Alterations in retinal tissue permeability are a hallmark of several ocular pathologies, including diabetic retinopathy and age-related macular degeneration. Although permeability plays a crucial role in maintaining retinal homeostasis, previously reported values vary widely due to differences in animal models, tissue preparation, and experimental configurations. In the present study, the hydraulic permeability of porcine retinal and subretinal layers was quantitatively characterized using a custom Ussing chamber, a two-component system for measuring trans-tissue fluid flow, enabling controlled physiological pressures. Tissue isolation was verified by scanning electron microscopy, which allowed identification of the targeted ocular layers based on their characteristic surface morphology. Neuroretina, BrM-choroid, and RPE-BrM-choroid samples were analyzed, and the effect of tissue fixation was systematically evaluated. Additional experiments were performed on fixed retina to investigate the pressure-permeability relationship. To complement the experimental measurements, a COMSOL Multiphysics model of fluid transport across the ocular layers was developed. The model reproduced the experimental configuration and incorporated experimentally determined transport properties together with tissue mechanical properties to (i) evaluate possible setup-induced artefacts, (ii) verify the analytical estimation of RPE permeability, and (iii) investigate the influence of strain-dependent retinal permeability on fluid transport across the tissue complex. Fixation with 4% formalin did not significantly alter retinal permeability while improving measurement reproducibility. The neuroretina exhibited relatively high intrinsic permeability (∼10-14 m2), consistent with its elevated metabolic demand, whereas the RPE-BrM-Choroid complex displayed markedly lower permeability, (∼10-17 m2), reflecting the barrier function of the outer blood-retina barrier. Numerical simulations indicated that the overall hydraulic response is dominated by the RPE-BrM-Choroid complex, while retinal strain-dependent permeability had a negligible influence on the predicted fluid transport, despite the dependence of the permeability-strain relationship on the adopted mechanical assumptions. Overall, this study provides an integrated experimental-computational framework for quantifying retinal and ocular tissue permeability. The proposed approach reduces variability associated with previous methodologies and supports the use of the porcine retina as a biologically relevant model for human tissue. These results provide physiologically relevant parameters for investigating fluid regulation, blood-retina barrier dysfunction, and ocular drug transport.

Dry eye disease (DED) is a heterogeneous, multifactorial disorder characterized by loss of tear-film and/or ocular-surface homeostasis, with epithelial stress, inflammation, and neurosensory abnormalities contributing to varying degrees across clinical phenotypes. This review synthesizes current evidence within a circuit-based sensory-autonomic-immune framework linking corneal sensory dysfunction to lacrimal dysfunction. We emphasize two anatomically and functionally distinct adrenergic checkpoints: norepinephrine (NE)-Adra1a-Ucp2 signaling in lacrimal acinar and myoepithelial cells, which restrains tear secretion, and epithelial NE-ADRB2 signaling, which reduces NGF/GDNF support and limits sensory nerve regeneration. We further examine Th1 CD4[+] T-cell/IFN-γ-mediated neural injury, TRPV1/TRPA1 nociceptor signaling, substance P/NK1R-mediated immune activation, and denervation-associated ferroptotic injury with respect to causal direction and strength of evidence. The relevance of these mechanisms varies across DED phenotypes: lacrimal-autonomic pathways are most pertinent to aqueous-deficient or mixed disease with tear hyposecretion, whereas MGD-predominant evaporative DED is driven primarily by meibomian-gland and lipid-layer abnormalities. Human evidence remains largely associative, and ferroptosis should currently be regarded as an exploratory downstream injury mechanism rather than an established therapeutic target. Intranasal varenicline provides clinical proof of concept for reflex-pathway modulation, whereas adrenergic, neuropeptide/TRP, neurotrophic, and ferroptosis-directed strategies require phenotype-specific mechanistic and clinical validation. This evidence-weighted framework links source cells and receptors to target compartments, candidate biomarkers, and expected functional outcomes.

