The NeuroProtect Formula: A Preventive Approach to AD Targeting the HIF-1/PI3K-AKT Signaling Pathway Evaluated through In Vivo, In Vitro, and Network Pharmacology Approaches.

OBJECTIVE: Alzheimer's Disease (AD) is a progressive neurodegenerative disorder with limited options for reversing its middle-to-late stages. Early intervention is crucial to slow down disease progression. This study aimed to investigate the potential of the NeuroProtect (NP) formula, a combination of geniposide and Panax notoginseng saponins, in preventing AD. We evaluated the effects of the NP formula on amyloid plaque accumulation, neuronal degeneration, and molecular signaling pathways using in vivo and in vitro models. METHODS: To predict functional pathways and potential downstream targets of NP intervention, we employed network pharmacology. The preventative impact of the NP formula was assessed using APP/PS1 mice. We conducted HE staining, ELISA assay, Golgi staining, and immunohistochemistry to detect the protective effect of NP. Additionally, cell experiments were performed to assess cell activity and target protein expression. RESULTS: Network pharmacology analysis revealed 145 drug-disease interactions and identified 5 core active targets associated with AD. Molecular docking results demonstrated strong binding affinity between the components of the NP formula (GP, GN-Rb1, GN-Rg1, NS-R1) and target proteins (STAT3, HIF1A, TLR4, mTOR, VEGFA). Notably, the binding energy between NS-R1 and mTOR was -11.4kcal/mol. Among the top 10 enriched KEGG pathways, the HIF-1 and PI3K-AKT signaling pathways were highlighted. In vivo experiments demonstrated that the NP formula significantly ameliorated pathological changes, decreased the Aß42/Aß40 ratio in the hippocampus and cortex, and increased dendritic spine density in the CA1 region during the early stage of AD. In vitro experiments further illustrated the NP formula's ability to reverse the inhibitory effects of Aß25-35 on cell viability and regulate the expression of Tlr4, Mtor, Hif1a, Stat3, and Vegfa. CONCLUSION: Our findings suggest that NP exhibits neuroprotective effects during the early stages of AD, positioning it as a potential candidate for AD prevention. The NP formula may exert its preventive effects through the HIF-1/PI3K-AKT signaling pathway, with mTOR identified as a key target.

Nitrogen Minimization in Hydrothermal Liquefaction Biocrude from Sewage Sludge with Green Extraction Solvents.

This study explored the effectiveness of hydrothermal liquefaction (HTL) in converting sewage sludge (SS) into high-quality biocrude. It scrutinized the influence of various solvents, including conventional choices like dichloromethane (DCM) and hexane, alongside environmentally friendly alternatives, such as ethyl butyrate (EB) and ethyl acetate (EA). HTL experiments, conducted at 350 °C for 60 min in a 20 mL batch reactor, include solvent-based biocrude extraction. Notably, EB showed the highest extraction yield (50.1 wt %), the lowest nitrogen distribution (5.4% with 0.32 wt %), and a remarkable 74% energy recovery (ER), setting a noteworthy benchmark in nitrogen reduction. GCMS analysis reveals EB-derived biocrude's superiority in having the least heteroatoms and nitrogenous compounds compared to hexane, EA, and DCM. Solid residues from hexane, EB, and EA displayed the highest nitrogen distribution range (62-68%), hinting at potential applications in further processes. These findings significantly inform solvent selection for efficient and sustainable waste-to-energy conversion. While promising, the study emphasizes the need to explore solvent-solute interactions further to optimize biocrude quality, highlighting the pivotal role of solvent choice in advancing clean, cost-effective waste-to-energy technologies.

Cross-Linkable Fullerene Enables Elastic and Conductive Grain Boundaries for Efficient and Wearable Tin-Based Perovskite Solar Cells.

