Nur77 increases mitophagy and decreases aggregation of α-synuclein by modulating the p-c-Abl/p-PHB2 Y121 in α-synuclein PFF SH-SY5Y cells and mice.

Parkinson's disease (PD) is characterized by the progressive death of dopamine (DA) neurons and the pathological accumulation of α-synuclein (α-syn) fibrils. In our previous study, simulated PHB2 phosphorylation was utilized to clarify the regulatory role of c-Abl in PHB2-mediated mitophagy in PD models. In this investigation, we employed an independently patented PHB2Y121 phosphorylated antibody in the PD model to further verify that the c-Abl inhibitor STI571 can impede PHB2Y121 phosphorylation, decrease the formation of α-Syn polymers, and improve autophagic levels. The specific involvement of Nur77 in PD pathology has remained elusive. We also investigate the contribution of Nur77, a nuclear transcription factor, to α-syn and mitophagy in PD. Our findings demonstrate that under α-syn, Nur77 translocates from the cytoplasm to the mitochondria, improving PHB-mediated mitophagy by regulating c-Abl phosphorylation. Moreover, Nur77 overexpression alleviates the expression level of pS129-α-syn and the loss of DA neurons in α-syn PFF mice, potentially associated with the p-c-Abl/p-PHB2 Y121 axis. This study provides initial in vivo and in vitro evidence that Nur77 protects PD DA neurons by modulating the p-c-Abl/p-PHB2 Y121 axis, and STI571 holds promise as a treatment for PD.

A novel mechanism of PHB2-mediated mitophagy participating in the development of Parkinson's disease.

JOURNAL/nrgr/04.03/01300535-202408000-00037/figure1/v/2023-12-16T180322Z/r/image-tiff Endoplasmic reticulum stress and mitochondrial dysfunction play important roles in Parkinson's disease, but the regulatory mechanism remains elusive. Prohibitin-2 (PHB2) is a newly discovered autophagy receptor in the mitochondrial inner membrane, and its role in Parkinson's disease remains unclear. Protein kinase R (PKR)-like endoplasmic reticulum kinase (PERK) is a factor that regulates cell fate during endoplasmic reticulum stress. Parkin is regulated by PERK and is a target of the unfolded protein response. It is unclear whether PERK regulates PHB2-mediated mitophagy through Parkin. In this study, we established a 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-induced mouse model of Parkinson's disease. We used adeno-associated virus to knockdown PHB2 expression. Our results showed that loss of dopaminergic neurons and motor deficits were aggravated in the MPTP-induced mouse model of Parkinson's disease. Overexpression of PHB2 inhibited these abnormalities. We also established a 1-methyl-4-phenylpyridine (MPP+)-induced SH-SY5Y cell model of Parkinson's disease. We found that overexpression of Parkin increased co-localization of PHB2 and microtubule-associated protein 1 light chain 3, and promoted mitophagy. In addition, MPP+ regulated Parkin involvement in PHB2-mediated mitophagy through phosphorylation of PERK. These findings suggest that PHB2 participates in the development of Parkinson's disease by interacting with endoplasmic reticulum stress and Parkin.

The key roles of reactive oxygen species in microglial inflammatory activation: Regulation by endogenous antioxidant system and exogenous sulfur-containing compounds.

Aberrant innate immunity in the brain has been implicated in the pathogenesis of several central nervous system (CNS) disorders, including Alzheimer's disease, Huntington's disease, Parkinson's disease, stroke, amyotrophic lateral sclerosis, and depression. Except for extraparenchymal CNS-associated macrophages, which predominantly afford protection against peripheral invading pathogens, it has been reported that microglia, a population of macrophage-like cells governing CNS immune defense in nearly all neurological diseases, are the main CNS resident immune cells. Although microglia have been recognized as the most important source of reactive oxygen species (ROS) in the CNS, ROS also may underlie microglial functions, especially M1 polarization, by modulating redox-sensitive signaling pathways. Recently, endogenous antioxidant systems, including glutathione, hydrogen sulfide, superoxide dismutase, and methionine sulfoxide reductase A, were found to be involved in regulating microglia-mediated neuroinflammation. A series of natural sulfur-containing compounds, including S-adenosyl methionine, S-methyl-L-cysteine, sulforaphane, DMS, and S-alk(enyl)-l-cysteine sulfoxide, modulating endogenous antioxidant systems have been discovered. We have summarized the current knowledge on the involvement of endogenous antioxidant systems in regulating microglial inflammatory activation and the effects of sulfur-containing compounds on endogenous antioxidant systems. Finally, we discuss the possibilities associated with compounds targeting the endogenous antioxidant system to treat neuroinflammation-associated diseases.

