Pub Date : 2025-12-03DOI: 10.1016/j.exer.2025.110790
William F. Horrigan , Quinn Caron , Aidan Rodriguez , Rupesh Singh , Bela Anand-Apte
The laser induced model of choroidal neovascularization (LiCNV) is a commonly used in vivo rodent model to study neovascular age-related macular degeneration (nAMD), although progression of this model is not well understood. In this study we characterize and compare the longitudinal progression of wound healing of laser induced choroidal neovascular (CNV) lesions in young and old mice. Using 2-month and 12-month-old C57BL/6J mice and ocular computerized tomography (OCT), fluorescein and indocyanine green angiography we performed a longitudinal imaging analysis at 3-, 7-, 14- and 28-days following laser injury. This was compared with immunohistochemical analysis of the lesions at similar timepoints for markers of angiogenesis, fibrosis, epithelial-to-mesenchymal transition (EMT), and gliosis. OCT analysis determined an increased lesion volume in older mice. In contrast younger mice demonstrated earlier vascularization and fibrosis with no difference in neovascularization or leakage. Reactive gliosis occurs directly above the laser-induced CNV lesion in both ages. The RPE in this model encloses the lesion area by day 14 in both young and old mice. This study concludes that age is an important variable in some but not all aspects of laser-induced CNV.
{"title":"The effect of age on wound healing in the laser induced choroidal neovascularization model","authors":"William F. Horrigan , Quinn Caron , Aidan Rodriguez , Rupesh Singh , Bela Anand-Apte","doi":"10.1016/j.exer.2025.110790","DOIUrl":"10.1016/j.exer.2025.110790","url":null,"abstract":"<div><div>The laser induced model of choroidal neovascularization (LiCNV) is a commonly used in vivo rodent model to study neovascular age-related macular degeneration (nAMD), although progression of this model is not well understood. In this study we characterize and compare the longitudinal progression of wound healing of laser induced choroidal neovascular (CNV) lesions in young and old mice. Using 2-month and 12-month-old C57BL/6J mice and ocular computerized tomography (OCT), fluorescein and indocyanine green angiography we performed a longitudinal imaging analysis at 3-, 7-, 14- and 28-days following laser injury. This was compared with immunohistochemical analysis of the lesions at similar timepoints for markers of angiogenesis, fibrosis, epithelial-to-mesenchymal transition (EMT), and gliosis. OCT analysis determined an increased lesion volume in older mice. In contrast younger mice demonstrated earlier vascularization and fibrosis with no difference in neovascularization or leakage. Reactive gliosis occurs directly above the laser-induced CNV lesion in both ages. The RPE in this model encloses the lesion area by day 14 in both young and old mice. This study concludes that age is an important variable in some but not all aspects of laser-induced CNV.</div></div>","PeriodicalId":12177,"journal":{"name":"Experimental eye research","volume":"263 ","pages":"Article 110790"},"PeriodicalIF":2.7,"publicationDate":"2025-12-03","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145687369","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-12-03DOI: 10.1016/j.exer.2025.110789
Kate E. Tarvestad-Laise , Grant Manion , Lisandra G. Plana Ramos , Mark Doll , Noel Verjan Garcia , Wendy M. Kittle , Levi Beverly , Nobuyuki Matoba , Hamid Band , Brian P. Ceresa
Tissue culture studies indicate c-Cbl and Cbl-b are negative regulators of growth factor receptor signaling. Inhibiting c-Cbl and Cbl-b is a potential therapeutic strategy for enhancing growth factor-mediated corneal re-epithelialization. An inducible, tissue-specific mouse model was used to assess the role of c-Cbl and Cbl-b in the corneal epithelium to determine the therapeutic potential of c-Cbl/Cbl-b inhibitors. Mice were generated by breeding c-Cbl and Cbl-b floxed mice with Keratin 14 (Krt14) ERTCre expressing mice. Knockdown was assessed by PCR and immunofluorescence. Corneal morphology was assessed using H&E staining. Ki67 immunostaining was used to measure cell proliferation. The functional consequence of c-Cbl/Cbl-b knockout was determined by measuring HGF-dependent corneal re-epithelialization following a mechanical debridement wound. Knockout of c-Cbl and Cbl-b was tamoxifen-dependent and did not affect the overall structure of the cornea. In male c-Cbl/Cbl-b knockout mice, corneal epithelial thickness and cell density were unchanged from controls, and there was enhanced HGF-dependent corneal re-epithelialization. Female knockout mice had reduced corneal epithelial thickness and cell density and increases in basal epithelial cell proliferation. However, there was no increase in HGF-dependent corneal re-epithelialization. Despite experimental considerations due to the Cre and tamoxifen toxicities, c-Cblf/f/Cbl-bf/f/Krt14 ERTCre+ mice are a viable model for studying c-Cbl and Cbl-b in corneal epithelial biology. Knockout of c-Cbl and Cbl-b affects corneal epithelial homeostasis and regeneration in both sexes of mice. However, the role of c-Cbl/Cbl-b in male knockout mice is stronger in re-epithelialization, whereas in female mice it regulates homeostasis.