The mouse retina has become a central model for linking neural organization to degeneration and repair because it combines laminated anatomy, genetic access, longitudinal imaging, and measurable visual function. This review is organized around three methodological eras: anatomical foundations (1970s-1990s), genetic access and reproducible injury models (2000s-2010s), and cell-resolved omics and high-resolution in vivo imaging (2015-present). Four Method Boxes translate this history into practical guidance for histology, electrophysiology, genetic manipulation, and in vivo imaging. We then examine two therapeutic areas that expose the model's evidentiary limits: retinal ganglion cell (RGC) axon regeneration after optic nerve injury and stage-matched treatment of photoreceptor degeneration. Across both areas, survival, local function, long-distance regeneration, circuit reconnection, and behavior are treated as distinct outcomes. The mouse retina is therefore most informative when complementary readouts are combined and promising mechanisms are tested against human-relevant tissue and larger-eye systems before clinical inference.

Our goal was to restore age related loss in HA levels in the vitreous of the presbyopic monkey eye to determine if intravitreal injection of HA restores the anatomical (i.e. accommodative lens thickening, forward movement of the posterior zonule insertion zone and thereby ciliary muscle forward movement etc.) and dioptric accommodative response and if so the duration of the response increase. We injected 0.3ml of HA (14mg/ml; Healon GV) into the vitreous near the optic nerve region, quantified the accommodative response to i.v. pilocarpine (2mg/kg wt) before and two weeks after HA injection, and compared accommodative responses at both time points and at 5, 8 and 16 months post HA. Accommodative amplitude was measured by Hartinger coincidence refractometry. Accommodative movements of the lens, and ciliary muscle were measured by ultrasound biomicroscopy (UBM; 50, 20 MHz) and choroidal movements in the region of the optic nerve were measured by OCT. RESULTS: Two weeks following HA injection the accommodative response to pilocarpine increased dramatically compared to baseline (ipsilateral eye; pre-HA); lens thickening ([increased] by 0.24 ± 0.05mm;p<0.022), forward ciliary muscle movement (by 0.42 + 0.09mm; p<0.019), centrifugal choroidal movement around the optic nerve (by 0.47 + 0.19mm;p<0.09) and accommodative amplitude (by 1.19 ± 0.79 diopters, and the anterior lens surface became more sharply curved (by 2.43 ± 0.49 mm radius of curvature; p<0.016). The forward movement of the posterior zonule insertion zone of the vitreous zonule reflects both forward movement of the vitreous zonule and forward movement of the muscle. Between experimental sessions in the contralateral control eye (no HA) there was no change in anatomical accommodation response as measured by insertion zone forward movement (-0.01±0.13mm) nor for any other parameter.(Croft et al., 2024) The HA anatomical accommodation effect was maintained up to 16 months following injection. Further, in response to Edinger-Westphal (EW) nucleus electrical stimulation to induce accommodation - the increased tissue movement response was also still present 5 to 8 months post HA injection (i.e., forward muscle movement by 0.28 ± 0.03mm and lens thickening by 0.13 ± 0.03mm; n=2) in absence of pilocarpine administration. The data suggests that HA levels are important to accommodative movements of the lens, muscle and choroid. These experiments may provide insights into presbyopia therapy, the mechanisms of accommodation, presbyopia, accommodating IOL function and perhaps glaucomatous optic neuropathy.

Age-related cataract (ARC) develops through cumulative oxidative stress and progressive loss of lens epithelial homeostasis, but the small-RNA mechanisms that distinguish an adaptive response from sustained injury remain unclear. Using a time-resolved H2O2 model in HLEC-B3 cells, we identified tRF-31, a 31-nt 5' tRNA-Ala-derived small RNA, as a biphasic stress-responsive regulator that increased during an early oxidative-response state with preserved viability but declined during sustained sublethal injury. N-acetylcysteine blunted the early increase and counteracted the later decrease in tRF-31, whereas the full-length parental tRNA-Ala pool remained stable and quantitative immunofluorescence showed stress-dependent angiogenin redistribution that was temporally associated with tRF-31 expression. Functionally, tRF-31 replenishment reduced apoptosis and mitochondrial depolarization while supporting post-stress epithelial recovery and DNA synthesis, whereas tRF-31 inhibition aggravated apoptotic and mitochondrial injury after endogenous tRF-31 had declined. Transcriptomic profiling, target prediction, quantitative PCR validation, and wild-type and mutant 3' untranslated region reporter assays supported homeodomain-interacting protein kinase 2 (HIPK2) as a direct target of tRF-31. tRF-31 suppressed HIPK2 and p53 Ser46 phosphorylation, reduced BAX, BAD, p21, and cleaved caspase-3, and preserved Bcl-2, RB phosphorylation, CCNE2, and PCNA. HIPK2 overexpression counteracted the anti-apoptotic and pro-recovery effects of tRF-31 mimic. In 92 ARC anterior capsule specimens, lower tRF-31, higher HIPK2 and CDKN1A expression, and a higher BAX/Bcl-2 ratio were associated with greater nuclear hardness and poorer preoperative visual acuity after adjustment for age. These findings identify a temporally regulated tRF-31/HIPK2-p53 axis that links oxidative stress responses to lens epithelial injury and impaired recovery in ARC.