Rutin Attenuates Oxidative Stress Via PHB2-Mediated Mitophagy in MPP+-Induced SH-SY5Y Cells.

Oxidative stress plays a crucial role in the occurrence and development of Parkinson's disease (PD). Rutin, a natural botanical ingredient, has been shown to have antioxidant properties. Therefore, the aim of this study was to investigate the neuroprotective effects of rutin on PD and the underlying mechanisms. MPP+(1-methyl-4-phenylpyridinium ions)-treated SH-SY5Y cells were used as an in vitro model of PD. Human PHB2-shRNA lentiviral particles were transfected into SH-SY5Y cells to interfere with the expression of Prohibitin2 (PHB2). The oxidative damage of cells was analyzed by detecting intracellular reactive oxygen species (ROS), malondialdehyde (MDA), and mitochondrial membrane potential (MMP). Western blotting was used to detect the protein expression of antioxidant factors such as nuclear factor E2-related factor 2 (Nrf2), heme oxygenase-1 (HO-1), NADPH quinone oxidoreductase-1 (NQO-1), and mitophagy factors PHB2, translocase of outer mitochondrial membrane 20 (TOM20), and LC3II/LC3I (microtubule-associated protein II light chain 3 (LC3II) to microtubule-associated protein I light chain 3 (LC3I)). In addition, we also examined the expression of PHB2 and LC3II/LC3I by immunofluorescence staining. MPP+ treatment significantly increased the generation of ROS and MDA and the level of MMP depolarization and decreased the protein expression of Nrf2, HO-1, NQO1, TOM20, PHB2, and LC3II/LC3I. In MPP+-treated SH-SY5Y cells, rutin significantly decreased the generation of ROS and MDA and the level of MMP depolarization and increased the protein expression of Nrf2, HO-1, NQO-1, TOM20, PHB2, and LC3II/LC3I. However, the protective role of rutin was inhibited in PHB2-silenced cells. Rutin attenuates oxidative damage which may be associated with PHB2-mediated mitophagy in MPP+-induced SH-SY5Y cells. Rutin might be used as a potential drug for the prevention and treatment of PD.

Artemisinin Alleviates Cerebral Ischemia/Reperfusion-Induced Oxidative Damage via Regulating PHB2-Mediated Autophagy in the Human Neuroblastoma SH-SY5Y Cell Line.

Oxidative stress plays a key role in cerebral ischemia/reperfusion injury. Artemisinin (ART) has antioxidative stress activity in addition to its powerful antimalarial effects. In this article, we investigated the effect of ART on OGD/R-induced oxidative stress injury and its underlying mechanisms. We used oxygen-glucose deprivation/reoxygenation (OGD/R) to establish an in vitro model of cerebral ischemia/reperfusion (I/R) injury. CCK-8 and lactate dehydrogenase (LDH) release were used to assess cellular damage. Measurement of reactive oxygen species (ROS), malondialdehyde (MDA), superoxide dismutase (SOD), glutathione (GSH), and mitochondrial membrane potential (MMP) estimates oxidative stress-induced damage and protection from ART effect. OGD/R treatment aggravated oxidative stress damage, whereas ART reversed the effects of OGD/R. Autophagy is closely related to oxidative stress; in order to confirm whether the antioxidative stress effect of ART is related to PHB2-mediated autophagy, we examined the protein expression of prohibitin 2 (PHB2), TOMM20, p62, and the conversion of microtubule-associated protein light chain 3I (LC3I) to LC3II and found that the protein expression of PHB2, TOMM20, p62, and LC3II/LC3I was significantly correlated with OGD/R treatment. The colocalization of PHB2 and LC3, TOMM20, and LC3 was reduced after OGD/R treatment, and ART reversed this change. After silencing PHB2, the protective effect of ART against OGD/R-induced oxidative stress injury was reduced, the protein expressions of PHB2, TOMM20 and LC3II/LC3I and the colocalization of PHB2 and LC3, TOMM20, and LC3 were decreased. We used chloroquine to block the lysosomal pathway and found that ART increased the conversion of LC3I to LC3II, silencing PHB2 which inhibited the conversion of LC3I to LC3II, and impaired mitophagy. Our findings showed that ART attenuated OGD/R-induced oxidative stress damage through PHB2-mediated mitophagy. To the current knowledge, our study is the first to demonstrate that ART attenuates OGD/R-induced oxidative stress injury through PHB2-mediated autophagy in the human neuroblastoma SH-SY5Y cell line, which provided new insights into the treatment of OGD/R injury.

