitus (95% CI = 1.72-4.38, OR = 2.75). As for ECAS, five factors presented as independent risk factors: age (95% CI = 1.09-1.11, OR = 1.10), male sex (95% CI = 1.01-1.02, OR = 1.01), diabetes mellitus (95% CI = 1.10-2.12, OR = 1.53), systolic blood pressure (95% CI = 1.95-2.88, OR = 2.37), and total cholesterol (95% CI = 1.00-1.13, OR = 1.06). CONCLUSIONS: ICAS and ECAS were relatively common among asymptomatic rural Chinese subjects. Although they shared similar risk factors, differences still existed between them.

Autophagy inhibits C2-ceramide-mediated cell death by decreasing the reactive oxygen species levels in SH-SY5Y cells.

Ceramide has been recognized as a second messenger that regulates several intracellular processes in neuronal cells. However, its role in neuronal autophagy is not fully understood. In this study, we used a human neuroblastoma cell line (SH-SY5Y) to investigate the mechanisms underlying C2-ceramide-mediated cell death and autophagy. C2-ceramide induced caspase-3-independent cell death. In addition, C2-ceramide induced autophagy, decreased the activation of Akt and mTOR, and increased the activation of JNK and ERK1/2. However, only inhibition of ERK1/2 with PD98059 prevented C2-ceramide-induced autophagy, indicating that the ERK1/2 pathway contributes to ceramide-induced autophagy. According to the results of the flow cytometric assays, C2-ceramide-induced cell death was increased by 3-methyadenine (3-MA) and decreased by rapamycin. Furthermore, the generation of reactive oxygen species (ROS) in the cells was increased by 3-MA and decreased by rapamycin. Based on these datas, autophagy protected SH-SY5Y cells from C2-ceramide-induced cell death by decreasing ROS production. Therapeutic strategies that regulate autophagy may be used in the treatment of neurological disorders associated with ceramide-induced cell death.

Anti-ganglioside antibodies in Guillain-Barré syndrome and chronic inflammatory demyelinating polyneuropathy in Chinese patients.

INTRODUCTION: In this study we investigated the relationships between anti-ganglioside antibodies and Guillain-Barré syndrome (GBS) and chronic inflammatory demyelinating polyneuropathy (CIDP). METHODS: Samples from 48 Chinese patients diagnosed with GBS and 18 patients diagnosed with CIDP were retrospectively reviewed. RESULTS: In the GBS patients, 62.5% were classified as having acute inflammatory demyelinating polyneuropathy (AIDP), 27.1% were found to have acute motor axonal neuropathy (AMAN), and 10.4% were unclassified. Serum IgG anti-ganglioside antibodies were detected in 46.2% of the AMAN patients and in 6.7% of the AIDP patients (P < 0.05); 5.6% of the 18 CIDP patients were IgG antibody positive, and 27.8% were IgM antibody positive. Facial palsy and sensory impairment were significantly associated with IgM antibodies. CONCLUSIONS: These results suggest that IgG anti-GM1 antibodies are associated with AMAN, but not with AIDP, and that IgM antibodies against GM1, GM2, and GM3 are associated with facial nerve palsy. Muscle Nerve 55: 470-475, 2017.

Comparison of Risk Factor between Lacunar Stroke and Large Artery Atherosclerosis Stroke: A Cross-Sectional Study in China.

BACKGROUND: Stroke is the second most common cause of mortality in China. Although most subtypes of ischemic stroke share similar risk factors, they have different etiologies. Our study aimed to evaluate the different risk factor profiles between the stroke subtypes, lacunar infarcts (LI) and large-artery atherosclerosis (LAA), and clarify the characteristics of current acute ischemic stroke in China. METHODS: In this cross-sectional study, we analyzed the clinical characteristics of 1982 patients with acute ischemic stroke who were admitted to the neurology department at the Peking University First Hospital between 2007 and 2014. Ischemic stroke was further classified into LAA, LI, cardioembolism (CE) and undetermined causes of infarction (UDI) according to TOAST classification. Demographic characteristics, risk factors, as well as the findings of laboratory and imaging tests of 1773 patients with LAA and LI, were analyzed by univariate and multivariate logistic analysis. RESULTS: Of the 1982 ischemic stroke patients included in this study, 1207 were diagnosed with LAA, 566 with LI, 173 with cardioembolism (CE) and 36 with undetermined causes of infarction (UDI). By comparing the risk factors in multivariate logistic regression analysis, hypertension [odds ratio (OR) = 1.832] and white matter leukoaraiosis (WML) (OR = 1.865) were found to be more strongly correlated with LI than LAA. Low density lipoprotein- cholesterol (LDL-c) (OR = 0.774) were more strongly related to LAA than LI. CONCLUSIONS: This study found that hypertension and WML were more strongly correlated with LI than LAA. LDL-c was more strongly related to LAA than LI.

Phloretin induces cell cycle arrest and apoptosis of human glioblastoma cells through the generation of reactive oxygen species.

Phloretin, a flavonoid present in various plants, has been reported to exert anticarcinogenic effects. However, the mechanism of its chemo-preventive effect on human glioblastoma cells is not fully understood. This study aimed to investigate the molecular mechanism of phloretin and its associated chemo-preventive effect in human glioblastoma cells. The results indicate that phloretin inhibited cell proliferation by inducing cell cycle arrest at the G0-G1 phase and induced apoptosis of human glioblastoma cells. Phloretin-induced cell cycle arrest was associated with increased expression of p27 and decreased expression of cdk2, cdk4, cdk6, cyclinD and cyclinE. Moreover, the PI3K/AKT/mTOR signaling cascades were suppressed by phloretin in a dose-dependent manner. In addition, phloretin triggered the mitochondrial apoptosis pathway and generated reactive oxygen species (ROS). This was accompanied by the up-regulation of Bax, Bak and c-PARP and the down-regulation of Bcl-2. The antioxidant agents N-acetyl-L-cysteine and glutathione weakened the effect of phloretin on glioblastoma cells. In conclusion, these results demonstrate that phloretin exerts potent chemo-preventive activity in human glioblastoma cells through the generation of ROS.

Association between Low Estimated Glomerular Filtration Rate and Risk of Cerebral Small-Vessel Diseases: A Meta-Analysis.

BACKGROUND: Although chronic kidney disease has been linked to cerebral small-vessel disease (CSVD), a definite relationship between them has not been established. This study assessed whether low estimated glomerular filtration is associated with risk of different subtypes of CSVDs. METHODS: Electronic databases were systematically searched for studies reporting an odds ratio of the association between low estimated glomerular filtration and CSVD risk. Sixteen studies, including 10,534 participants, were identified. A fix effects model was applied and odds ratios (ORs) with 95% confidence intervals were presented. RESULTS: Overall, risk of CSVDs was greater in individuals with low estimated glomerular filtration (OR = 2.20). Stratified analyses consistently showed significant associations across different subtypes, with pooled OR being greatest in subjects with silent cerebral infarction (SCI) (OR = 2.71) and cerebral microbleed (OR = 2.70). A pooled estimate of studies showing OR as a continuous variable showed results consistent with the former analysis (OR = .98 per standard deviation decrease) in low estimated glomerular filtration. CONCLUSIONS: This study revealed that low estimated glomerular filtration was significantly associated with risk of CSVDs. Low estimated glomerular filtration was most strongly associated with SCI (OR = 2.71) among subtypes of CSVDs.