Label-free OIRD detection of protein microarrays on high dielectric constant substrate with enhanced intrinsic sensitivity.

Oblique-incidence reflectivity difference (OIRD) is a dielectric constant-sensitive technique and exhibits intriguing applications in label-free and high-throughput detection of protein microarrays. With the outstanding advantage of being compatible with arbitrary substrates, however, the effect of the substrate, particularly its dielectric constant on the OIRD sensitivity has not been fully disclosed. In this paper, for the first time we investigated the dependence of OIRD sensitivity on the dielectric constant of the substrate under top-incident OIRD configuration by combining theoretical modeling and experimental evaluation. Optical modeling suggested that the higher dielectric constant substrate exhibits a higher intrinsic sensitivity. Experimentally, three substrates including glass, fluorine-doped tin oxide (FTO) and silicon (Si) with different dielectric constants were selected as microarray substrates and their detection performances were evaluated. In good agreement with the modeling, high dielectric constant Si-based microarray exhibited the highest sensitivity among three chips, reaching a detection limit of as low as 5 ng mL-1 with streptavidin as the model target. Quantification of captured targets on three chips with on-chip enzyme-linked immunosorbent assay (ELISA) further confirmed that the enhanced performance originates from the high dielectric constant enhanced intrinsic OIRD sensitivity. This work thus provides a new way to OIRD-based label-free microarrays with improved sensitivity.

Causal associations between circulating cytokines and risk of sepsis and related outcomes: a two-sample Mendelian randomization study.

Introduction: Sepsis represents a critical medical condition that arises due to an imbalanced host reaction to infection. Central to its pathophysiology are cytokines. However, observational investigations that explore the interrelationships between circulating cytokines and susceptibility to sepsis frequently encounter challenges pertaining to confounding variables and reverse causality. Methods: To elucidate the potential causal impact of cytokines on the risk of sepsis, we conducted two-sample Mendelian randomization (MR) analyses. Genetic instruments tied to circulating cytokine concentrations were sourced from genome-wide association studies encompassing 8,293 Finnish participants. We then evaluated their links with sepsis and related outcomes using summary-level data acquired from the UK Biobank, a vast multicenter cohort study involving over 500,000 European participants. Specifically, our data spanned 11,643 sepsis cases and 474,841 controls, with subsets including specific age groups, 28-day mortality, and ICU-related outcomes. Results and Discussion: MR insights intimated that reduced genetically-predicted interleukin-10 (IL-10) levels causally correlated with a heightened sepsis risk (odds ratio [OR] 0.68, 95% confidence interval [CI] 0.52-0.90, P=0.006). An inverse relationship emerged between monocyte chemoattractant protein-1 (MCP-1) and sepsis-induced mortality. Conversely, elevated macrophage inflammatory protein 1 beta (MIP1B) concentrations were positively linked with both sepsis incidence and associated mortality. These revelations underscore the causal impact of certain circulating cytokines on sepsis susceptibility and its prognosis, hinting at the therapeutic potential of modulating these cytokine levels. Additional research is essential to corroborate these connections.

NASP gene contributes to autism by epigenetic dysregulation of neural and immune pathways.