Nonreceptor Tyrosine Kinase c-Abl-Mediated PHB2 Phosphorylation Aggravates Mitophagy Disorder in Parkinson's Disease Model.

Mitophagy and oxidative stress play important roles in Parkinson's disease (PD). Dysregulated mitophagy exacerbates mitochondrial oxidative damage; however, the regulatory mechanism of mitophagy is unclear. Here, we provide a potential mechanistic link between c-Abl, a nonreceptor tyrosine kinase, and mitophagy in PD progression. We found that c-Abl activation reduces the interaction of prohibitin 2 (PHB2) and microtubule-associated protein 1 light chain 3 (LC3) and decreases the expressive level of antioxidative stress proteins, including nuclear factor erythroid 2-related factor 2 (Nrf2), NADPH quinone oxidoreductase-1 (NQO-1), and the antioxidant enzyme heme oxygenase-1 (HO-1) in 1-methyl-4-phenylpyridinium- (MPP+-) lesioned SH-SY5Y cells. Importantly, we found that MPP+ can increase the expression of phosphorylated proteins at the tyrosine site of PHB2 and the interaction of c-Abl with PHB2. We showed for the first time that PHB2 by changing tyrosine (Y) to aspartate (D) at site 121 resulted in impaired binding of PHB2 and LC3 in vitro. Moreover, silencing of PHB2 can decrease the interaction of PHB2 and LC3 and exacerbate the loss of dopaminergic neurons. We also found that STI 571, a c-Abl family kinase inhibitor, can decrease dopaminergic neuron damage and ameliorate MPTP-induced behavioral deficits in PD mice. Taken together, our findings highlight a novel molecular mechanism for aberrant PHB2 phosphorylation as an inhibitor of c-Abl activity and suggest that c-Abl and PHB2 are potential therapeutic targets for the treatment of individuals with PD. However, these results need to be further validated in PHB2 Y121D mice.

Autophagy-Dependent Ferroptosis-Related Signature is Closely Associated with the Prognosis and Tumor Immune Escape of Patients with Glioma.

BACKGROUND: Ferroptosis is an autophagy-dependent form of cell death, sometimes called "ferritinophagy". Its related pathway has been proven to regulate the programmed death of glioma stem cells. Mining autophagy-dependent ferroptosis-related gene (AD-FRG) signature could facilitate the discovery of mechanisms and therapeutic targets showing drug resistance to chemotherapeutic drugs. METHODS: We exhaustively searched HADB, MSigDB and FerrDb datasets and obtained 25 genes confirmed to exist in autophagy and ferroptosis death pathways. Glioma gene expression and clinicopathological data were collected from TCGA and CGGA datasets. RESULTS: Lasso regression and Cox regression analysis were carried out to construct a nine AD-FRGs signature (SIRT1, MTDH, HSPB1, CISD2, HMOX1, ATG7, MTOR, PRKAA2 and EIF2AK4). ROC curve showed that nine genes signature could effectively predict 1- (AUC = 0.869), 3- (AUC = 0.922) and 5-year (AUC = 0.870) survival rates. Immunohistochemical images confirmed the protein expression level of the gene model. The prognostic nomogram of risk score, age, WHO grade, isocitrate dehydrogenase (IDH) wild-type condition, 1p/19q co-deletion state was built. The calibration curve demonstrated that the prediction of the nomogram is highly consistent with the actual results. Moreover, tumor microenvironment analysis showed that the high-risk group was associated with high immune infiltration status and high tumor purity. Correlation analysis showed that the expression of SIRT1, CISD2 and HSPB1 might be related to macrophage infiltration and immunotolerance in glioma tissues. CONCLUSION: Based on autophagy-dependent ferroptosis-related genes, we established gene signature and nomogram that maybe effectively predict the overall survival rate of glioma and correlate with the immunosuppressive tumor microenvironment (TME).