Autophagy represents a lysosomal degradative pathway through which cells systematically break down and recycle their own constituents, including proteins, lipid droplets, and entire organelles, by sequestering them within double-membrane autophagosomes prior to lysosomal fusion. Beyond its role as a fundamental quality control mechanism, this process critically shapes cellular metabolism and preserves intracellular equilibrium. In ocular biology, autophagic activity is involved in structural morphogenesis during eye development and becomes dysregulated across a spectrum of vision-threatening disorders. This review synthesises current evidence on autophagy throughout distinct ocular compartments, with particular attention given to its pathogenic contributions to corneal disease, cataract formation, glaucomatous neurodegeneration, and major retinopathies, with the goal of advancing the understanding of the underlying mechanism in this evolving field.

Epithelial-to-mesenchymal transition (EMT) of lens epithelial cells (LECs) drives fibrotic cataractogenesis, including anterior subcapsular cataract (ASC). Although non-canonical p38 MAPK signaling is markedly elevated during TGF-β-mediated EMT, defining its precise role has historically been complicated by off-target effects of first-generation inhibitors (such as SB203580) on ERK1/2 and Smad3 pathways. To clarify the functional necessity of p38 activity in lens EMT, we evaluated the effects of next-generation, highly selective p38 inhibitors with distinct modes of action (Skepinone-L, VX-745 and BIRB796). Pharmacological blockade using Skepinone-L, VX-745, or BIRB796 reproducibly preserved the lens epithelial phenotype and prevented TGF-β-mediated transdifferentiation into myofibroblasts. Despite their distinct structural mechanisms and binding modes, all three compounds yielded identical protective phenotypes. This functional concordance across mechanistically diverse inhibitors confirms that the observed anti-fibrotic effects are strictly attributable to p38 inhibition rather than off-target activity. Of great interest, selective p38 inhibition uncouples key events in the EMT process. While p38 blockade prevents downstream phenotypic transition and extracellular matrix remodeling, it does not block the upstream induction of α-smooth muscle actin (α-SMA). These findings demonstrate that p38 MAPK acts as an essential co-regulatory hub for cytoskeletal reorganization, and extracellular matrix remodeling, rather than an absolute switch for all transdifferentiation markers. Consequently, targeting p38 offers a refined strategy for preventing fibrotic cataract formation by specifically uncoupling the pathological manifestations of lens EMT leading to fibrotic cataract formation.

Dry eye disease (DED) is characterized by tear film instability and chronic inflammation, with the cornea serving as a key target site vulnerable to inflammatory damage. This study integrated single-cell and bulk transcriptomics combined with Scissor and CIBERSORTx algorithms to screen corneal key cell populations and biomarkers in murine DED. Corneal wing cells were identified as core disease-associated cells, among which interferon-response wing cells (IFN-response-wing) were the pivotal pathogenic subtype. Three IFN-response-wing signature biomarkers Upk3b, Dusp1 and Ly6e were screened, enriched in immune pathways including antigen processing and presentation, primary immunodeficiency, and cytokine receptor interaction. We constructed a chemical-induced DED mouse model for multi-layered validation via immunofluorescence, Real-time polymerase chain reaction (RT-qPCR) and Western blot. Immunofluorescence revealed tissue-specific protein patterns: DED corneal epithelium exhibited elevated IRF1, Dusp1 and Ly6e protein yet reduced Upk3b, with Ly6e co-localized with IRF1-positive cells. However, RT-qPCR and Western blot of whole corneal tissue demonstrated significant upregulation of Upk3b, Dusp1 and Ly6e at both mRNA and total protein levels. Collectively, IFN-response-wing drives DED pathogenesis, and Upk3b, Dusp1, Ly6e act as tissue-specific DED biomarkers with inconsistent trends between epithelial localized protein and global corneal molecular abundance, providing novel mechanistic clues for DED diagnosis and targeted therapy.