BACKGROUND: Epigenetics makes substantial contribution to the aetiology of autism spectrum disorder (ASD) and may harbour a unique opportunity to prevent the development of ASD. We aimed to identify novel epigenetic genes involved in ASD aetiology. METHODS: Trio-based whole exome sequencing was conducted on ASD families. Genome editing technique was used to knock out the candidate causal gene in a relevant cell line. ATAC-seq, ChIP-seq and RNA-seq were performed to investigate the functional impact of knockout (KO) or mutation in the candidate gene. RESULTS: We identified a novel candidate gene NASP (nuclear autoantigenic sperm protein) for epigenetic dysregulation in ASD in a Chinese nuclear family including one proband with autism and comorbid atopic disease. The de novo likely gene disruptive variant tNASP(Q289X) subjects the expression of tNASP to nonsense-mediated decay. tNASP KO increases chromatin accessibility, promotes the active promoter state of genes enriched in synaptic signalling and leads to upregulated expression of genes in the neural signalling and immune signalling pathways. Compared with wild-type tNASP, tNASP(Q289X) enhances chromatin accessibility of the genes with enriched expression in the brain. RNA-seq revealed that genes involved in neural and immune signalling are affected by the tNASP mutation, consistent with the phenotypic impact and molecular effects of nasp-1 mutations in Caenorhabditis elegans. Two additional patients with ASD were found carrying deletion or deleterious mutation in the NASP gene. CONCLUSION: We identified novel epigenetic mechanisms mediated by tNASP which may contribute to the pathogenesis of ASD and its immune comorbidity.

SQSTM1/p62 promotes the progression of gastric cancer through epithelial-mesenchymal transition.

Background: SQSTM1/p62 is an autophagy-related receptor protein that participates in regulating tumorigenesis and multiple signaling pathways. Gastric cancer (GC) is a common tumor in the digestive tract and continues to pose a significant threat to human health. Therefore, this study aims to investigate the impact of p62 on gastric cancer. Methods: Immunohistochemistry and Western blotting were employed to assess the expression level of the p62 protein in gastric cancer tissues and its correlation with prognosis. Subsequently, in vitro cell experiments were conducted to determine the role of p62 in gastric cancer cell proliferation, migration, and metastasis. Result: The expression of p62 in gastric cancer tissues was significantly higher than in normal tissues. The expression of p62 was positively correlated with poor prognosis in gastric cancer patients. In vitro cell experiments indicated that p62 promotes gastric cancer cell proliferation and migration. Mechanistically, elevated p62 expression induced epithelial-mesenchymal transition (EMT), leading to upregulation of E-cadherin and downregulation of N-cadherin and vimentin. Conclusion: This study provides novel and robust evidence for the mechanism by which elevated p62 expression promotes the progression of gastric cancer. It offers promising therapeutic targets for anti-tumor treatment strategies in gastric cancer patients.

Revealing Novel Genomic Insights and Therapeutic Targets for Juvenile Idiopathic Arthritis through Omics.

BACKGROUND: The genetic architecture of juvenile idiopathic arthritis (JIA) remains only partially comprehended. There is a clear imperative for continued endeavors to uncover insights into the underlying causes of JIA. METHODS: This study encompassed a comprehensive spectrum of endeavors, including conducting a JIA GWAS meta-analysis that incorporated data from 4,550 JIA cases and 18 446 controls. We employed in silico and genome-editing approaches to prioritize target genes. To investigate pleiotropic effects, we conducted phenome-wide association studies. Cell-type enrichment analyses were performed by integrating bulk and single-cell sequencing data. Finally, we delved into potential druggable targets for JIA. RESULTS: Fourteen genome-wide significant non-HLA loci were identified including four novel loci, each exhibiting pleiotropic associations with other autoimmune diseases or musculoskeletal traits. We uncovered strong genetic correlation between JIA and bone mineral density (BMD) traits at 52 genomic regions, including three GWAS loci for JIA. Candidate genes with immune functions were captured by in silico analyses at each novel locus, with additional findings identified through our experimental approach. Cell-type enrichment analysis revealed 21 specific immune cell types crucial for affected organs in JIA, indicating their potential contribution to the disease. Finally, 24 known or candidate druggable target genes were prioritized. CONCLUSIONS: Our identification of four novel JIA associated genes, CD247, RHOH, COLEC10 and IRF8, broadens novel potential drug repositioning opportunities. We established a new genetic link between COLEC10, TNFRSF11B and JIA/BMD. Additionally, the identification of RHOH underscores its role in positive thymocyte selection, thereby illuminating a critical facet of JIA's underlying biological mechanisms.