Atorvastatin improves motor function, anxiety and depression by NOX2-mediated autophagy and oxidative stress in MPTP-lesioned mice.

Parkinson's disease (PD) is a neurodegenerative disease caused by the loss of dopaminergic neurons. It is characterized by static tremors, stiffness, slow movements, and gait disturbances, but it is also accompanied by anxiety and depression. Our previous study showed that atorvastatin could reduce the risk of PD, but the mechanism is still unclear. In this paper, Our findings showed that atorvastatin increased muscle capacity and the coordination of movement and improved anxiety and depression. Atorvastatin could decrease the expression of α-synuclein Ser129 and NADPH oxidase 2 (NOX2), increase the protein expression of LC3II/I, and promote autophagy flow. To further confirm that atorvastatin protection was achieved by inhibiting NOX2, we injected at midbrain with NOX2 shRNA (M) lentivirus and found that silent NOX2 produced the same effect as atorvastatin. Further research found that atorvastatin could reduce MPTP-induced oxidative stress damage, while inhibiting NOX2 decreased the antioxidative stress effect of atorvastatin. Our results suggest that atorvastatin can improve muscle capacity, anxiety and depression by inhibiting NOX2, which may be related to NOX2-mediated oxidative stress and autophagy. Atorvastatin may be identified as a drug that can effectively improve behavioral disorders. NOX2 may be a potential gene target for new drug development in PD.

Nur77 attenuates inflammatory responses and oxidative stress by inhibiting phosphorylated IκB-α in Parkinson's disease cell model.

Neuroinflammation and oxidative stress play key roles in the pathological development of Parkinson's disease (PD). Nerve growth factor-induced gene B (Nur77) is closely related to dopamine neurotransmission, and its pathogenesis is unclear. This study aims to investigate the role and mechanism of Nur77 in a cell model of Parkinson's disease. Silencing Nur77 with siRNA can aggravate intracellular LDH release, increase the expression of pro-inflammatory genes (such as tumor necrosis factor α, nuclear factor κB (p65), monocyte chemotactic protein 1, interleukin-6), and decrease cell survival, decrease expression of nuclear factor E2-related factor(Nrf2), heme oxygenase 1, NADPH quinineoxidoreductase-1. Cytosporone B (Nur77 agonist) has the opposite effect to Nur77 silencing. PDTC (NF-κB inhibitor / antioxidant) can also inhibit pro-inflammatory genes to a similar degree as Cytosporone B. Phosphorylated IκB-α can be inhibited by Cytosporone B, while silencing Nur77 can increase the protein expression level of phosphorylated IκB-α. After silencing IκB-α, both Cytosporone B and siNur77 did not affect pro-inflammatory genes and antioxidant stress. These findings reveal the first evidence that Nur77 exerts anti-inflammatory and antioxidant stress effects by inhibiting IκB-α phosphorylation expression in a Parkinson cell model. Nur77 may be a potential therapeutic target for Parkinson's disease.

MsrA Suppresses Inflammatory Activation of Microglia and Oxidative Stress to Prevent Demyelination via Inhibition of the NOX2-MAPKs/NF-κB Signaling Pathway.

INTRODUCTION: Demyelination causes neurological deficits involving visual, motor, sensory symptoms. Deregulation of several enzymes has been identified in demyelination, which holds potential for the development of treatment strategies for demyelination. However, the specific effect of methionine sulfoxide reductase A (MsrA) on demyelination remains unclear. Hence, this study aims to explore the effect of MsrA on oxidative stress and inflammatory response of microglia in demyelination. METHODS: Initially, we established a mouse model with demyelination induced by cuprizone and a cell model provoked by lipopolysaccharide (LPS). The expression of MsrA in wild-type (WT) and MsrA-knockout (MsrA-/-) mice were determined by RT-qPCR and Western blot analysis. In order to further explore the function of MsrA on inflammatory response, and oxidative stress in demyelination, we detected the expression of microglia marker Iba1, inflammatory factors TNF-α and IL-1ß and intracellular reactive oxygen species (ROS), superoxide dismutase (SOD) activity, as well as expression of the NOX2-MAPKs/NF-κB signaling pathway-related genes in MsrA-/- mice and LPS-induced microglia following different treatments. RESULTS: MsrA expression was downregulated in MsrA-/- mice. MsrA silencing was shown to produce severely injured motor coordination, increased expressions of Iba1, TNF-α, IL-1ß, ROS and NOX2, and extent of ERK, p38, IκBα, and p65 phosphorylation, but reduced SOD activity. Conjointly, our study suggests that Tat-MsrA fusion protein can prevent the cellular inflammatory response and subsequent demyelination through negative regulation of the NOX2-MAPKs/NF-κB signaling pathway. CONCLUSION: Our data provide a profound insight on the role of endogenous antioxidative defense systems such as MsrA in controlling microglial function.