Tin-based perovskite solar cells (TPSCs) have received increasing attention due to their low toxicity, high theoretical efficiency, and potential applications as wearable devices. However, the inherent fast and uncontrollable crystallization process of tin-based perovskites results in high defect density in the film. Meanwhile, when fabricated into flexible devices, the prepared perovskite film exhibits inevitable brittleness and high Young's modulus, seriously weakening the mechanical stability. In this work, we design and synthesize a cross-linkable fullerene, thioctic acid functionalized C60 fulleropyrrolidinium iodide (FTAI), which has multiple interactions with perovskite components and can finely regulate the crystallization quality of perovskite film. The obtained perovskite film shows an increased grain size and a more matched energy level with the electron transport material, effectively improving the carrier extraction efficiency. The FTAI-based rigid device achieves a champion efficiency of 14.91 % with enhanced stability. More importantly, the FTAI located at the perovskite grain boundaries could spontaneously cross-link during the perovskite annealing process, which effectively improves the conductivity and elasticity of grain boundaries, thereby giving the film excellent bending resistance. Finally, the FTAI-based wearable device yields a record efficiency of 12.35 % and displays robust bending durability, retaining about 90 % of the initial efficiency after 10,000 bending times.

Pseudoamaolide P, a 1,2:3,4:9,10:9,19-tetraseco-cycloartane triterpene spiroketal lactone from seeds of Pseudolarix amabilis.

A 1,2:3,4:9,10:9,19-tetraseco-cycloartane triterpene spiroketal lactone, pseudoamaolide P (1), two new labdane-type diterpenoids, pseudoamains A and B (2-3), and four known cembrane-type diterpenoids (4-7) were isolated from the seeds of Pseudolarix amabilis. The structures of these compounds were elucidated by spectroscopic analyses, including HRESIMS, 1D-, and 2D-NMR. The anti-inflammatory activities of the compounds were evaluated by suppressing the transcription of the NF-κB-dependent reporter gene in LPS-induced 293 T/NF-κB-luc cells. All compounds do not show potent activity.

GLS and GOT2 as prognostic biomarkers associated with dendritic cell and immunotherapy response in breast cancer.

Breast cancer is the females' most common cancer. Targeting the immune microenvironment is a new and promising treatment method for breast cancer. Nevertheless, only a small section of patients can profit by immunotherapy, and improving the ability to accurately predict the potential for immunotherapy response is still awaiting further exploration. In this study, we found that the key factors of glutamine metabolism, glutaminase 1 (GLS) and mitochondrial aspartate transaminase (GOT2), showed opposite expression patterns in breast cancer samples. Based on the expression level of GLS and GOT2, we divided the breast cancer samples into two clusters: Cluster 2 showed GLS expressed higher and GOT2 expressed lower, whereas Cluster 1 showed GOT2 expressed higher and GLS expressed lower. GSEA showed that the clusters were related to pathways of immunity. Further analysis showed that Cluster 2 was positively associated with immunity infiltration. Through WGCNA, we identified a module strongly correlated with glutamine metabolism and immunity and identified 11 dendritic cell-associated genes involved in dendritic cell development, maturation, activation and other functions. In addition, Cluster 2 also showed higher immune checkpoint gene expression, which suggest the Cluster 2 had even better response to immunotherapy. The validation dataset could also be clustered into two groups. Cluster 2 (GLS expressed higher and GOT2 expressed lower) of the validation dataset was also positively associated with dendritic cells and a better immunotherapy response. Thus, these data indicate that GLS and GOT2 are prognostic biomarkers which closely related to dendritic cells and better reacted to immunotherapy in breast cancer.

Gastrodin alleviates NTG-induced migraine-like pain via inhibiting succinate/HIF-1α/TRPM2 signaling pathway in trigeminal ganglion.