{"title":"Knockout of c-Cbl and Cbl-b enhances corneal re-epithelialization in male, but not female, mice","authors":"Kate E. Tarvestad-Laise , Grant Manion , Lisandra G. Plana Ramos , Mark Doll , Noel Verjan Garcia , Wendy M. Kittle , Levi Beverly , Nobuyuki Matoba , Hamid Band , Brian P. Ceresa","doi":"10.1016/j.exer.2025.110789","DOIUrl":"10.1016/j.exer.2025.110789","url":null,"abstract":"<div><div>Tissue culture studies indicate c-Cbl and Cbl-b are negative regulators of growth factor receptor signaling. Inhibiting c-Cbl and Cbl-b is a potential therapeutic strategy for enhancing growth factor-mediated corneal re-epithelialization. An inducible, tissue-specific mouse model was used to assess the role of c-Cbl and Cbl-b in the corneal epithelium to determine the therapeutic potential of c-Cbl/Cbl-b inhibitors. Mice were generated by breeding c-Cbl and Cbl-b floxed mice with Keratin 14 (Krt14) ERTCre expressing mice. Knockdown was assessed by PCR and immunofluorescence. Corneal morphology was assessed using H&E staining. Ki67 immunostaining was used to measure cell proliferation. The functional consequence of c-Cbl/Cbl-b knockout was determined by measuring HGF-dependent corneal re-epithelialization following a mechanical debridement wound. Knockout of c-Cbl and Cbl-b was tamoxifen-dependent and did not affect the overall structure of the cornea. In male c-Cbl/Cbl-b knockout mice, corneal epithelial thickness and cell density were unchanged from controls, and there was enhanced HGF-dependent corneal re-epithelialization. Female knockout mice had reduced corneal epithelial thickness and cell density and increases in basal epithelial cell proliferation. However, there was no increase in HGF-dependent corneal re-epithelialization. Despite experimental considerations due to the Cre and tamoxifen toxicities, c-Cbl<sup>f/f</sup>/Cbl-b<sup>f/f</sup>/Krt14 ERTCre<sup>+</sup> mice are a viable model for studying c-Cbl and Cbl-b in corneal epithelial biology. Knockout of c-Cbl and Cbl-b affects corneal epithelial homeostasis and regeneration in both sexes of mice. However, the role of c-Cbl/Cbl-b in male knockout mice is stronger in re-epithelialization, whereas in female mice it regulates homeostasis.</div></div>","PeriodicalId":12177,"journal":{"name":"Experimental eye research","volume":"263 ","pages":"Article 110789"},"PeriodicalIF":2.7,"publicationDate":"2025-12-03","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145687389","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-12-02DOI: 10.1016/j.exer.2025.110778
Arturs Zemitis , Liva Caikovska , Juris Vanags , Jingzhi Fan , Kristaps Klavins , Guna Laganovska
Glaucoma is a leading cause of irreversible blindness worldwide and is primarily managed through IOP reduction, though its underlying pathophysiology remains incompletely understood. This study investigates metabolic alterations in glaucoma via targeted metabolomic profiling to identify potential biomarkers and pathogenic mechanisms. Aqueous humor samples were collected from 191 patients at Pauls Stradins Clinical University Hospital immediately before the initiation of cataract surgery, ensuring preoperative conditions. Quantitative metabolite profiling was conducted using liquid chromatography coupled to a Thermo Orbitrap Exploris 120 mass spectrometer. A significant association was observed between open-angle glaucoma and the presence of PEXS (χ2(1) = 9.96, p = 0.002, Cramer's V = 0.228). Several metabolites, including tryptophan, leucine, phenylalanine, and glutamine, were significantly upregulated in glaucoma patients (all p < 0.003, FDR = 0.0358), with tyrosine showing a similar trend (p = 0.0034, FDR = 0.0358). These findings suggest dysregulation of aromatic amino acid metabolism and potential impairment of amino acid hydroxylases, possibly linked to reduced tetrahydrobiopterin (BH4) availability. Disruption in BH4 regeneration—driven by oxidative stress or MTHFR polymorphisms—may impair nitric oxide synthesis and contribute to disease progression. Elevated glutamine and leucine levels could reflect compensatory neuroprotective mechanisms against excitotoxic damage. Our findings suggest that altered biopterin-related cofactor metabolism in aqueous humor may disrupt nitric oxide production and exacerbate oxidative stress, both of which are key factors in glaucoma pathogenesis. These insights highlight the potential of oxidative stress-related biomarkers and antioxidant-based strategies for future glaucoma diagnosis and therapy.