Simvastatin Improves Behavioral Disorders and Hippocampal Inflammatory Reaction by NMDA-Mediated Anti-inflammatory Function in MPTP-Treated Mice.

The cognitive function impairment may be related to the inflammation of the hippocampus in Parkinson's disease. Simvastatin can play a positive role in Parkinson's disease. The purpose of this study was to investigate whether simvastatin could improve behavioral disorders, especially depression, anxiety and cognitive function in mouse PD models, and further explore the molecular mechanism. In the present study, C57BL-6 mice underwent intraperitoneal injection of MPTP (30 mg/kg) once a day for 5 consecutive days. At the same time, simvastatin (10 mg/kg) was pretreated for 2 days before the Parkinson's disease model was established, and then continued for 5 days, and the control group underwent intraperitoneal injection of MK801 (dizocilpine, 0.2 mg/kg) and saline solution. Depression status was tested by a tail suspension test and a sucrose splash test, followed by an open-field test and an elevated plus maze test to determine anxiety levels. Spatial behavior and muscle status were measured with a water maze and a rotarod test. The expression of RNA and protein of N-methyl-D-aspartate receptor subtype 2B (NMDAR2B), nerve growth factor IB (Nur77), cyclooxygenase-2 (COX-2), and tumor necrosis factor (TNF) α were assayed by real-time polymerase chain reaction and Western blot. Our results showed that simvastatin can improve the cognitive function, anxiety, and depression of PD mice with MPTP injury. Simvastatin reversed the NMDAR2B increase, restored Nur77 downward, and reduced the expression of COX-2 and TNF-α in MPTP-treated mice. This role of simvastatin was consistent with MK801 in increasing the expression of Nur77 and inhibiting NMDAR2B and cytokines in MPTP-lesioned PD mice. These findings suggest that reversed the NMDAR2B increase, restored Nur77 downward, and reduced the expression of COX-2 and TNF-α in MPTP-treated mice may be one of the mechanisms that simvastatin improves cognitive functions, depression, and anxiety in MPTP-lesioned mice.

Interleukin-17 receptor C gene polymorphism reduces treatment effect and promotes poor prognosis of ischemic stroke.

OBJECTIVE: To study the relationship between Interleukin-17 receptor C (IL-17RC) gene polymorphism and ischemic stroke (IS). METHODS: Three hundred cases of IS patients and 300 cases of the healthy controls were selected. Serum of IS patients and the controls was collected. The relative mRNA levels of IL-17, IL-17RC, IL-6, IL-8, G-CSF and granulocyte-macrophage colony stimulating factor (GM-CSF) by qRT-PCR. The protein expression of IL-17 and IL-17RC was determined by Western blotting. IL-17RC genotype was identified by PCR amplification. The proportion of IL-17RC, SNP and re37511 in IS and control group was determined. The treatment effect on IS and prognosis of patients with IL-17RC, SNP and re37511 was compared. RESULTS: The relative mRNA levels of IL-17, IL-17RC, IL-6, IL-8, G-CSF and GM-CSF in IS group were significantly higher than the control group. The protein expression of IL-17 and IL-17RC in IS group was also markedly higher than the control group. The proportion of IL-17RC re37511 in IS group was much larger than control group and proportion of IL-17RC much less. The percent of poor treatment effect in re37511 was much larger than IL-17RC. The percent of death and recrudescence in patients with IL-17RC re37511 was the highest. CONCLUSION: IS up-regulates the expression of IL-17 and IL-17RC. IL-17RC re37511 indicates the patients have a poorer treatment effect and prognosis.

Protective effects of Progranulin against focal cerebral ischemia-reperfusion injury in rats by suppressing endoplasmic reticulum stress and NF-κB activation in reactive astrocytes.