Target genes regulated by CLEC16A intronic region associated with common variable immunodeficiency.

BACKGROUND: CLEC16A intron 19 has been identified as a candidate locus for common variable immunodeficiency (CVID). OBJECTIVES: This study sought to elucidate the molecular mechanism by which variants at the CLEC16A intronic locus may contribute to the pathogenesis of CVID. METHODS: The investigators performed fine-mapping of the CLEC16A locus in a CVID cohort, then deleted the candidate functional SNP in T-cell lines by the CRISPR-Cas9 technique and conducted RNA-sequencing to identify target gene(s). The interactions between the CLEC16A locus and its target genes were identified using circular chromosome conformation capture. The transcription factor complexes mediating the chromatin interactions were determined by proteomic approach. The molecular pathways regulated by the CLEC16A locus were examined by RNA-sequencing and reverse phase protein array. RESULTS: This study showed that the CLEC16A locus is an enhancer regulating expression of multiple target genes including a distant gene ATF7IP2 through chromatin interactions. Distinct transcription factor complexes mediate the chromatin interactions in an allele-specific manner. Disruption of the CLEC16A locus affects the AKT signaling pathway, as well as the molecular response of CD4+ T cells to immune stimulation. CONCLUSIONS: Through multiomics and targeted experimental approaches, this study elucidated the underlying target genes and signaling pathways involved in the genetic association of CLEC16A with CVID, and highlighted plausible molecular targets for developing novel therapeutics.

Label-Free and Real-Time Optical Detection of Affinity Binding of the Antibody on Adherent Live Cells.

Oblique-incidence reflectivity difference (OIRD) is a novel real-time, label-free, and nondestructive optical detection method and exhibits encouraging application in the detection of antibody/DNA microarrays. In this study, for the first time, an OIRD label-free immunoassay was achieved by using adherent live cells as the probe. The cells were cultured on glass cells, and the affinity binding of antibodies targeted on the HLA class I antigen of the cell surface was detected with an OIRD. The results show that an OIRD is able to detect the binding process of anti-human HLA-A, B, and C antibodies on MDA-MB-231 cells and HUVEC cells. Control experiments and complementary fluorescence analysis confirmed the high detection specificity and good quantitative virtue of the OIRD label-free immunoassay. Label-free OIRD imaging analysis of cell microarrays was further demonstrated successfully, and the underlying optical mechanism was revealed by combining the theoretical modeling. This work explores the use of live cells as probes for an OIRD immunoassay, thus expanding the potential applications of the OIRD in the field of pathological analysis, disease diagnosis, and drug screening, among others.

A Lateral Flow Assay Based on Streptavidin-biotin Amplification System with Recombinase Polymerase Amplification for Rapid and Quantitative Detection of Salmonella enteritidis.

Salmonella constitutes a prevalent alimentary pathogen, instigating zoonotic afflictions. Consequently, the prompt discernment of Salmonella in sustenance is of cardinal significance. Lateral flow assays utilizing colorimetric methodologies adequately fulfill the prerequisites of point-of-care diagnostics, however, their detection threshold remains elevated, generally permitting only qualitative discernment, an impediment to the preliminary screening of nascent pathogens. In response to this conundrum, we propose a lateral flow diagnostic predicated upon a streptavidin-biotin amplification system with recombinase polymerase amplification engineered for the expeditious and quantitative discernment of Salmonella enteritidis. Trace nucleic acids within a sample undergo exponential amplification via recombinase polymerase amplification to a level discernable, constituting the initial signal amplification. Subsequently, along the test line (T-line) of the lateral flow strip, the chromatic signal undergoes augmentation by securing a greater quantity of AuNPs through the magnification capacity of the streptavidin-biotin mechanism, affecting the second signal amplification. Quantitative results are procured via smartphone capture and transferred to computer software for precise calculation of the targeted quantity. The lateral flow strip exhibits a LOD at 19.41 CFU/mL for cultured S. enteritidis. The RSD of three varying concentrations were respectively 3.74 %, 5.96 %, and 4.25 %.