{"title":"Altered biopterin-related cofactor metabolism in the aqueous humor of glaucoma patients","authors":"Arturs Zemitis , Liva Caikovska , Juris Vanags , Jingzhi Fan , Kristaps Klavins , Guna Laganovska","doi":"10.1016/j.exer.2025.110778","DOIUrl":"10.1016/j.exer.2025.110778","url":null,"abstract":"<div><div>Glaucoma is a leading cause of irreversible blindness worldwide and is primarily managed through IOP reduction, though its underlying pathophysiology remains incompletely understood. This study investigates metabolic alterations in glaucoma via targeted metabolomic profiling to identify potential biomarkers and pathogenic mechanisms. Aqueous humor samples were collected from 191 patients at Pauls Stradins Clinical University Hospital immediately before the initiation of cataract surgery, ensuring preoperative conditions. Quantitative metabolite profiling was conducted using liquid chromatography coupled to a Thermo Orbitrap Exploris 120 mass spectrometer. A significant association was observed between open-angle glaucoma and the presence of PEXS (χ<sup>2</sup>(1) = 9.96, p = 0.002, Cramer's V = 0.228). Several metabolites, including tryptophan, leucine, phenylalanine, and glutamine, were significantly upregulated in glaucoma patients (all p < 0.003, FDR = 0.0358), with tyrosine showing a similar trend (p = 0.0034, FDR = 0.0358). These findings suggest dysregulation of aromatic amino acid metabolism and potential impairment of amino acid hydroxylases, possibly linked to reduced tetrahydrobiopterin (BH4) availability. Disruption in BH4 regeneration—driven by oxidative stress or MTHFR polymorphisms—may impair nitric oxide synthesis and contribute to disease progression. Elevated glutamine and leucine levels could reflect compensatory neuroprotective mechanisms against excitotoxic damage. Our findings suggest that altered biopterin-related cofactor metabolism in aqueous humor may disrupt nitric oxide production and exacerbate oxidative stress, both of which are key factors in glaucoma pathogenesis. These insights highlight the potential of oxidative stress-related biomarkers and antioxidant-based strategies for future glaucoma diagnosis and therapy.</div></div>","PeriodicalId":12177,"journal":{"name":"Experimental eye research","volume":"263 ","pages":"Article 110778"},"PeriodicalIF":2.7,"publicationDate":"2025-12-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145676683","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-12-01DOI: 10.1016/j.exer.2025.110776
Jieqing Yuan , Lingling Meng , Yuting Chen
Background
Graves' Ophthalmopathy (GO) is classified as an autoimmune condition that involves inflammation and structural changes within the orbit, frequently linked to thyroid abnormalities. The objective of this research is to clarify the genetic foundations associated with GO.
Methods
We employed summary statistics derived from a genome-wide association study (GWAS) focusing on Graves' Ophthalmopathy (GO), which included 753 affected individuals and 499,595 controls. To explore the genetic associations with the disease, we conducted an extensive transcriptome-wide association study (TWAS) that merged GWAS findings related to GO with expression quantitative trait loci (eQTL) data obtained from thyroid tissues featured in the Genotype-Tissue Expression Version 8 (GTEx V8) dataset. The validation of significant genes was carried out using three complementary methodologies: FUSION, FOCUS, and Multi-marker Analysis of GenoMic Annotation (MAGMA). To gain further insights into the genetic framework linked to GO, we utilized conditional and joint analysis in conjunction with fine-mapping approaches. Colocalization analysis was performed to identify potential overlapping single nucleotide polymorphisms (SNPs) between the significant genes and GO. In order to determine causality, we executed a Mendelian randomization analysis. The GeneMANIA tool was applied to elucidate gene interactions and their functional roles. Subsequently, we predicted transcription factor target genes, with a particular emphasis on potential ZSCAN9 targets. Finally, functional enrichment analysis revealed significant pathways associated with ZSCAN9 and its related targets.
Results
By employing a combination of three methodologies for TWAS—namely FUSION, FOCUS, and MAGMA—we have identified two candidate genes that demonstrate a significant correlation with GO. Mendelian randomization analysis yielded a suggestive but non-significant association between genetically predicted ZSCAN9 expression and GO, which warrants further investigation. Moreover, colocalization analysis indicates that the ZSCAN9 and HLA-DMA genes may share genetic signals with eQTL, suggesting that these genes could play a role in a shared pathway that contributes to the pathogenesis of GO.
Conclusion
Our findings highlight the genetic architecture of GO, identifying ZSCAN9 and HLA - DMA as potential causal genes, and underscore the importance of integrating multi-omics data to unravel complex disease mechanisms. Subsequent investigations ought to concentrate on corroborating these results and identifying potential therapeutic targets for GO.