The aim of this study is to explore the effect progranulin (PGRN) has on endoplasmic reticulum (ER) stress and the NF-κB activation in reactive astrocytes found in rat models with focal cerebral ischemia-reperfusion (I/R) injury. Sprague-Dawley (SD) rats were grouped into the sham, I/R, PGRN-high dose, PGRN-low dose, and negative control (NC) groups. TTC staining was applied in order to detect the cerebral infarction volume, a TUNEL assay to detect the apoptosis rate of neurons, an ELISA to measure MDA, SOD, iNOS, LDH, TNF-α, IL-1ß, IL-6, and IL-10 levels, and both RT-qPCR and western blotting methods in order to detect PGRN, GFAP, GRP78, CHOP, and NF-κB p65 expressions. The astrocytes (AST) cells were then assigned into the normal, I/R, negative control (NC), PGRN-high dose, and PGRN-low dose groups. After completing the transfection process, the proliferative capacity of AST cells was detected by use of the CCK-8 assay. Both the in vivo and in vitro results, in comparison with the I/R and the NC groups, the PGRN-high dose and PGRN-low dose groups both presented a decrease in cerebral infarction volume, apoptosis rate of neurons, MDA, LDH, iNOS, TNF-α, IL-1ß, IL-6 levels, and GFAP, GRP78, CHOP, NF-κB p65 expressions, and an increase in SOD, IL-10, and PGRN levels as well as cell proliferation depending on dosage. Based on our results, we came to the confirmation that PGRN can reduce neuronal apoptosis by mitigating ER stress in the reactive astrocytes as well as downregulating the inflammatory levels by suppressing the NF-κB signaling pathway.

Comparative transcriptome analysis of the global circular RNAs expression profiles between SHEE and SHEEC cell lines.

Esophageal squamous cell carcinoma (ESCC) is widely regarded as one of the most lethal types of cancer around the world. The fact that early detection of ESCC could dramatically improve the treatment outcome of the patients has sparked considerable interest in searching for reliable and accurate diagnostic biomarkers. Recently, circular RNAs (circRNA) have emerged as a new type of non-coding RNAs with significant RNase resistance, wide abundance and remarkable internal diversity. There is also increasing evidence suggesting that circRNAs could be implicated in the pathogenesis of cancer and other diseases. In this study, we performed a comparative analysis of the global circRNA expression profiles in normal and malignant esophageal epithelial cell lines by a combination of RNA sequencing and bioinformatics analysis. We identified 813 significantly up-regulated and 445 down-regulated circRNA candidates, of which 32 were subsequently validated by quantitative real-time reverse transcription polymerase chain reaction analysis. The differentially expressed circRNAs were found to be associated with pathways involved in metabolism, cell apoptosis, proliferation and migration, which are commonly altered in cancer cells. Based on the obtained data, we constructed a circRNA-miRNA interaction network, in which circRNA9927-NBEAL1 represented the biggest node. Our study could lay the groundwork for further investigation concerning the pathological roles of circRNAs in ESCC.

Effect of ticagrelor versus prasugrel on platelet reactivity: a meta-analysis.

BACKGROUND AND OBJECTIVE: Whether ticagrelor is superior to prasugrel in inhibiting platelet reactivity (PR) has remained unclear, possibly because different test methods have been used to determine this. Therefore, using a different test method, we performed a meta-analysis comparing the effects of ticagrelor and prasugrel on PR. METHODS AND RESULTS: PubMed, Embase, Web of Science, and Google Scholar were searched - without language restrictions (last updated on 26 February 2017) - for randomized trials comparing the effects of prasugrel with those of ticagrelor in patients with coronary artery disease. Selected studies were chosen for pooled analysis according to the inclusion and exclusion criteria. Data are presented as mean difference (MD) and 95% confidence interval. For the loading dose, using the VerifyNow-P2Y12 (VN) test, the PR was similar for both the prasugrel and ticagrelor groups [MD=10.80 (-9.81-31.40), P=0.30]. Using the vasodilatorstimulated phosphoprotein test, the PR was also similar for both the ticagrelor and prasugrel groups [MD=-2.87 (-6.35-0.60), P=0.10]. For the maintenance dose, using the VN test, the PR was lower in the ticagrelor group than in the prasugrel group [MD=-43.37 (-60.53 to -26.21), P<0.01]. Finally, using the vasodilator-stimulated phosphoprotein test, the PR was lower in the ticagrelor group than in the prasugrel group [MD=-9.23 (-15.82 to -2.64), P<0.01]. CONCLUSION: There was no difference between ticagrelor and prasugrel in terms of PR under the loading dose, but ticagrelor had a lower degree of PR under the maintenance dose. The results were not affected by the different PR test methods.

