SB431542 partially inhibits high glucose-induced EMT by restoring mitochondrial homeostasis in RPE cells.

BACKGROUND: The epithelial-mesenchymal transition (EMT) of retinal pigment epithelial (RPE) cells participated in the development of retinal fibrosis. SB431542 is a small molecule inhibitor with inhibitory effects on the ALK4, ALK5 and ALK7. Our study aimed to explore the effect of SB431542 on the EMT of RPE cells and to provide new ideas for the treatment of retinal fibrosis. METHODS: We performed fundus fluorescein angiography, optical coherence tomography and hematoxylin-eosin staining in vivo to observe the effect of SB431542 on choroidal neovascularization (CNV)-induced retinopathy. The proliferation, migration, cytoskeleton, adhesion, reactive oxygen species (ROS), mitochondrial morphology and membrane potential of RPE cells were observed in vitro through fluorescein diacetate staining, Cell Counting Kit-8 experiment, wound healing assay, phalloidin staining, immunofluorescence, MitoSOX, DCFH-DA, MitoTracker and JC-10 staining. Western blot, reverse transcription quantitative and immunofluorescence were used to detect the expression of EMT-related markers, pERK1/2, pGSK3ß and ß-catenin. RESULTS: SB431542 significantly alleviated retinopathy in the CNV model. The proliferation, migration and adhesion in RPE cells decreased to a certain extent in SB431542 treatment. SB431542 partially normalized the structure of RPE cells. The expression levels of E-cadherin increased, while the expression levels of laminin and N-cadherin decreased with SB431542 treatment. SB431542 reduced the production of total ROS, mitochondrial SOX and recovered the mitochondrial membrane potential to a certain degree. In addition, our study showed that SB431542 downregulated the phosphorylation of ERK1/2, GSK3ß and the expression of ß-catenin. CONCLUSION: SB431542 improved EMT in RPE cells by maintaining mitochondrial homeostasis via the ERK1/2 and GSK3ß/ß-catenin pathways. Video Abstract SB431542 inhibits EMT in RPE cells under high glucose conditions.

Endocytosis of GluN2B-containing NMDA receptors mediates NMDA-induced excitotoxicity.

Abstract: N-methyl-D-aspartate (NMDA) receptor overactivation is involved in neuronal damage after stroke. However, the mechanism underlying NMDA receptor-mediated excitotoxicity remains unclear. In this study, we confirmed that excessive activation of NMDARs led to cell apoptosis in PC12 cells and in primary cultured cortical neurons, which was mediated predominantly by the GluN2B-containing, but not the GluN2A-containing NMDARs. In addition, Clathrin-dependent endocytosis participated in NMDA-induced excitotoxicity. Furthermore, we identified that GluN2B-containing NMDARs underwent endocytosis during excessive NMDA treatment. Peptides specifically disrupting the interaction between GluN2B and AP-2 complex not only blocked endocytosis of GluN2B induced by NMDA treatment but also abolished NMDA-induced excitotoxicity. These results demonstrate that Clathrin-dependent endocytosis of GluN2B-containing NMDARs is critical to NMDA-induced excitotoxicity in PC12 cells and in primary cultured cortical neurons, and therefore provide a novel target for blocking NMDAR-mediated excitotoxicity.

Construction and validation of a neovascular glaucoma nomogram in patients with diabetic retinopathy after pars plana vitrectomy.