BACKGROUND: Increasing evidence highlights the involvement of metabolic disorder and calcium influx mediated by transient receptor potential channels in migraine; however, the relationship between these factors in the pathophysiology of migraine remains unknown. Gastrodin is the major component of the traditional Chinese medicine Tianma, which is extensively used in migraine therapy. PURPOSE: Our work aimed to explore the analgesic action of gastrodin and its regulatory mechanisms from a metabolic perspective. METHODS/RESULTS: After being treated with gastrodin, the mice were given nitroglycerin (NTG) to induce migraine. Gastrodin treatment significantly raised the threshold of sensitivity in response to both mechanical and thermal stimulus evidenced by von Frey and hot plate tests, respectively, and decreased total contact numbers in orofacial operant behavioral assessment. We found that the expression of transient receptor potential melastatin 2 (TRPM2) channel was increased in the trigeminal ganglion (TG) of NTG-induced mice, resulting in a sustained Ca2+ influx to trigger migraine pain. The content of succinate, a metabolic biomarker, was elevated in blood samples of migraineurs, as well as in the serum and TG tissue from NTG-induced migraine mice. Calcium imaging assay indicated that succinate insult elevated TRPM2-mediated calcium flux signal in TG neurons. Mechanistically, accumulated succinate upregulated hypoxia inducible factor-1α (HIF-1α) expression and promoted its translocation into nucleus, where HIF-1α enhanced TRPM2 expression through transcriptional induction in TG neurons, evidenced by luciferase reporter measurement. Gastrodin treatment inhibited TRPM2 expression and TRPM2-dependent Ca2+ influx by attenuating succinate accumulation and downstream HIF-1α signaling, and thereby exhibited analgesic effect. CONCLUSION: This work revealed that succinate was a critical metabolic signaling molecule and the key mediator of migraine pain through triggering TRPM2-mediated calcium overload. Gastrodin alleviated NTG-induced migraine-like pain via inhibiting succinate/HIF-1α/TRPM2 signaling pathway in TG neurons. These findings uncovered the anti-migraine effect of gastrodin and its regulatory mechanisms from a metabolic perspective and provided a novel theoretical basis for the analgesic action of gastrodin.

Relation between hydrophilic/hydrophobic characteristics of sludge extracellular polymeric substances and sludge moisture-holding capacity in hot-pressing drying.

Sludge poses a serious threat to the environmental health. Hot-pressing drying has been proven efficient in sludge treatment because of the reduced thermal contact resistance, rapid increase in sludge temperature, and high drying rate. Sludge extracellular polymeric substances (EPS) significantly influence moisture transfer. However, whether in hot-pressing or traditional thermal drying, the effect of EPS on sludge moisture-holding capacity is rarely reported. Thereby, this study investigated the relationship between hydrophilic/hydrophobic characteristics of EPS and sludge moisture-holding capacity at various drying time and mechanical compression using XAD resin fractionation. Thermodynamic analysis indicated that sludge moisture desorption isotherms, net isosteric heat of desorption, and differential entropy presented a downward trend with the increase in drying time and mechanical compression, suggesting reduced sludge moisture-holding capacity. EPS analysis showed that at the same drying time, applying 25 kPa mechanical compression increased sludge temperature by 16 % and protein content by 13.8 %. At the same sludge temperature, protein content rose by 7.3 % compared to the drying without mechanical compression. It was concluded that the fast rise in sludge temperature and the mechanical extrusion facilitated the destruction of sludge microbial flocs, accelerating the release of intracellular and EPS-bound moisture and contributing to the decrease in moisture-holding capacity. Besides, tryptophan protein-like substances were the major source of hydrophilic/hydrophobic organic matter, compared to polysaccharide and humic acid-like substances. The gradually reduced sludge moisture-holding capacity was divided into three stages. Below 67 °C, the moisture desorption was dominated by the release of intracellular moisture. Below 85 °C, the increase in protein and the enhanced exposure of hydrophobic functional groups in protein improved the hydrophobicity of EPS. Above 85 °C, protein consumption due to thermal decomposition and browning reaction facilitated the desorption of EPS-bound moisture. Hence, this study provided novel insights into the mechanism of sludge drying.

Adverse Effects of Gefitinib on Skin and Colon in a Lung Cancer Mouse Model.

BACKGROUND: Gefitinib, an Epidermal Growth Factor Receptor Tyrosine Kinase Inhibitor (EGFR-TKI), frequently causes side effects when used to treat non-small cell lung cancer. OBJECTIVE: The purpose of this experiment was to investigate the side effect of gefitinib on the skin and colon of mice. METHODS: Male Balb/c nu-nu nude mice aged 4-5 weeks were used as xenograft tumor models, and gefitinib at 150 mg/kg and 225 mg/kg was started at 9 days after the xenograft tumor grew out. The mice's weights and tumor volumes were tracked concurrently, and the mouse skin adverse reactions and diarrhea were observed during the treatment. The animal tissues were subjected to biochemical and pathological evaluations after 14 days. RESULTS: Gefitinib effectively decreased the size and weight of transplanted tumors in nude mice, while also lowering body weight and raising indexes of the liver and spleen. Gefitinib could cause skin adverse reactions and diarrhea in mice. Further pathological investigation revealed tight junction- related markers in the mice's skin and colon to be reduced and macrophages and neutrophils to be increased after gefitinib treatment. CONCLUSION: The findings imply that gefitinib has negative effects on the skin and colon. Gefitinib- induced skin and colon adverse reactions in mice have been successfully modeled in this study.