Targeting of miR-150 on Gli1 gene to inhibit proliferation and cell cycle of esophageal carcinoma EC9706.

OBJECTIVE: Glioma-associated oncogene homolog 1 (Gli1) in Hedgehog signal pathway regulates Cyclin D1 expression, cell cycle or proliferation modulation. Esophageal cancer patients had significantly elevated Gli1 expression, which is related with survival and prognosis. It has been demonstrated that the level of miR-150 was decreased in esophageal cancer patients compared to normal control. As a complementary relationship exists between miR-150 and 3'-UTR of Gli1, this study investigated if miR-150 played a role in regulating Gli1 expression, and proliferation or cell cycle of esophageal cancer cells. PATIENTS AND METHODS: Esophageal squamous cell carcinoma (ESCC) patients from our hospital were recruited to collect tumor and adjacent tissues for miR-150 and Gli1 expression. Esophageal carcinoma cell line EC9706 and normal esophageal epithelial cell line HEEC were compared for expression of miR-150, Gli1 and Cyclin D1. Dual luciferase reporter gene assay examined the targeted relationship between miR-150 and 3'-UTR of Gli1. In vitro cultured EC9706 cells were treated with miR-150 mimic, si-Gli1 or the combination of miR-150 mimic and si-Gli1, respectively, to check their gene expression, cell cycle and proliferation. RESULTS: ESCC tissues had significantly higher Gli1 expression and lower miR-150 expression. EC9706 cell also had higher Gli1 expression than that in HEEC, whilst miR-150 was down-regulated. Via targeting 3'-UTR of Gli1 gene, miR-150 inhibited its expression. Transfection of miR-150 mimic, si-Gli1 or the combination of miR-150 mimic and si-Gli1, respectively, remarkably decreased expression of Gli1 and Cyclin D1 expression in EC9706 cells, whose cell cycle arresting at G0/G1 phase was enhanced with weakened proliferation. CONCLUSIONS: MiR-150 can induce G0/G1 cell cycle arresting and weaken proliferation of esophageal carcinoma cells via targeted inhibition on Gli1 and downstream expression of Cyclin D1.

Geniposide promotes autophagy to inhibit insulin resistance in HepG2 cells via P62/NF­κB/GLUT­4.

Insulin resistance (IR) is known to be an important factor, which can lead to the onset of type 2 diabetes. Autophagy is a cellular process, which sequesters senescent or damaged proteins in autophagosomes for recycling of their products. Insulin and intracellular molecules, including mammalian target of rapamycin (mTOR), are well­known inhibitors of autophagy. In patients with type 2 diabetes, the expression levels of glucose transporter 4 (GLUT­4) in skeletal muscles are significantly decreased, indicating decreased glucose­processing ability. Geniposide is an iridoid compound isolated from Gardenia jasminoides Ellis. Previously, it was reported that geniposide significantly promoted glucose uptake. In the present study, a HepG2 cell model of IR was constructed to determine whether geniposide can promote autophagy to inhibit insulin resistance in HepG2 cells via P62/nuclear factor (NF)­κB/GLUT­4. Cell proliferation was analyzed by performing an MTT assay, and the mRNA expression levels of NF­κB and GLUT­4 were assessed using semi­quantitative polymerase chain reaction and immunohistochemical staining. In addition, the protein levels of GLUT­4, P62 and phosphorylated­P65 were assessed by western blotting. The expression of GLUT­4 was initially increased following geniposide treatment, decreasing in time to its lowest level at 8 h. The expression levels of NF­κB and GLUT­4 in the IR cells treated with and without geniposide were significantly different, compared with those in the control group. Geniposide promoted autophagy in the IR HepG2 cells and significantly improved IR in the HepG2 cells, which may be associated with the dynamic regulation of the P62/NF­κB/GLUT­4 pathway.

Sinomenine Protects PC12 Neuronal Cells against H2O2-induced Cytotoxicity and Oxidative Stress via a ROS-dependent Up-regulation of Endogenous Antioxidant System.