{"title":"Transcriptome-wide association study identifies key genes in Graves' ophthalmopathy","authors":"Jieqing Yuan , Lingling Meng , Yuting Chen","doi":"10.1016/j.exer.2025.110776","DOIUrl":"10.1016/j.exer.2025.110776","url":null,"abstract":"<div><h3>Background</h3><div>Graves' Ophthalmopathy (GO) is classified as an autoimmune condition that involves inflammation and structural changes within the orbit, frequently linked to thyroid abnormalities. The objective of this research is to clarify the genetic foundations associated with GO.</div></div><div><h3>Methods</h3><div>We employed summary statistics derived from a genome-wide association study (GWAS) focusing on Graves' Ophthalmopathy (GO), which included 753 affected individuals and 499,595 controls. To explore the genetic associations with the disease, we conducted an extensive transcriptome-wide association study (TWAS) that merged GWAS findings related to GO with expression quantitative trait loci (eQTL) data obtained from thyroid tissues featured in the Genotype-Tissue Expression Version 8 (GTEx V8) dataset. The validation of significant genes was carried out using three complementary methodologies: FUSION, FOCUS, and Multi-marker Analysis of GenoMic Annotation (MAGMA). To gain further insights into the genetic framework linked to GO, we utilized conditional and joint analysis in conjunction with fine-mapping approaches. Colocalization analysis was performed to identify potential overlapping single nucleotide polymorphisms (SNPs) between the significant genes and GO. In order to determine causality, we executed a Mendelian randomization analysis. The GeneMANIA tool was applied to elucidate gene interactions and their functional roles. Subsequently, we predicted transcription factor target genes, with a particular emphasis on potential ZSCAN9 targets. Finally, functional enrichment analysis revealed significant pathways associated with ZSCAN9 and its related targets.</div></div><div><h3>Results</h3><div>By employing a combination of three methodologies for TWAS—namely FUSION, FOCUS, and MAGMA—we have identified two candidate genes that demonstrate a significant correlation with GO. Mendelian randomization analysis yielded a suggestive but non-significant association between genetically predicted ZSCAN9 expression and GO, which warrants further investigation. Moreover, colocalization analysis indicates that the ZSCAN9 and HLA-DMA genes may share genetic signals with eQTL, suggesting that these genes could play a role in a shared pathway that contributes to the pathogenesis of GO.</div></div><div><h3>Conclusion</h3><div>Our findings highlight the genetic architecture of GO, identifying ZSCAN9 and HLA - DMA as potential causal genes, and underscore the importance of integrating multi-omics data to unravel complex disease mechanisms. Subsequent investigations ought to concentrate on corroborating these results and identifying potential therapeutic targets for GO.</div></div>","PeriodicalId":12177,"journal":{"name":"Experimental eye research","volume":"263 ","pages":"Article 110776"},"PeriodicalIF":2.7,"publicationDate":"2025-12-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145667808","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-12-01DOI: 10.1016/j.exer.2025.110777
Yunjing Ma , Zijie Zhang , Zhe Jia , Qian Guo , Yue Huang , Hui Liu , Chen Zhang , Ruibo Yang , Xiaoxiao Lu , Shaozhen Zhao
This study aimed to investigate the therapeutic potential and anti-inflammatory mechanisms of tacrolimus (FK506) in moderate to severe dry eye by clinical, in vivo, and in vitro studies. We evaluated the effect of 0.1 % FK506 versus 0.1 % fluorometholone eye drops on patients with moderate to severe dry eye. Ocular surface disease index questionnaire (OSDI), tear breakup time (TBUT), corneal erosion, Schirmer's I test, and conjunctival impression cytology were measured at 2, 4, and 8 weeks. Tear samples from patients before and after FK506 treatment were subjected to proteomic analysis. The underlying mechanisms of FK506 were clarified using dry eye rat model and hypertonic-stimulated human corneal epithelial cells. Results demonstrated that FK506 showed transiently superior efficacy over fluorometholone in OSDI at week 2 (P = 0.041) and TBUT at week 4 (P = 0.001), despite both groups achieving significant improvements in all parameters from baseline to week 8 (P < 0.001). Tear proteomics revealed FK506-mediated modulation of autophagy-lysosomal pathway. FK506 improved ocular surface damage, reduced TNF-α and IL-1β, and upregulated LC3B II, TFEB, and LAMP1 protein expression, accompanied by reduced P62 in the rat cornea. In vitro, FK506 suppressed pro-inflammatory cytokines via enhanced autophagic flux, promoted TFEB nuclear translocation, restored lysosomal structural and functional integrity, while inhibited mTOR phosphorylation. Knockdown of TFEB reversed the FK506-mediated benefits. Overall, FK506 alleviates the symptoms and signs of moderate to severe dry eye by suppressing the inflammatory response in corneal epithelial cells. This therapeutic effect is mediated through the activation of the TFEB-driven autophagy-lysosomal pathway.
{"title":"Tacrolimus exerts anti-inflammatory effects in moderate to severe dry eye through activation of autophagy-lysosomal pathway","authors":"Yunjing Ma , Zijie Zhang , Zhe Jia , Qian Guo , Yue Huang , Hui Liu , Chen Zhang , Ruibo Yang , Xiaoxiao Lu , Shaozhen Zhao","doi":"10.1016/j.exer.2025.110777","DOIUrl":"10.1016/j.exer.2025.110777","url":null,"abstract":"<div><div>This study aimed to investigate the therapeutic potential and anti-inflammatory mechanisms of tacrolimus (FK506) in moderate to severe dry eye by clinical, in vivo, and in vitro studies. We evaluated the effect of 0.1 % FK506 versus 0.1 % fluorometholone eye drops on patients with moderate to severe dry eye. Ocular surface disease index questionnaire (OSDI), tear breakup time (TBUT), corneal erosion, Schirmer's I test, and conjunctival impression cytology were measured at 2, 4, and 8 weeks. Tear samples from patients before and after FK506 treatment were subjected to proteomic analysis. The underlying mechanisms of FK506 were clarified using dry eye rat model and hypertonic-stimulated human corneal epithelial cells. Results demonstrated that FK506 showed transiently superior efficacy over fluorometholone in OSDI at week 2 (P = 0.041) and TBUT at week 4 (P = 0.001), despite both groups achieving significant improvements in all parameters from baseline to week 8 (P < 0.001). Tear proteomics revealed FK506-mediated modulation of autophagy-lysosomal pathway. FK506 improved ocular surface damage, reduced TNF-α and IL-1β, and upregulated LC3B II, TFEB, and LAMP1 protein expression, accompanied by reduced P62 in the rat cornea. In vitro, FK506 suppressed pro-inflammatory cytokines via enhanced autophagic flux, promoted TFEB nuclear translocation, restored lysosomal structural and functional integrity, while inhibited mTOR phosphorylation. Knockdown of TFEB reversed the FK506-mediated benefits. Overall, FK506 alleviates the symptoms and signs of moderate to severe dry eye by suppressing the inflammatory response in corneal epithelial cells. This therapeutic effect is mediated through the activation of the TFEB-driven autophagy-lysosomal pathway.</div></div>","PeriodicalId":12177,"journal":{"name":"Experimental eye research","volume":"263 ","pages":"Article 110777"},"PeriodicalIF":2.7,"publicationDate":"2025-12-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145667796","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-11-27DOI: 10.1016/j.exer.2025.110774