Efficient Tin-Based Perovskite Solar Cells Enabled by Precisely Synthesized Single-Isomer Fullerene Bisadducts with Regulated Molecular Packing.

Designing and synthesizing fullerene bisadducts with a higher-lying conduction band minimum is promising to further improve the device performance of tin-based perovskite solar cells (TPSCs). However, the commonly obtained fullerene bisadduct products are isomeric mixtures and require complicated separation. Moreover, the isomeric mixtures are prone to resulting in energy alignment disorders, interfacial charge loss, and limited device performance improvement. Herein, we synthesized single-isomer C60- and C70-based diethylmalonate functionalized bisadducts (C60BB and C70BB) by utilizing the steric-hindrance-assisted strategy and determined all molecular structures involved by single crystal diffraction. Meanwhile, we found that the different solvents used for processing the fullerene bisadducts can effectively regulate the molecular packing in their films. The dense and amorphous fullerene bisadduct films prepared by using anisole exhibited the highest electron mobility. Finally, C60BB- and C70BB-based TPSCs showed impressive efficiencies up to 14.51 and 14.28%, respectively. These devices also exhibited excellent long-term stability. This work highlights the importance of developing strategies to synthesize single-isomer fullerene bisadducts and regulate their molecular packing to improve TPSCs' performance.

Histidine Triad Nucleotide-Binding Protein 1 Improves Critical Limb Ischemia by Regulating Mitochondrial Homeostasis.

Critical limb ischemia (CLI) is a common complication of diabetes mellitus that typically occurs in the later stages of the disease. Vascularization is indeed an important physiological process involving the formation of new blood vessels from existing ones. It occurs in response to various normal and pathophysiological conditions, and one of its critical roles is to compensate for inadequate oxygen supply, which is often seen in situations like chronic limb ischemia (CLI). Histidine triad nucleotide-binding protein 1 (Hint1) is a member of the Hint family that has been shown to attenuate cardiac hypertrophy, but its role in vascularization still needs to be clarified. In this study, we investigated the role of Hint1 in CLI. We found that Hint1 is significantly reduced in the muscle tissue of STZ-induced diabetic mice and high-glucose (HG)-treated endothelial cells (ECs). Hint1 deletion impaired blood flow recovery and vascularization, whereas Hint1 overexpression promoted these processes. In addition, our in vitro study showed that Hint1 deficiency aggravated mitochondrial dysfunction in ECs, as evidenced by impaired mitochondrial respiration, decreased mitochondrial membrane potential, and increased reactive oxygen species. Our findings suggest that Hint1 deficiency impairs blood perfusion by damaging mitochondrial function and that Hint1 may represent a potential therapeutic target for treating CLI.

Preparation and SERS applications of Ta2O5 composite nanostructures.

Noble metal and semiconductor composite substrates possess high sensitivity, excellent stability, good biocompatibility, and selective enhancement, making them an important research direction in the field of surface-enhanced Raman scattering (SERS). Ta2O5, as a semiconductor material with high thermal stability, corrosion resistance, outstanding optical properties, and catalytic performance, has great potential in SERS research. This study aims to design and fabricate a composite SERS substrate based on Ta2O5 nanostructures, achieving optimal detection performance by combining the urchin-like structure of Ta2O5 with silver nanoparticles (Ag NPs). The urchin-like Ta2O5 nanostructures were prepared using a hydrothermal reaction method. The bandgap was modulated through structure design and the self-doping technique, the charge transfer efficiency and surface plasmon resonance effects were improved, thereby achieving better SERS performance. The composite substrate enables highly sensitive quantitative detection. This composite SERS substrate combines the electromagnetic enhancement mechanism (EM) and chemical enhancement mechanism (CM), achieving ultra-low detection limits of 10-13 M for R6G. Within the concentration range above 10-12 M, there is a good linear relationship between concentration and peak intensity, demonstrating excellent quantitative analysis capabilities. Furthermore, this composite SERS substrate is capable of precise detection of analytes such as crystal violet (CV) and methylene blue (MB), holding broad application prospects in areas such as food safety and environmental monitoring.