BACKGROUND: Neovascular glaucoma (NVG) is likely to occur after pars plana vitrectomy (PPV) for diabetic retinopathy (DR) in some patients, thus reducing the expected benefit. Understanding the risk factors for NVG occurrence and building effective risk prediction models are currently required for clinical research. AIM: To develop a visual risk profile model to explore factors influencing DR after surgery. METHODS: We retrospectively selected 151 patients with DR undergoing PPV. The patients were divided into the NVG (NVG occurrence) and No-NVG (No NVG occurrence) groups according to the occurrence of NVG within 6 months after surgery. Independent risk factors for postoperative NVG were screened by logistic regression. A nomogram prediction model was established using R software, and the model's prediction accuracy was verified internally and externally, involving the receiver operator characteristic curve and correction curve. RESULTS: After importing the data into a logistic regression model, we concluded that a posterior capsular defect, preoperative vascular endothelial growth factor ≥ 302.90 pg/mL, glycosylated hemoglobin ≥ 9.05%, aqueous fluid interleukin 6 (IL-6) ≥ 53.27 pg/mL, and aqueous fluid IL-10 ≥ 9.11 pg/mL were independent risk factors for postoperative NVG in patients with DR (P < 0.05). A nomogram model was established based on the aforementioned independent risk factors, and a computer simulation repeated sampling method was used to internally and externally verify the nomogram model. The area under the curve (AUC), sensitivity, and specificity of the model were 0.962 [95% confidence interval (95%CI): 0.932-0.991], 91.5%, and 82.3%, respectively. The AUC, sensitivity, and specificity of the external validation were 0.878 (95%CI: 0.746-0.982), 66.7%, and 95.7%, respectively. CONCLUSION: A nomogram constructed based on the risk factors for postoperative NVG in patients with DR has a high prediction accuracy. This study can help formulate relevant preventive and treatment measures.

Microinvasive pars plana vitrectomy combined with internal limiting membrane peeling versus anti-VEGF intravitreal injection for treatment-naïve diabetic macular edema (VVV-DME study): study protocol for a randomized controlled trial.

BACKGROUND: Diabetic macular edema (DME) is the main cause of vision loss in diabetic patients. Currently, anti-vascular endothelial growth factor (VEGF) intravitreal injection stands as the first-line therapy for DME. However, some patients exhibit insufficient response to anti-VEGF agents and often require multiple injections, imposing psychological and economic burdens. While microinvasive pars plana vitrectomy (PPV) has been shown to be safe and effective in treating refractory DME, scant research has explored its application to treatment-naïve DME. The purpose of this study is to determine whether early PPV combined with internal limiting membrane (ILM) peeling can lessen the therapeutic burden of DME patients, prevent vision loss, and maintain long-term stabilization of diabetic retinopathy. METHODS: This is a single-center, prospective, parallel-group, non-inferiority randomized controlled trial involving 102 DME participants. Participants will be randomly assigned to either the study group (PPV combined with ILM peeling) or the control group (conbercept intravitreal injection (IVC)) at a 1:1 ratio, with a scheduled follow-up at 12 months post-operation. Comparative analysis of results between the two groups will be conducted at months 1, 3, 6, and 12 after the intervention. The primary outcomes involve evaluating the changes in central subfield thickness (CST) and best corrected visual acuity (BCVA). The secondary outcomes include assessment of optical coherence tomography (OCT) and OCT angiography (OCTA) biomarkers, re-treatment and adverse events rates, diabetic retinopathy (DR) development, cost-effectiveness analysis, and vision-related quality of life (VRQL). DISCUSSION: Some patients do not respond well to anti-VEGF drugs and repeated intravitreal injections increase the treatment burden for patients. The VVV study aims to explore whether PPV combined with ILM peeling could become an initial treatment option for treatment-naïve DME patients. TRIAL REGISTRATION: ClinicalTrials.gov NCT05728476. Registered on 15 February 2023.

Postsynaptic RIM1 modulates synaptic function by facilitating membrane delivery of recycling NMDARs in hippocampal neurons.

NMDA receptors (NMDARs) are crucial for excitatory synaptic transmission and synaptic plasticity. The number and subunit composition of synaptic NMDARs are tightly controlled by neuronal activity and sensory experience, but the molecular mechanism mediating NMDAR trafficking remains poorly understood. Here, we report that RIM1, with a well-established role in presynaptic vesicle release, also localizes postsynaptically in the mouse hippocampus. Postsynaptic RIM1 in hippocampal CA1 region is required for basal NMDAR-, but not AMPA receptor (AMPAR)-, mediated synaptic responses, and contributes to synaptic plasticity and hippocampus-dependent memory. Moreover, RIM1 levels in hippocampal neurons influence both the constitutive and regulated NMDAR trafficking, without affecting constitutive AMPAR trafficking. We further demonstrate that RIM1 binds to Rab11 via its N terminus, and knockdown of RIM1 impairs membrane insertion of Rab11-positive recycling endosomes containing NMDARs. Together, these results identify a RIM1-dependent mechanism critical for modulating synaptic function by facilitating membrane delivery of recycling NMDARs.