Timon Ax
{"title":"Significant errors in the paper by Lee R, Ong J, Waisberg E, Suh A, Kadipasaoglu M, Mader T, Gibson CR, Berdahl J, Lee AG; SADES Study Group. Defining spaceflight associated dry eye syndrome (SADES): Mechanisms, complications, and countermeasures. Exp eye res. 2025 Sep;258:110513. doi: 10.1016/j.exer.2025.110513. Epub 2025 Jul 7. PMID: 40633645","authors":"Timon Ax","doi":"10.1016/j.exer.2025.110774","DOIUrl":"10.1016/j.exer.2025.110774","url":null,"abstract":"","PeriodicalId":12177,"journal":{"name":"Experimental eye research","volume":"263 ","pages":"Article 110774"},"PeriodicalIF":2.7,"publicationDate":"2025-11-27","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145631845","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-11-26DOI: 10.1016/j.exer.2025.110768
Vaibav Nandeesh , Xinyue Zhang , Katherine Zheng , Rahmat Asfiya , Paramanantham Anjugam , Nelson Dzidzorgbe Kwaku Akpabli-Tsigbe , Valeria Diaz , Thi Thao Nguyen , Brian Mooney , Akhil Srivastava , Jian-Xing Ma , Hu Huang
Purpose
Diabetic retinopathy (DR) is a medical complication of diabetes in which damage occurs to the retina. DR involves microvascular dysfunction of the retina, with impaired communication between pericytes and endothelial cells (EC) contributing to disease progression. This study investigated how small extracellular vesicles (sEV) released by human retinal pericytes (HRP) under diabetes-like stress conditions (high glucose (HG) + hypoxia) influence EC function.
Methods
HRP were cultured under HG + hypoxia and mannitol (control) conditions. sEV were isolated using differential ultracentrifugation and characterized using nanoparticle tracking analysis and transmission electron microscopy. Human retinal endothelial cells (HREC) were treated with HRP-derived sEV, and assessed for metabolic activity (MTT), barrier integrity (electric cell-impedance sensing), permeability (Transwell assay), migration (scratch assay), and angiogenic potential (tube formation). Protein expression was evaluated using Western blot and immunofluorescence staining. Proteomic profiles were performed using mass spectrometry and bioinformatics analyses.
Results
HRP sEV from diabetes-like stress conditions and control conditions showed no differences in size, concentration, or morphology. HREC uptake of HRP sEV was efficient and comparable across the two conditions. However, HRP sEV from diabetes-like stress conditions impaired HREC metabolic activity and barrier function while increasing permeability, migration, and angiogenesis. Conversely, HRP sEV from control conditions enhanced barrier integrity and metabolism without affecting permeability or angiogenesis. Proteomic analysis identified 86 sEV proteins that are differentially abundant between the two conditions. These proteins are enriched in pathways involved in extracellular matrix remodeling, inflammation, signaling, and metabolism.
Conclusions
HRP-derived sEV from HG + hypoxia conditions elicit endothelial dysfunctions relevant to DR pathology, in contrast to sEV from control conditions. The functional and proteomic alterations by diabetes-like stress suggest a mechanistic role for pericyte-derived sEV in DR progression and provide insights into potential therapeutic targets.
{"title":"Small extracellular vesicles derived from human retinal pericytes under high glucose and hypoxia conditions promote endothelial cell dysfunction in vitro","authors":"Vaibav Nandeesh , Xinyue Zhang , Katherine Zheng , Rahmat Asfiya , Paramanantham Anjugam , Nelson Dzidzorgbe Kwaku Akpabli-Tsigbe , Valeria Diaz , Thi Thao Nguyen , Brian Mooney , Akhil Srivastava , Jian-Xing Ma , Hu Huang","doi":"10.1016/j.exer.2025.110768","DOIUrl":"10.1016/j.exer.2025.110768","url":null,"abstract":"<div><h3>Purpose</h3><div>Diabetic retinopathy (DR) is a medical complication of diabetes in which damage occurs to the retina. DR involves microvascular dysfunction of the retina, with impaired communication between pericytes and endothelial cells (EC) contributing to disease progression. This study investigated how small extracellular vesicles (sEV) released by human retinal pericytes (HRP) under diabetes-like stress conditions (high glucose (HG) + hypoxia) influence EC function.</div></div><div><h3>Methods</h3><div>HRP were cultured under HG + hypoxia and mannitol (control) conditions. sEV were isolated using differential ultracentrifugation and characterized using nanoparticle tracking analysis and transmission electron microscopy. Human retinal endothelial cells (HREC) were treated with HRP-derived sEV, and assessed for metabolic activity (MTT), barrier integrity (electric cell-impedance sensing), permeability (Transwell assay), migration (scratch assay), and angiogenic potential (tube formation). Protein expression was evaluated using Western blot and immunofluorescence staining. Proteomic profiles were performed using mass spectrometry and bioinformatics analyses.