Prediction of Higher Heating Values in Bio-Oil from Solvothermal Biomass Conversion and Bio-Oil Upgrading Given Discontinuous Experimental Conditions.

Both the conversion of lignocellulosic biomass to bio-oil (BO) and the upgrading of BO have been the targets of many studies. Due to the large diversity and discontinuity seen in terms of reaction conditions, catalysts, solvents, and feedstock properties that have been used, a comparison across different publications is difficult. In this study, machine learning modeling is used for the prediction of final higher heating value (HHV) and ΔHHV for the conversion of lignocellulosic feedstocks to BO, and BO upgrading. The models achieved coefficient of determination (R2) scores ranging from 0.77 to 0.86, and the SHapley Additive exPlanations (SHAP) values were used to obtain model explainability, revealing that only a few experimental parameters are largely responsible for the outcome of the experiments. In particular, process temperature and reaction time were overwhelmingly responsible for the majority of the predictions, for both final HHV and ΔHHV. Elemental composition of the starting feedstock or BO dictated the upper possible HHV value obtained after the experiment, which is in line with what is known from previous methodologies for calculating HHV for fuels. Solvent used, initial moisture concentration in BO, and catalyst active phase showed low predicting power, within the context of the data set used. The results of this study highlight experimental conditions and variables that could be candidates for the creation of minimum reporting guidelines for future studies in such a way that machine learning can be fully harnessed.

Machine Learning Model Insights into Base-Catalyzed Hydrothermal Lignin Depolymerization.

Lignin, an abundant component of plant matter, can be depolymerized into renewable aromatic chemicals and biofuels but remains underutilized. Homogeneously catalyzed depolymerization in water has gained attention due to its economic feasibility and performance but suffers from inconsistently reported yields of bio-oil and solid residues. In this study, machine learning methods were used to develop predictive models for bio-oil and solid residue yields by using a few reaction variables and were subsequently validated by doing experimental work and comparing the predictions to the results. The models achieved a coefficient of determination (R2) score of 0.83 and 0.76, respectively, for bio-oil yield and solid residue. Variable importance for each model was calculated by two different methodologies and was tied to existing studies to explain the model predictive behavior. Based on the outcome of the study, the creation of concrete guidelines for reporting in lignin depolymerization studies was recommended. Shapley additive explanation value analysis reveals that temperature and reaction time are generally the strongest predictors of experimental outcomes compared to the rest.

Effects of virgin and BaP-adsorbed microplastics ingestion by Manila clams (Ruditapes philippinarum).

Numerous microplastic-related studies have investigated the impact of plastic materials on the marine food chain. In this study, Manila clams were exposed to microplastic (MP) of various polymer types, shapes, and concentrations to determine the ingestion selectivity and adverse effects caused. Benzo[a]pyrene was introduced as the second stressor to investigate the role of MP as a vector of contaminant. The result of a 2-day acute exposure showed that clams are more likely to ingest those in sphere shapes due to their similarity to microalgae. The feeding rate continuously declined when clams were exposed to at least 2to/L particles. Additionally, co-exposure of MP and B[a]P resulted in higher DNA fragmentation but lower catalase activity compared to single exposure to MP. Our study revealed that the uptake of MP by clams is not only determined by its shape and concentration but also by the presence of existing contaminants.

Dianthrone derivatives from Polygonum multiflorum Thunb: Anti-diabetic activity, structure-activity relationships (SARs), and mode of action.