Association between artificial sweeteners-aspartame consumption and colorectal cancer risk: evidence-based strategies.

In this study, clinical trials were generalized, summarized, and meta-analyzed to evaluate correlations between artificial sweeteners (ASs) and colorectal cancer (CRC). PubMed, Web of Science, EMBASE (Ovid platform), MEDLINE, and the Cochrane Library databases were searched from inception until July 24, 2023. The association between AS exposure and CRC incidence was assessed using odds ratios (ORs) and 95% confidence intervals (CIs). STATA software (version 12.0) was used to perform the meta-analysis. Ten studies (three case-control studies and seven cohort studies) involving 711,537 participants were identified. Results showed that the intake of ASs reduced the incidence of CRC (OR=0.93, 95% CI=[0.87-0.99]) and was not significantly associated with mortality (OR=0.93, 95% CI=[0.83-1.05]). Subgroup analyses showed that low doses of ASs were associated with lower CRC incidence (OR=0.90, 95% CI=[0.83-0.99]), and medium/high doses were not associated with CRC incidence (OR=1.11, 95% CI=[0.93-1.33]; OR=0.89, 95% CI=[0.79-1.00], respectively). Moreover, low, medium, and high exposures were not associated with an increased risk of mortality due to CRC (OR=0.95, 95% CI=[0.80-1.14]; OR=0.99, 95% CI=[0.88-1.11]; OR=0.93, 95% CI=[0.71-1.21], respectively). The results of our meta-analysis showed that a low intake of ASs may be associated with a lower risk of CRC.

Comprehensive evaluation of water resources carrying capacity in Henan Province based on entropy weight TOPSIS - coupling coordination - obstacle model.

This study aims to investigate the water resource carrying capacity (WRCC) of Henan Province, identify its main obstacles, and provide suggestions for optimizing its WRCC. The article constructs a WRCC evaluation system with 20 indicators for the four subsystems of water resources, economy, society, and ecology based on literature and the actual situation of Henan Province. The entropy weighted TOPSIS method is used to calculate the WRCC of Henan Province from 2005 to 2021. The coupling coordination model is used to explore the degree of coupling coordination among internal systems, while the obstacle model is used to study its restrictive influencing factors. The study found that (1) the WRCC fluctuated in a U-shaped pattern around 0.5 during the study period; (2) the coupling and coordination degree of each subsystem is generally good, except for 2012 and 2013, which showed basic coordination; (3) currently, the main obstacles to the WRCC are ecosystems and water resources. The main indicators are afforestation area, proportion of the tertiary industry, fertilizer usage, and urban sewage treatment rate. Therefore, Henan Province should take measures such as reducing fertilizer usage, standardizing urban sewage treatment, improving water efficiency, and optimizing industrial structure to optimize its WRCC and promote comprehensive utilization of water resources.

Rational Design of Mesoporous Coordination Polymer Nanophotosensitizers for Photodynamic Tumor Ablation.

Effective clinical practice of precise photodynamic therapy (PDT) is severely impeded by the inherent drawbacks and aggregation propensity of conventional photosensitizers. An all-in-one approach is highly desired to optimize structural features, photophysical properties, and pharmacokinetic behaviors of photosensitizers. Herein, we have fabricated mesoporous boron dipyrromethene-bridged coordination polymer nanophotosensitizers (BCP-NPs) for high-performance PDT via a unique solvent-assisted assembly strategy. Distinctive photophysical and structural characteristics of BCP-NPs confer enhanced photodynamic activities, together with high cellular uptake and ultrahigh stability. Moreover, BCP-NPs showed excellent tumor accumulation and prolonged tumor retention, achieving eradication of the triple-negative breast cancer (TNBC) model under low-power-density LED irradiation. This work has provided a valuable paradigm for the construction of mesoporous photoactive nanomaterials for biophotonic applications.