</div></div><div><h3>Results</h3><div>HRP sEV from diabetes-like stress conditions and control conditions showed no differences in size, concentration, or morphology. HREC uptake of HRP sEV was efficient and comparable across the two conditions. However, HRP sEV from diabetes-like stress conditions impaired HREC metabolic activity and barrier function while increasing permeability, migration, and angiogenesis. Conversely, HRP sEV from control conditions enhanced barrier integrity and metabolism without affecting permeability or angiogenesis. Proteomic analysis identified 86 sEV proteins that are differentially abundant between the two conditions. These proteins are enriched in pathways involved in extracellular matrix remodeling, inflammation, signaling, and metabolism.</div></div><div><h3>Conclusions</h3><div>HRP-derived sEV from HG + hypoxia conditions elicit endothelial dysfunctions relevant to DR pathology, in contrast to sEV from control conditions. The functional and proteomic alterations by diabetes-like stress suggest a mechanistic role for pericyte-derived sEV in DR progression and provide insights into potential therapeutic targets.</div></div>","PeriodicalId":12177,"journal":{"name":"Experimental eye research","volume":"263 ","pages":"Article 110768"},"PeriodicalIF":2.7,"publicationDate":"2025-11-26","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145631854","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-11-26DOI: 10.1016/j.exer.2025.110775
Yurdagul Girgin , Gozde Sahin Vural , Yucel Yigit , Basak Isildar , Pakize Nur Akkaya , Gamze Tanriverdi , Muhammed Dara Tas , Ozlem Barut Selver
Aim
This study aimed to investigate the regenerative efficacy of topically administered insulin on corneal epithelial and stromal healing in a well-established rabbit model of ocular surface damage induced by chemical injury.
Methods
A standardized alkali burn model was created in the right eyes of 16 New Zealand white rabbits. Animals were randomly assigned into four groups: insulin-treated group (I group, n = 6), polyvinyl alcohol + povidone-treated group (P group, n = 6), untreated control group (C group, n = 4), and uninjured healthy controls (H group, n = 3).
Results
No statistically significant differences were observed among experimental groups regarding the epithelial defect area on day 1. By day 3, complete epithelial closure was achieved in all treated groups. There was no major intergroup differences were observed in surface integrity or clarity; in fact, insulin did not enhance epithelial healing or improve clinical scores compared to the control group However, stromal edema persisted more prominently in the insulin and control groups compared to healthy controls. Although α-SMA immunoreactivity did not show a significant intergroup difference, its relative increase in the insulin group may reflect early fibrotic remodeling. Electron microscopy demonstrated more compact and aligned collagen fibrils, as well as a greater density of telocyte-like interstitial cells in the insulin group—suggestive of enhanced stromal restructuring and repair.
Discussion
Topical insülin was associated with ultrastructural changes suggestive of stromal remodeling following chemical injury, although histological findings (including marked stromal edema) and the absence of clinical improvement indicate that its effect on overall corneal restoration remains uncertain. The presence of telocyte-like cells and improved collagen fibril organization in the insulin-treated corneas indicates a potential role for insulin in promoting physiological stromal remodeling. These findings support further investigation into insulin-based topical therapies for corneal wound healing.
{"title":"Regenerative effects of topical insulin on corneal wound healing: From surface restoration to stromal remodeling","authors":"Yurdagul Girgin , Gozde Sahin Vural , Yucel Yigit , Basak Isildar , Pakize Nur Akkaya , Gamze Tanriverdi , Muhammed Dara Tas , Ozlem Barut Selver","doi":"10.1016/j.exer.2025.110775","DOIUrl":"10.1016/j.exer.2025.110775","url":null,"abstract":"<div><h3>Aim</h3><div>This study aimed to investigate the regenerative efficacy of topically administered insulin on corneal epithelial and stromal healing in a well-established rabbit model of ocular surface damage induced by chemical injury.</div></div><div><h3>Methods</h3><div>A standardized alkali burn model was created in the right eyes of 16 New Zealand white rabbits. Animals were randomly assigned into four groups: insulin-treated group (I group, n = 6), polyvinyl alcohol + povidone-treated group (P group, n = 6), untreated control group (C group, n = 4), and uninjured healthy controls (H group, n = 3).</div></div><div><h3>Results</h3><div>No statistically significant differences were observed among experimental groups regarding the epithelial defect area on day 1. By day 3, complete epithelial closure was achieved in all treated groups. There was no major intergroup differences were observed in surface integrity or clarity; in fact, insulin did not enhance epithelial healing or improve clinical scores compared to the control group However, stromal edema persisted more prominently in the insulin and control groups compared to healthy controls. Although α-SMA immunoreactivity did not show a significant intergroup difference, its relative increase in the insulin group may reflect early fibrotic remodeling. Electron microscopy demonstrated more compact and aligned collagen fibrils, as well as a greater density of telocyte-like interstitial cells in the insulin group—suggestive of enhanced stromal restructuring and repair.</div></div><div><h3>Discussion</h3><div>Topical insülin was associated with ultrastructural changes suggestive of stromal remodeling following chemical injury, although histological findings (including marked stromal edema) and the absence of clinical improvement indicate that its effect on overall corneal restoration remains uncertain. The presence of telocyte-like cells and improved collagen fibril organization in the insulin-treated corneas indicates a potential role for insulin in promoting physiological stromal remodeling. These findings support further investigation into insulin-based topical therapies for corneal wound healing.</div></div>","PeriodicalId":12177,"journal":{"name":"Experimental eye research","volume":"263 ","pages":"Article 110775"},"PeriodicalIF":2.7,"publicationDate":"2025-11-26","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145622642","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-11-25DOI: 10.1016/j.exer.2025.110773
