Molecular Vision最新文献

Purpose: To explore the protective effects of cassia polysaccharides on myopia by examining their influence on ARPE-19 cells with reduced PAX6 expression.

Methods: The ARPE-19 cell line with diminished PAX6 expression was established using a lentiviral approach and the addition of XAV-939, an inhibitor of Wnt/β-catenin. We assessed the expression of genes and proteins involved in Wnt/β-catenin, scleral remodeling, and cell cycle regulation following treatment with cassia polysaccharides. Gene and protein expression were quantified using reverse transcription PCR and western blot analyses, respectively. Additionally, the migratory capabilities of these cells were evaluated using a scratch assay.

Results: Optimal transduction was achieved with a multiplicity of infection of 20, successfully generating a stable ARPE-19 cell line with low PAX6 expression. Cassia polysaccharides did not significantly alter the expression of Wnt2 compared to control groups. Similarly, when treated with XAV-939, β-catenin levels were modified in PAX6-shRNA and XAV-939 but remained unchanged in the cassia polysaccharides. Scleral remodeling markers, including MMP-2 and TGF-β, were elevated, and COL1A1 was decreased in PAX6-shRNA, with no significant changes observed in the cassia polysaccharides. Cell cycle analysis indicated reduced cyclin-dependent kinase 1 and proliferating cell nuclear antigen levels in PAX6-shRNA, with cassia polysaccharides showing no significant effect. Scratch assay results demonstrated slower wound healing in PAX6-shRNA compared to controls over 72 h, with no significant differences observed in the cassia polysaccharides.

Conclusions: Cassia polysaccharides may mitigate ARPE-19 cell damage induced by low PAX6 expression through modulation of Wnt/β-catenin, potentially slowing the progression of myopia and offering a protective effect on vision.

Purpose: Diabetic retinopathy (DR), a severe microvascular complication of both type 1 and type 2 diabetes, is one of the leading causes of blindness. Prolonged hyperglycemia leads to vascular endothelial changes, inflammation, neovascularization, and apoptosis through multiple mechanisms, including increased aldose reductase (AR) activity and formation of advanced glycation end products, which contribute to the development of DR. Based on our previous studies with various functional foods that showed AR inhibition and prevented the formation of advanced glycation end products, in this study, a functional food (FF) mix was formulated and investigated its efficacy against DR progression in a rat model.

Methods: An FF mix was prepared with powders of amla pericarp, turmeric rhizome, ginger rhizome, cinnamon bark, and black pepper seeds in a specific proportion. Two-month-old Sprague-Dawley rats were grouped into control (C), streptozotocin-induced diabetes (D), and diabetes fed with FF at two levels (FF1, 0.85 g/100 g diet; FF2, 4.25 g/100 g diet) for 6 months from the induction of diabetes. At the end of the experiment, electroretinography was performed, and the eyes were dissected after the animals were sacrificed. A set of eyes was formalin-fixed for histology and immunohistochemistry examination, and the retina from the remaining eyes was used for immunoblotting analysis.

Results: Supplementation of FF mix in the diet to diabetic rats has improved retinal function (electroretinography), as well as prevented histomorphological changes and loss of photoreceptor cells (rhodopsin), compared to untreated diabetic rats. Further, FF mix ameliorated hyperglycemia-induced angiogenesis (vascular endothelial growth factor, hypoxia-inducible factor 1α) and gliosis (glial fibrillary acidic protein) in the diabetic rats, accompanied by decreased inflammation (phosphorylated nuclear factor κB, tumor necrosis factor α, monocyte chemoattractant protein 1) and apoptosis (Bax, Bcl2, caspase3, and caspase12).

Conclusions: This study illustrates the potential of an FF mix, attributed to the synergistic effects of its components, alleviating the progression of diabetic retinopathy by reducing diabetes-induced hypoxia, gliosis, and inflammation, while also inhibiting apoptosis in retinal cells.

Purpose: Diabetes is a chronic inflammatory disease that may damage the blood-retinal barrier, leading to diabetic retinopathy (DR). Blood-retinal barrier rupture may subject the retinal pigmented epithelial cells to a hyperosmolar stress (HOS), activating the transcription factor nuclear factor of activated T cells 5 (NFAT5). In addition, inflammatory cytokines, such as monocyte chemoattractant protein 1 (MCP-1/CCL2), play a crucial role in DR. The aims of our study were to determine whether HOS induces MCP-1 levels in arising retinal pigmented epithelial 19 (ARPE-19) cells and to decipher the responsible intracellular cascade involved in such stimulation.