Non-fouling polymer brush grafted fluorine-doped tin oxide enabled optical and chemical enhancement for sensitive label-free antibody microarrays.

Oblique-incidence reflectivity difference (OIRD) is a compelling technique for real-time, label-free and non-destructive detection of antibody microarray chips, but its sensitivity needs essential improvement for clinical diagnosis. In this study, we report an innovative high-performance OIRD microarray by using poly[oligo(ethylene glycol) methacrylate-co-glycidyl methacrylate] (POEGMA-co-GMA) brush grafted fluorine-doped tin oxide (FTO) as the chip substrate. The polymer brush enhances the interfacial binding reaction efficiency of targets from the complicated sample matrix due to its high antibody loading and excellent anti-fouling merits; the FTO-polymer brush layered structure, on the other hand, excites the interference enhancement effect of OIRD to achieve enhanced intrinsic optical sensitivity. Synergistically, the sensitivity of this chip is significantly improved compared to rival chips, achieving a limit of detection (LOD) as low as 25 ng mL-1 for the model target C-reactive protein (CRP) in 10% human serum. This work explores the tremendous influence of the chip interfacial structure on the OIRD sensitivity and proposes a rational interfacial engineering strategy to boost the performance of the label-free OIRD based microarray and other bio-devices.

Influencing Factors of Bidens pilosa L. Hyperaccumulating Cadmium Explored by the Real-Time Uptake of Cd2+ Influx around Root Apexes under Different Exogenous Nutrient Ion Levels.

Though Bidens pilosa L. has been confirmed to be a potential Cd hyperaccumulator, the accumulation mechanism is not yet clear. The dynamic and real-time uptake of Cd2+ influx by B. pilosa root apexes was determined using non-invasive micro-test technology (NMT), which partly explored the influencing factors of the Cd hyperaccumulation mechanism under the conditions of different exogenous nutrient ions. The results indicated that Cd2+ influxes at 300 µm around the root tips decreased under Cd treatments with 16 mM Ca2+, 8 mM Mg2+, 0.5 mM Fe2+, 8 mM SO42- or 18 mM K+ compared to single Cd treatments. The Cd treatments with a high concentration of nutrient ions showed an antagonistic effect on Cd2+ uptake. However, Cd treatments with 1 mM Ca2+, 0.5 mM Mg2+, 0.5 mM SO42- or 2 mM K+ had no effect on the Cd2+ influxes as compared with single Cd treatments. It is worth noting that the Cd treatment with 0.05 mM Fe2+ markedly increased Cd2+ influxes. The addition of 0.05 mM Fe2+ exhibited a synergistic effect on Cd uptake, which could be low concentration Fe2+ rarely involved in blocking Cd2+ influx and often forming an oxide membrane on the root surface to help the Cd uptake by B. pilosa. The results also showed that Cd treatments with high concentration of nutrient ions significantly increased the concentrations of chlorophyll and carotenoid in leaves and the root vigor of B. pilosa relative to single Cd treatments. Our research provides novel perspectives with respect to Cd uptake dynamic characteristics by B. pilosa roots under different exogenous nutrient ion levels, and shows that the addition of 0.05 mM Fe2+ could promote the phytoremediation efficiency for B. pilosa.

The effects of salinity and pH variation on hyperaccumulator Bidens pilosa L. accumulating cadmium with dynamic and real-time uptake of Cd2+ influx around its root apexes.