PTP1B plays an important role as a key negative regulator of tyrosine phosphorylation associated with insulin receptor signaling in the therapy for diabetes and obesity. In this study, the anti-diabetic activity of dianthrone derivatives from Polygonum multiflorum Thunb., as well as the structure-activity relationships, mechanism, and molecular docking were explored. Among these analogs, trans-emodin dianthrone (compound 1) enhances insulin sensitivity by upregulating the insulin signaling pathway in HepG2 cells and displays considerable anti-diabetic activity in db/db mice. By using photoaffinity labeling and mass spectrometry-based proteomics, we discovered that trans-emodin dianthrone (compound 1) may bind to PTP1B allosteric pocket at helix α6/α7, which provides fresh insight into the identification of novel anti-diabetic agents.

Convolutional Neural Network-Based Lane-Change Strategy via Motion Image Representation for Automated and Connected Vehicles.

The lane-change decision-making module of automated and connected vehicles (ACVs) is one of the most crucial and challenging issues to be addressed. Motivated by human beings' underlying driving paradigm and the convolutional neural network's (CNN) dramatic capability of extracting features and learning strategies, this article proposes a CNN-based lane-change decision-making method via the dynamic motion image representation. Human drivers take proper driving maneuvers after they subconsciously construct the dynamic traffic scene representation in their brains, so this study first proposes the dynamic motion image representation method to reveal informative traffic situations in the motion-sensitive area (MSA), which provides a full view of surrounding cars. Then, this article develops a CNN model to extract the underlying features and learn driving policies from labeled datasets of MSA motion images. Besides, a safety-constrained layer is added to avoid vehicle collisions. We build a simulation platform based on the simulation of urban mobility (SUMO) to collect traffic datasets and test our proposed method. In addition, real-world traffic datasets are also involved to further investigate the proposed method's performance. The rule-based strategy and reinforcement learning (RL)-based method are used to compare with our approach. All results demonstrate that the proposed method performs lane-change decision-making much better than prevailing methods, which suggests our scheme has huge potential to accelerate the deployment of ACVs and is worth further study.

Switchable MPC-based multi-objective regenerative brake control via flow regulation for electric vehicles.

Recent investigations of the electric braking booster (E-Booster) focus on its potential to enhance brake energy regeneration. A vehicle's hydraulic system is composed of the E-Booster and electric stability control to control the master cylinder and wheel cylinders. This paper aims to address the independent closed-loop control of the position and pressure as well as the maintenance of the pedal feel. To track both the reference signals related to piston displacement and the wheel cylinder pressure, an explicit model predictive control (MPC) is developed. First, the new flow model is introduced as the foundation for controller design and simulation. Next, in accordance with the operational conditions, the entire system is divided into three switchable subsystems. The three distributed MPCs are constructed based on the linearized subsystems, and a state machine is used to perform the state jump across the controllers. A linear piecewise affine control law can then be obtained by solving the quadratic program (QP) of explicit MPC. Afterwards, the non-linear extended Kalman filter including the recorded time-variant process noise is used to estimate all the state variables. The effectiveness of the explicit MPC is evidenced by the simulations compared with a single MPC in regenerative and dead-zone conditions. The proposed controller decreases the latency significantly by 85 milliseconds, which also helps to improve accuracy by 22.6%. Furthermore, the pedal feel remains consistent, even when factoring in the number of vibrations caused by the inherent hydraulic characteristic of pressure versus volume.

[Diterpenoid constituents in Pseudolarix amabilis and their antitumor activities in vitro].

By various chromatographic techniques and extensive spectroscopic methods, 17 abietane diterpenoids were isolated from the dichloromethane fraction of the 95% ethanol cold-soak extracts of the seeds of Pseudolarix amabilis, namely pseudoamaol A(1), 12α-hydroxyabietic acid(2), 12-methoxy-7,13-abietadien-18-oic acid(3), 13-hydroxy-8,11,13-podocarpatrien-18-oic acid(4), 15-hydroxy-7,13-abietadien-12-on-18-oic acid(5), 8(14)-podocarpen-13-on-18-oic acid(6), holophyllin K(7), metaglyptin B(8), 7α-hydroxydehydroabietinsaure-methylester(9), 7-oxodehydroabietic acid(10), 15-hydroxy-7-oxodehydroabietinsaure-methy-lester(11), 15-methoxydidehydroabietic acid(12), 7-oxo-15-hydroxy-dehydroabietic acid(13), 15-hydroxydehydroabietic acid(14), 8,11,13-abietatriene-15,18-diol(15), 8,11,13-abietatriene-15-hydroxy-18-succinic acid(16), and 7ß-hydroxydehydroabie-tic acid(17). Compound 1 was a new compound. The isolated compounds were evaluated for their antitumor activities(HepG2, SH-SY5Y, K562), and compounds 8 and 17 showed potential cytotoxic activity against K562 cells, with IC_(50) values of 26.77 and 37.35 µmol·L~(-1), respectively.