Methods: ARPE-19 cells or ARPE-19 cells transfected with dominant negative NFAT5 plasmid or NFAT5 short hairpin RNA plasmids were preincubated or not for 1 h in the absence or presence of a protein kinase or transcription factor inhibitor and then incubated for 8 h with iso-osmolar or hyperosmolar medium in the absence or presence of inhibitor. NFAT5 reporter gene activity was quantified by luminescence. MCP-1 messenger RNA (mRNA) and protein levels were determined by quantitative real-time PCR and enzyme-linked immunosorbent assay, respectively. Biologically active MCP-1 was assessed by a calcium mobilization assay performed using Chinese hamster ovary cells expressing or not the MCP-1 receptor and apoaequorin.

Results: In response to HOS, ARPE-19 cells showed a significant increase in MCP-1 mRNA levels independent of NFAT5 activation. Moreover, the MCP-1 protein secreted by ARPE-19 in response to HOS is biologically active. The use of various inhibitors of protein kinase and transcription factors suggest that the HOS-induced increase in MCP-1 mRNA levels is dependent on a protein kinase C (PKC) and/or a MEK1/2-p38 pathway activating p53, as well as a PKC-p38-PI3K-PDK1-AKT activating hypoxia-inducible factor 1 alpha (HIF1α).

Conclusion: HOS increases the expression of MCP-1 mRNA and protein levels in ARPE-19 cells, and the secreted MCP-1 is biologically active. The HOS-induced increase of MCP-1 mRNA appears to be independent of NFAT5 activation. Despite the activation of NFAT5 upon HOS and the presence of NFAT5 binding sites in the MCP-1 gene promoter, activated NFAT5 may not be sufficient to induce MCP-1 gene transactivation in response to HOS in ARPE-19 cells. The intracellular cascade involved in the HOS-induced increase of MCP-1 mRNA in ARPE-19 cells may consist of a PKC-p38-PI3K-PDK1-AKT-HIF1α axis and/or a MEK1/2-p38-p53 axis.

Purpose: Diabetic retinopathy results from damage to small blood vessels and retinal neurons due to the overproduction of reactive oxygen species and overexpression of TRPM2 and TRPV1. Hence, inhibition of these events by ginger and selenium may reduce diabetes-induced ocular damage. Therefore, the aim of this study was to investigate the therapeutic effects of ginger, selenium, and their combinations on apoptosis, inflammation, insulin resistance, oxidative damage, and the expression of TRPM2 and TRPV1.

Methods: Seventy-two adult male Wistar rats were divided into nine groups as follows: control, diabetes, diabetes-ginger (100 mg/kg), diabetes-selenium (50, 100, and 150 µg/kg), and diabetes-ginger (100 mg/kg)-selenium (50, 100, and 150 µg/kg).

Results: Diabetes increased the expression of protein and genes of TRPM2 and TRPV1, and it induced oxidative damage by increasing malondialdehyde levels and decreasing superoxide dismutase, glutathione peroxidase, and catalase enzyme activities. Diabetes induced apoptosis by increasing BAX and caspase-3 gene expression and decreasing Bcl2 in eye tissue when compared to the control group. However, treatment with ginger (100 mg/kg), selenium (50, 100, and 150 µg/kg), and their combinations improved these situations in the diabetic groups compared to the diabetic group.

Conclusions: Diabetes induced retinopathy by inducing oxidative damage, inflammation, apoptosis, and upregulation of TRPM2 and TRPV1. However, treatments with selenium, ginger, and their combinations improved diabetic retinopathy by inhibiting oxidative damage, inflammation, and apoptosis and downregulating protein and gene expression of TRPM2 and TRPV1. The results of this study suggest that ginger and selenium can be a good treatment to inhibit the progression of diabetic retinopathy.

Purpose: Our goal was to explore whether inhibition of sortilin could protect the retina against ischemia/reperfusion (I/R) damage, as well as explore whether this inhibition could reduce inflammatory mediators in retinal Müller cells.

Methods: We used both primary human Müller cells and a rat Müller cell line (rMC-1) grown in normal (5 mM) or high (25 mM) glucose. Some cells were treated with AF38469, a small-molecule inhibitor of sortilin. We performed western blotting for the inflammatory mediators, tumor necrosis factor α, and NOD-like receptor protein 3. We also measured protein levels of lysosome-associated membrane glycoprotein 2 (LAMP2), a marker of autophagy, and cleaved caspase 3, a marker of apoptosis, in the cells. We then tested the actions of eye drops containing AF38469 on mice exposed to I/R. We assessed neuronal damage at 2 days post-I/R and vascular damage at 10 days post-I/R.

Results: High-glucose culturing conditions significantly increased inflammatory, autophagic, and apoptotic markers in both primary human Müller and rat Müller cells. All markers were reduced by treating the cells with AF38469. AF38469 eye drops also significantly reduced I/R-induced neuronal and vascular damage.

Conclusion: These studies demonstrate that sortilin regulates the inflammatory, autophagic, and apoptotic pathways in Müller cells grown in high glucose. Inhibition of sortilin using AF38469 eye drops also reduced I/R-induced retinal damage.

Purpose: To determine whether nitazoxanide (NZT) can increase mitochondrial nuclear retrograde regulator 1 (MNRR1) in retinal endothelial cells (RECs) grown in normal or high glucose and thereby reduce inflammation.