Bidens pilosa L. has been confirmed to be a potential Cd hyperaccumulator by some researchers, but the dynamic and real-time uptake of Cd2+ influx by B. pilosa root apexes was a conundrum up to now. The aim of our study was to investigate the effects of salinity and pH variations on the characteristics of Cd2+ influx around the root apexes of B. pilosa. The tested seedlings of B. pilosa were obtained by sand culture experiments in a greenhouse after 1 month from germination, and the Cd2+ influxes from the root apex of B. pilosa under Cd treatments with different salinity and pH levels were determined with application of non-invasive micro-test technology (NMT). The results showed that Cd2+ influxes at 300 µm from the root tips decreased under Cd treatments with 5 mM and 10 mM NaCl, as compared to Cd stress alone. However, Cd treatments with 2.5 mM NaCl had little effect on the net Cd2+ influxes, as compared to Cd treatments alone. Importantly, Cd treatments at pH = 4.0 markedly increased Cd2+ influxes in roots, and Cd treatment at pH = 7.0 had no significant effect on the net Cd2+ influxes compared to Cd treatments at pH = 5.5. Results also showed that Cd treatments with 10 mM NaCl significantly decreased concentrations of chlorophyll (Chl) a and b in leaves and root vigor of B. pilosa relative to Cd treatments alone, while there were no significant differences between Cd treatments with 2.5 mM NaCl and Cd treatments alone. But root vigor was inhibited significantly under Cd treatments with 5 mM and 10 mM NaCl. A significant increase of root vigor was observed in Cd treatments at pH = 4.0, as compared to pH = 5.5. The Cd treatments with high and medium concentrations of NaCl inhibited the uptake of Cd by B. pilosa roots and affected the Chl and root vigor further. But the Cd treatments at pH = 4.0 could promote the Cd uptake and root vigor. Our results revealed the uptake mechanisms of B. pilosa as a potential phytoremediator under different salinity and pH levels combined with Cd contamination and provided a new idea for screening ideal hyperaccumulator and constructing evaluation system.

Ammonia fiber expansion-assisted deep eutectic solvent treatment for wheat straw fraction separation and bioconversion.

In this study, an ammonia fiber expansion (AFEX)-assisted deep eutectic solvent (DES) pretreatment method was developed for the rapid separation of wheat straw fractions, which reduced the pretreatment time for DES and improved the pretreatment efficiency. This study describes the feasibility of the AFEX-assisted DES pretreatment in terms of both progressive and parallel relationships and analyzes the subsequent enzymatic effect in generating glucose from cellulose. Ammonia fiber expansion-assisted DES one-pot pretreatment at 120 °C, for 1.5 h resulted in an enzymatic efficiency of 98.0 ± 3.1 %. Moreover, the enzyme efficiency remained greater than 85 % after three recovery cycle experiments. The comparison between regenerated-lignin (d-lignin) and alkaline-lignin showed that regenerated lignin has a lower molecular weight and belongs to para-hydroxy-phenyl-guaiacyl-syringyl (H-G-S) type lignin. This study developed is a green and efficient pretreatment process with great potential in the separation and utilization of biomass fractions.

Effect of Ammoniated Fiber Explosion Combined with H2O2 Pretreatment on the Hydrogen Production Capacity of Herbaceous and Woody Waste.

An appropriate pretreatment process is an important part of the preparation of biomass energy from agricultural and forestry waste. Compared to physical and chemical pretreatments alone, the combined ammoniated fiber explosion (AFEX) + hydrogen peroxide (H2O2) pretreatment process can significantly improve the lignin degradation rate and saccharification efficiency, thus improving the hydrogen production capacity during medium-temperature dark fermentation. This study showed that the combined pretreatment increased the saccharification efficiency of herbaceous, hardwood, and softwood biomass by 58.7, 39.5, and 20.6% and the corresponding gas production reached 145.49, 80.75, and 57.52 mL/g, respectively. In addition, X-ray diffraction, scanning electron microscopy, and Fourier-transform infrared spectroscopy showed that AFEX + H2O2 disrupted the structure of the feedstock and was more favorable for lignin removal. Soluble metabolites indicated that AFEX + H2O2 pretreatment enhanced the butyrate metabolic pathway of the substrate and biohydrogen generation and increased the levels of extracellular polymers and microbial community structure.