Zhangxing Xu, Lan Yang, Hong Gu
In diabetes, diabetic retinopathy (DR) is the most prevalent consequence. Despite continuous advancements in treatment, current therapeutic approaches remain limited in efficacy and may carry side effects. A deeper understanding of DR pathogenesis is essential for generating more effective strategies. Bioinformatics tools were employed to analyze single-cell data from the GEO and SRA databases related to DR, aiming to identify target genes. STRING database was used to analyze PPI network of COL4A2. COL4A2 expression in retinal endothelial cells under high glucose conditions was detected using qRT-PCR and Western blot. Cells with COL4A2 knockdown or overexpression were constructed, and effects of COL4A2 expression levels on cell proliferation, migration, and angiogenesis were assessed through CCK-8 assays, Annexin V/PI staining, scratch assays, and tube formation assays. COL4A2-overexpressing cells were treated with an AKT inhibitor, and regulatory role of AKT signaling on proliferative properties of retinal vascular endothelial cells was validated by measuring cell viability, apoptosis, migration, and angiogenic capacity. High glucose conditions facilitated production of COL4A2 in human retinal capillary endothelial cells (HRCECs). COL4A2 level was correlated with proliferative properties of HRCECs, and overexpression of COL4A2 enhanced proliferation, migration, and angiogenesis of retinal endothelial cells. COL4A2 augmented proliferative-related characteristics of HRCECs by activating AKT signaling pathway. This study revealed critical role of COL4A2 in DR and elucidated its mechanism of regulating angiogenesis and cell proliferation through AKT signaling pathway. Our research gave us fresh perspectives on molecular causes of DR and a theoretical basis for creating COL4A2-targeting treatment plans.
{"title":"COL4A2 activation of AKT signaling drives endothelial cell proliferation, migration, and angiogenesis in diabetic retinopathy","authors":"Zhangxing Xu, Lan Yang, Hong Gu","doi":"10.1016/j.exer.2025.110773","DOIUrl":"10.1016/j.exer.2025.110773","url":null,"abstract":"<div><div>In diabetes, diabetic retinopathy (DR) is the most prevalent consequence. Despite continuous advancements in treatment, current therapeutic approaches remain limited in efficacy and may carry side effects. A deeper understanding of DR pathogenesis is essential for generating more effective strategies. Bioinformatics tools were employed to analyze single-cell data from the GEO and SRA databases related to DR, aiming to identify target genes. STRING database was used to analyze PPI network of COL4A2. COL4A2 expression in retinal endothelial cells under high glucose conditions was detected using qRT-PCR and Western blot. Cells with COL4A2 knockdown or overexpression were constructed, and effects of COL4A2 expression levels on cell proliferation, migration, and angiogenesis were assessed through CCK-8 assays, Annexin V/PI staining, scratch assays, and tube formation assays. COL4A2-overexpressing cells were treated with an AKT inhibitor, and regulatory role of AKT signaling on proliferative properties of retinal vascular endothelial cells was validated by measuring cell viability, apoptosis, migration, and angiogenic capacity. High glucose conditions facilitated production of COL4A2 in human retinal capillary endothelial cells (HRCECs). COL4A2 level was correlated with proliferative properties of HRCECs, and overexpression of COL4A2 enhanced proliferation, migration, and angiogenesis of retinal endothelial cells. COL4A2 augmented proliferative-related characteristics of HRCECs by activating AKT signaling pathway. This study revealed critical role of COL4A2 in DR and elucidated its mechanism of regulating angiogenesis and cell proliferation through AKT signaling pathway. Our research gave us fresh perspectives on molecular causes of DR and a theoretical basis for creating COL4A2-targeting treatment plans.</div></div>","PeriodicalId":12177,"journal":{"name":"Experimental eye research","volume":"263 ","pages":"Article 110773"},"PeriodicalIF":2.7,"publicationDate":"2025-11-25","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145631862","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-11-25DOI: 10.1016/j.exer.2025.110770
Tang Shuijing , Wu Jiakang , Yang Tingting , Jing Ma
Background
The genetic architecture of keratoconus (KC) is complex, and the interplay between its biomechanical and inflammatory etiological components remains poorly defined. We aimed to quantitatively resolve these distinct genetic pathways to provide a mechanistic framework for disease pathogenesis.
Methods
We applied genomic structural equation modeling (genomic SEM) to large-scale GWAS summary statistics to model two latent genetic factors: a Corneal Biomechanical Stability Factor (CBSF) and an Atopic-Inflammatory Liability (AIL) factor. Pathway-specific multivariate GWAS were subsequently generated, from which candidate causal genes were inferred and then validated for differential expression in two independent patient transcriptomic cohorts. Downstream regulatory effects were interrogated using virtual knockout simulations on single-cell RNA-seq data, followed by pathway enrichment analysis. Functional validation was performed at the protein level using human corneal keratocytes cultured on a soft substrate designed to mimic the KC biomechanical environment.