Activation of retinal glial cells contributes to the degeneration of ganglion cells in experimental glaucoma.

Elevation of intraocular pressure (IOP) is a major risk factor for neurodegeneration in glaucoma. Glial cells, which play an important role in normal functioning of retinal neurons, are well involved into retinal ganglion cell (RGC) degeneration in experimental glaucoma animal models generated by elevated IOP. In response to elevated IOP, mGluR I is first activated and Kir4.1 channels are subsequently inhibited, which leads to the activation of Müller cells. Müller cell activation is followed by a complex process, including proliferation, release of inflammatory and growth factors (gliosis). Gliosis is further regulated by several factors. Activated Müller cells contribute to RGC degeneration through generating glutamate receptor-mediated excitotoxicity, releasing cytotoxic factors and inducing microglia activation. Elevated IOP activates microglia, and following morphological and functional changes, these cells, as resident immune cells in the retina, show adaptive immune responses, including an enhanced release of pro-inflammatory factors (tumor neurosis factor-α, interleukins, etc.). These ATP and Toll-like receptor-mediated responses are further regulated by heat shock proteins, CD200R, chemokine receptors, and metabotropic purinergic receptors, may aggravate RGC loss. In the optic nerve head, astrogliosis is initiated and regulated by a complex reaction process, including purines, transmitters, chemokines, growth factors and cytokines, which contributes to RGC axon injury through releasing pro-inflammatory factors and changing extracellular matrix in glaucoma. The effects of activated glial cells on RGCs are further modified by the interplay among different types of glial cells. This review is concluded by presenting an in-depth discussion of possible research directions in this field in the future.

EphA4/ephrinA3 reverse signaling mediated downregulation of glutamate transporter GLAST in Müller cells in an experimental glaucoma model.

Deficiency of glutamate transporter GLAST in Müller cells may be culpable for excessive extracellular glutamate, which involves in retinal ganglion cell (RGC) damage in glaucoma. We elucidated how GLAST was regulated in rat chronic ocular hypertension (COH) model. Western blot and whole-cell patch-clamp recordings showed that GLAST proteins and GLAST-mediated current densities in Müller cells were downregulated at the early stages of COH. In normal rats, intravitreal injection of the ephrinA3 activator EphA4-Fc mimicked the changes of GLAST in COH retinas. In purified cultured Müller cells, EphA4-Fc treatment reduced GLAST expression at mRNA and protein levels, which was reversed by the tyrosine kinase inhibitor PP2 or transfection with ephrinA3-siRNA (Si-EFNA3), suggesting that EphA4/ephrinA3 reverse signaling mediated GLAST downregulation. EphA4/ephrinA3 reverse signaling-induced GLAST downregulation was mediated by inhibiting PI3K/Akt/NF-κB pathways since EphA4-Fc treatment of cultured Müller cells reduced the levels of p-Akt/Akt and NF-κB p65, which were reversed by transfecting Si-EFNA3. In Müller cells with ephrinA3 knockdown, the PI3K inhibitor LY294002 still decreased the protein levels of NF-κB p65 in the presence of EphA4-Fc, and the mRNA levels of GLAST were reduced by LY294002 and the NF-κB inhibitor SN50, respectively. Pre-injection of the PI3K/Akt pathway activator 740 Y-P reversed the GLAST downregulation in COH retinas. Western blot and TUNEL staining showed that transfecting of Si-EFNA3 reduced Müller cell gliosis and RGC apoptosis in COH retinas. Our results suggest that activated EphA4/ephrinA3 reverse signaling induces GLAST downregulation in Müller cells via inhibiting PI3K/Akt/NF-κB pathways, thus contributing to RGC damage in glaucoma.