Competition-Induced Binding Spherical Nucleic AcidFluorescence Amplifier for the Detection of Di (2-ethylhexyl) Phthalate in the Aquatic Environment.

Di-2-ethylhexyl phthalate (DEHP) is a toxic plasticizer and androgen antagonist. Its accumulation in water exceeds national drinking water standards and it must be continuously and effectively regulated. Currently, methods used to detect DEHP are still unsatisfactory because they usually have limited detection sensitivity and require complex operating procedures. A competition-induced fluorescence detection method was developed for the selective detection of DEHP in an aquatic environment. An aptamer with walking function was used as the recognition element for DEHP, and its quantification was induced by competition to change the fluorescence signal. The detection range was 0.01~100 µg/L, and the detection limit was 1.008 µg/L. This high-sensitivity DEHP detection capability and simplified process facilitates real-time fields and other monitoring tasks.

Effect of Ammonia Fiber Expansion Combined with NaOH Pretreatment on the Resource Efficiency of Herbaceous and Woody Lignocellulosic Biomass.

The most essential issue facing the world today is the provision of energy and sustainable consumption of natural resources. Pretreatment is an essential step to produce biofuels from lignocellulosic biomass. In this study, ammonia fiber explosion (AFEX) combined with NaOH (A-NaOH) pretreatment effects on the characteristics of Pennisetum sinese (herbaceous), oak (hardwood), and camphor wood (softwood) were assessed using enzymatic efficiency analysis, thereby identifying the composition properties of subsequent bio-H2 production. The results show that the lignin removal (84.2%, 59.7%, and 36.7%, respectively) at 5%A-NaOH conditions and enzymatic efficiency (36.2%, 9.7%, and 6.5%, respectively) of Pennisetum sinese (P. sinese), oak, and camphor wood were significantly increased under 4% A-NaOH conditions. Further A-NaOH pretreatment significantly promoted dark fermentation bio-H2 production (152.3, 99.1, and 76.9 mL/g TS, respectively) and volatile acid production (4660.2, 3720.2, and 3496.2 mg/L, respectively) of P. sinese, oak, and camphor wood. These findings show that A-NaOH pretreatment is an effective means of utilization of lignocellulose resources.

MiR-494-3p Upregulation Exacerbates Cerebral Ischemia Injury by Targeting Bhlhe40.

PURPOSE: Cerebral ischemia is related to insufficient blood supply and is characterized by abnormal reactive oxygen species (ROS) production and cell apoptosis. Previous studies have revealed a key role for basic helix-loop-helix family member e40 (Bhlhe40) in oxidative stress and cell apoptosis. This study aimed to investigate the roles of miR-494-3p in cerebral ischemia/reperfusion (I/R) injury. MATERIALS AND METHODS: A mouse middle cerebral artery occlusion (MCAO/R) model was established to mimic cerebral ischemia in vivo. Brain infarct area was assessed using triphenyl tetrazolium chloride staining. Oxygen-glucose deprivation/reoxygenation (OGD/R) operation was adopted to mimic neuronal injury in vitro. Cell apoptosis was analyzed by flow cytometry. The relationship between miR-494-3p and Bhlhe40 was validated by luciferase reporter and RNA immunoprecipitation assays. RESULTS: Bhlhe40 expression was downregulated both in MCAO/R animal models and OGD/R-induced SH-SY5Y cells. Bhlhe40 overexpression inhibited cell apoptosis and reduced ROS production in SH-SY5Y cells after OGD/R treatment. MiR-494-3p was verified to bind to Bhlhe40 and negatively regulate Bhlhe40 expression. Additionally, cell apoptosis and ROS production in OGD/R-treated SH-SY5Y cells were accelerated by miR-494-3p overexpression. Rescue experiments suggested that Bhlhe40 could reverse the effects of miR-494-3p overexpression on ROS production and cell apoptosis. CONCLUSION: MiR-494-3p exacerbates brain injury and neuronal injury by regulating Bhlhe40 after I/R.