Results
The genomic SEM supported a dual-pathway architecture, with the CBSF exerting a strong protective effect on KC (β = −0.48, p < 0.01) and the AIL factor conferring significant risk (β = 0.26, p < 0.05), together explaining 32.0 % of KC's genetic variance. From a pool of 112 candidate loci, we prioritized four high-confidence candidate genes. The protective effects of COL1A1, LOXL2, and ITGA1 were traced to their roles in matrix homeostasis, while the risk-conferring activity of SFRP1 was linked to inflammatory signaling. This mechanistic divergence, first revealed through in silico perturbation, was subsequently corroborated at the proteomic level, where our in vitro KC model displayed molecular signatures consistent with the predicted disease state.
Conclusion
By resolving the dual genetic pathways of keratoconus, we successfully identified high-confidence candidate genes whose pathogenic relevance was further supported by functional validation. This provides a robust mechanistic framework and identifies high-priority targets for precision therapy.
圆锥角膜（KC）的遗传结构是复杂的，其生物力学和炎症病原学成分之间的相互作用仍然不清楚。我们的目标是定量地解决这些不同的遗传途径，为疾病的发病机制提供一个机制框架。方法应用基因组结构方程模型（genomic SEM）进行大规模GWAS汇总统计，对角膜生物力学稳定因子（CBSF）和特应性炎症倾向因子（AIL）两个潜在遗传因素进行建模。随后产生了通路特异性多变量GWAS，从中推断出候选致病基因，然后在两个独立的患者转录组学队列中验证差异表达。通过对单细胞RNA-seq数据进行虚拟敲除模拟，研究下游调控效应，然后进行途径富集分析。在蛋白质水平上，使用在软底物上培养的人角膜角质细胞进行功能验证，以模拟KC生物力学环境。结果基因组SEM支持双通路结构，CBSF对KC具有较强的保护作用（β = - 0.48, p < 0.01），而AIL因子具有显著的风险（β = 0.26, p < 0.05），共同解释了32.0%的KC遗传变异。从112个候选基因座中，我们优先选择了4个高置信度的候选基因。COL1A1、LOXL2和ITGA1的保护作用可以追溯到它们在基质稳态中的作用，而SFRP1的风险赋予活性与炎症信号传导有关。这种机制上的差异，首先通过硅微扰揭示出来，随后在蛋白质组学水平上得到证实，我们的体外KC模型显示出与预测的疾病状态一致的分子特征。结论通过解析圆锥角膜的双重遗传通路，成功鉴定出高置信度的候选基因，其致病相关性得到功能验证的进一步支持。这提供了一个强大的机制框架，并确定了精确治疗的高优先级目标。
{"title":"Biomechanical and inflammatory pathways underlying the genetic architecture of keratoconus: A genomic SEM study","authors":"Tang Shuijing , Wu Jiakang , Yang Tingting , Jing Ma","doi":"10.1016/j.exer.2025.110770","DOIUrl":"10.1016/j.exer.2025.110770","url":null,"abstract":"<div><h3>Background</h3><div>The genetic architecture of keratoconus (KC) is complex, and the interplay between its biomechanical and inflammatory etiological components remains poorly defined. We aimed to quantitatively resolve these distinct genetic pathways to provide a mechanistic framework for disease pathogenesis.</div></div><div><h3>Methods</h3><div>We applied genomic structural equation modeling (genomic SEM) to large-scale GWAS summary statistics to model two latent genetic factors: a Corneal Biomechanical Stability Factor (CBSF) and an Atopic-Inflammatory Liability (AIL) factor. Pathway-specific multivariate GWAS were subsequently generated, from which candidate causal genes were inferred and then validated for differential expression in two independent patient transcriptomic cohorts. Downstream regulatory effects were interrogated using virtual knockout simulations on single-cell RNA-seq data, followed by pathway enrichment analysis. Functional validation was performed at the protein level using human corneal keratocytes cultured on a soft substrate designed to mimic the KC biomechanical environment.</div></div><div><h3>Results</h3><div>The genomic SEM supported a dual-pathway architecture, with the CBSF exerting a strong protective effect on KC (β = −0.48, p < 0.01) and the AIL factor conferring significant risk (β = 0.26, p < 0.05), together explaining 32.0 % of KC's genetic variance. From a pool of 112 candidate loci, we prioritized four high-confidence candidate genes. The protective effects of <em>COL1A1</em>, <em>LOXL2</em>, and <em>ITGA1</em> were traced to their roles in matrix homeostasis, while the risk-conferring activity of <em>SFRP1</em> was linked to inflammatory signaling. This mechanistic divergence, first revealed through <em>in silico</em> perturbation, was subsequently corroborated at the proteomic level, where our <em>in vitro</em> KC model displayed molecular signatures consistent with the predicted disease state.</div></div><div><h3>Conclusion</h3><div>By resolving the dual genetic pathways of keratoconus, we successfully identified high-confidence candidate genes whose pathogenic relevance was further supported by functional validation. This provides a robust mechanistic framework and identifies high-priority targets for precision therapy.</div></div>","PeriodicalId":12177,"journal":{"name":"Experimental eye research","volume":"263 ","pages":"Article 110770"},"PeriodicalIF":2.7,"publicationDate":"2025-11-25","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145622640","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
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