Methods: We used control and diabetic human retinal protein samples, control and diabetic mouse retinal samples, and RECs grown in normal (5 mM) and high (25 mM) glucose protein samples to explore levels of MNRR1, high mobility group box 1, interleukin 1β, tumor necrosis factor α, and Tom20 by western blotting. We used immunostaining to localize MNRR1 in the retina. We also used a Seahorse XF^e24 Bioanalyzer to measure the oxygen consumption rate in RECs under different conditions. Some cells were treated with NZT to increase MNRR1 levels.

Results: MNRR1 protein levels were reduced in the diabetic retina of both humans and mice. MNRR1 was localized to RECs. RECs grown in normal or high glucose and treated with NZT had significantly higher MNRR1 levels. NZT reduced inflammatory markers in RECs grown in high glucose. NZT increased the oxygen consumption rate in RECs grown in high glucose, which was associated with an increase in Tom20.

Conclusions: MNRR1 is reduced in the diabetic retina. NZT increased MNRR1 levels in RECs. NZT protected RECs grown in high glucose by reducing inflammatory mediators and mitochondrial dysfunction and increasing mitochondrial mass. NZT may offer a new therapeutic option for diabetic retinopathy.

The fovea is an anatomic specialization of the human retina critical for high visual acuity, color vision, and contrast sensitivity. The molecular and cellular pathways directing this focal topography are still to be determined. Abnormalities of the fovea (e.g., foveal hypoplasia in children) are considered a significant contributor to reduced quality of life. In addition, the fovea is often damaged in common retinal diseases, such as age-related macular degeneration and diabetic retinopathy, with a global economic burden of $500 billion USD. Currently, there are no treatments for foveal defects. Most genes contributing to foveal abnormalities have been identified but are yet to be characterized and studied. This is because common laboratory animals do not have a fovea, and only rare human tissue samples are available during the major phase of foveal maturation, from birth to the end of the fourth year of life. We discuss validation of the anole lizard, which has a foveal structure, for research studies into foveal development. Since foveal development continues after birth, it may be possible to stimulate new foveal maturation where there is developmental damage. From this review, we propose an evidence-based cellular mechanism that offers new possibilities for testing future therapies for foveal defects.

Purpose: To evaluate the association between suboptimal glycemic control and central macular thickness in central retinal vein occlusion patients over a 1-year follow-up period.

Methods: This retrospective cohort study included adult patients with central retinal vein occlusion diagnosed at the Helsinki University Hospital, Finland, with HbA1c levels measured within 6 months before or at diagnosis. Patients were divided into two groups: those with good (≤42 mmol/mol) and poor glycemic control (>42 mmol/mol). Central macular thickness and best-corrected visual acuity were assessed at baseline, 3 months, and 1 year.

Results: Among 40 patients, 10 had suboptimal glycemic control. At 3 months and 1 year, central macular thickness was significantly higher in the poor glycemic control group compared with the good glycemic control group (367.0 ± 43.3 µm vs. 288.2 ± 36.6 µm, p = 0.016 and 380.8 ± 44.8 µm vs. 302.0 ± 71.3 µm, p = 0.035, respectively). HbA1c levels correlated with central macular thickness at 3 months (R ² = 0.514, p = 0.045) but did not reach statistical significance at 1 year (R ² = 0.246, p = 0.071).

Conclusions: Poor glycemic control in patients with central retinal vein occlusion is associated with greater central macular thickness at both 3 months and 1 year. These findings emphasize the importance of optimal glycemic control to improve retinal outcomes in central retinal vein occlusion.

Purpose: To evaluate the genetic characteristics using whole-exome sequencing (WES), aiming to assess the potential of this approach for accurate diagnoses and to explore the genetic factors underlying these conditions.

Methods: A total of 28 patients, including 6 with primary congenital glaucoma and 22 with juvenile open-angle glaucoma, were studied. Genetic analysis involved initial Sanger sequencing for the CYP1B1 and MYOC genes. WES was subsequently performed in 11 patients with negative initial results, using a panel of genes most associated with glaucoma and related ophthalmic syndromes. Variant interpretation was performed based on American College of Medical Genetics and Genomics guidelines. Segregation analysis was performed when possible.

Results: Pathogenic variants in CYP1B1 and MYOC genes were identified in three patients (10%). WES identified disease-causing variants in three additional patients (27% of those with negative results from the initial testing conducted so far), all of them with syndromic features. In families where segregation analysis was possible, variants were confirmed to segregate with the clinical presentation. The diagnostic yield was 21%.

Conclusions: WES is an effective diagnostic tool for early-onset glaucoma, enhancing clinical management and genetic counseling. It supports its use in routine diagnostics to enable early detection and intervention for at-risk relatives. Further research is needed to uncover additional genetic factors and refine testing guidelines.