Sinomenine (SN), a purified alkaloid from Chinese herb Sinomenium acutum that was used preferentially in the treatment of rheumatoid diseases, has exerted neuroprotective effects and anti-inflammatory properties in many previous studies. Some studies have revealed that the antioxidant property of SN, acting mainly through inhibiting NADPH oxidase activation, was involved in the beneficial effects of SN. However, SN belongs to the family of dextrorotatory morphinan analogues, which may initiate elevation of reactive oxygen species (ROS) levels. Thus in the present report, we conducted studies to examine its impact and mechanism on the resistance of PC12 neuronal cells to oxidative stress. Precondition with SN (0.1-5 µM) for 12 h significantly decreased H2O2-induced cytotoxicity and remarkably alleviated oxidative injury. However, SN exhibited little direct free radical scavenging property in vitro and induced "appropriate" production of ROS in PC12 cell. Interestingly, the SN-triggering ROS production served as a signal to activate the Nrf2 antioxidant system including Nrf2, HO-1, and NQO-1, which was inhibited by the antioxidant trolox. Furthermore, Nrf2 knockdown largely attenuated the beneficial effects of SN precondition on oxidative stress. In conclusion, our findings suggested that SN increased the resistance to oxidative stress in neuronal cells via a ROS-dependent up-regulation of endogenous antioxidant system, and this mechanism may be involved in the neuroprotection of SN.

Mir-483-5p promotes the malignant transformation of immortalized human esophageal epithelial cells by targeting HNF4A.

MicroRNAs (miRNAs) are small non-coding RNAs that promote the progression of cancer by negatively regulating gene expression. Down-regulation of miR-483-5p was reported in a number of cancers. However, the biological functions of miR-483-5p in esophageal squamous cell carcinomas are not fully understood. In this study, the expression levels of miRNAs in the immortalized human esophageal epithelial cell line SHEE and the malignantly transformed esophageal carcinoma cell line SHEEC were examined by miRNA microarray chip. The expression level of miR-483-5p was verified by a quantitative reverse transcription-polymerase chain reaction. Growth, apoptosis, and colony formation ability were also examined in SHEEC cells after transfection with inhibitors targeting miR-483-5p. And the target genes of miR-483-5p were predicted using bioinformatics approaches and the expression profile of SHEEC cells transfected with the miRNA inhibitors. protein levels of the target gene in SHEEC cells with a control or miRNA inhibitors were measured using Western blotting. The expression of miR-483-5p was elevated in SHEEC cells as compared to the SHEE cells. Silencing of miR-483-5p expression in SHEEC cells inhibited both the proliferation and formation of colonies and increased apoptosis. We also identified hepatocyte nuclear factor 4α (HNF4A) as a target of miR-483-5p in SHEEC cells. Knockdown of HNF4A recapitulated the effects of miR-483-5p. Our data showed that the miR-483-5p/HNF4A axis affected the malignant transformation of immortalized human esophageal epithelial cells and is a potential therapeutic target for ESCC.

Prognostic significance of PD-L1 expression in patients with gastric cancer in East Asia: a meta-analysis.

The overexpression of programmed cell death-ligand 1(PD-L1) has been observed in gastric cancer (GC). However, whether the expression of PD-L1 in tumor cells or blood serum is associated with the prognosis of patients with GC remains unclear. Therefore, we performed a meta-analysis to evaluate the prognostic significance of PD-L1 expression in GC. Electronic databases were searched systematically. Studies that met the inclusion criteria were included in the meta-analysis. Data concerning the hazard ratio (HR) for overall survival and disease-free survival with a 95% confidence interval (CI) according to the expression status of PD-L1 evaluated by immunohistochemistry or enzyme-linked immunosorbent assay were extracted. The data were analyzed using a random effects model. Subgroup analyses were proposed. Our results showed that eight studies with 950 patients met the inclusion criteria and were included in the meta-analysis. The pooled HR for overall survival indicated that patients with PD-L1-positive expression had significantly shorter survival time compared with the PD-L1-negative group (HR 1.60, 95% CI 1.09-2.36, P=0.012). The pooled HR for disease-free survival demonstrated that the difference between the two groups was not statistically significant (HR 1.02, 95% CI 0.32-3.20, P=0.98). In conclusion, our results indicate that the evaluation of PD-L1 overexpression in GC tissue or blood serum may be useful in the future as a novel prognostic factor.