Eye field specification requires precise spatiotemporal signaling. Among these signals, the morphogen Noggin blocks both BMP and Activin signaling pathways to generate retinal tissue from pluripotent frog animal cap cells. We previously found that Pax6 and Tbx3 are sufficient to replace Noggin in generating retinal tissue, but it was unclear which signaling pathway affected tbx3 expression. We had also found that Noggin, while activating all other eye field transcription factors (EFTFs), repressed tbx3 expression in animal cap cells, and that Otx2, a transcription factor essential for anterior neural plate induction, is expressed prior to tbx3 and can overcome Noggin-mediated repression to induce tbx3 in vitro. Since we have also found that Tbx3 is required for eye formation, we sought to determine how it is regulated. Here, we show that in vivo tbx3 expression is affected by both BMP and Activin signaling, and that the BMP/Activin chemical inhibitor LDN-193189 preferentially expands tbx3 expression in the eye field while repressing the cement gland, the anterior-most structure in the frog embryo. We further show that otx2, tbx3 and rax expression are all expanded in the eye field following LDN-193189 treatment. To determine how tbx3 is regulated in the eye, we identify evolutionarily conserved regions of the tbx3 gene that drive GFP expression in the eyes of transgenic tadpoles. We observed direct binding by myc-tagged Otx2 to activate tbx3 and one region that myc-tagged Otx2 directly binds. Together, these findings close the regulatory loop, demonstrating that tbx3 expression and early eye formation are directly regulated by Otx2-mediated activation.

X-linked juvenile retinoschisis (XLRS) is a hereditary retinal dystrophy resulting from mutations in the retinoschisin (RS1) gene, which lead to progressive structural abnormalities in the retina and consequent visual impairment. Despite this, the molecular mechanisms through which RS1 mutations induce cell type-specific transcriptional remodeling during disease progression remain inadequately elucidated. To delineate transcriptional changes associated with RS1 mutations, we conducted single-cell RNA sequencing (scRNA-seq) on a CRISPR/Cas9-engineered Rs1[emR209C] knock-in mouse model. Integrative analysis with the Mouse Retina Cell Atlas demonstrated that rod photoreceptors exhibited the most significant transcriptional dysregulation among retinal cell types. Notably, dipeptidyl peptidase-like protein 10 (DPP10) was progressively and markedly upregulated, predominantly within photoreceptors, as the disease advanced. These findings were corroborated by immunofluorescence staining, quantitative PCR, and western blot analyses, which confirmed increased retinal DPP10 expression concurrent with a gradual decline in RS1 levels. To explore the functional implications of DPP10 dysregulation, we developed Y79 retinal cell models expressing either wild-type RS1 or the XLRS-associated RS1 R209C variant. Cells expressing the RS1 R209C mutation exhibited elevated DPP10 expression alongside reduced viability. Crucially, siRNA-mediated knockdown of DPP10 partially rescued cell viability and was associated with reduced GRP78/BiP expression in RS1 R209C-expressing cells, supporting potential functional relevance between DPP10 upregulation and mutant RS1-induced cellular stress. Furthermore, circulating DPP10 concentrations were significantly increased in patients with XLRS compared to control subjects. Collectively, these results identify DPP10 as a molecular alteration associated with RS1 mutation and suggest its potential as a circulating biomarker candidate for XLRS.

Corneal fibrosis is a dysregulated wound-healing response in which epithelial and basement membrane failure, limbal dysfunction, unresolved inflammation, persistent myofibroblasts, vascular amplification, and extracellular matrix disorganisation impair corneal transparency and vision. Although transforming growth factor-β is central, persistence reflects interconnected epithelial, stromal, immune, vascular, and mechanobiological networks. This review frames fibrosis as a stage-dependent continuum from early fibrotic commitment through active stromal remodelling to established structural scar and evaluates interventions according to their most plausible biological window. Anti-inflammatory and pathway-directed treatments are considered primarily as preventive or active-stage interventions, whereas limbal restoration, mesenchymal stromal cell- and extracellular vesicle-based approaches, amniotic membrane-derived interventions, bioengineered constructs, and keratoplasty address surface stabilisation, regeneration, or structural replacement. Current evidence supports prevention and modulation of early or active fibrosis more strongly than pharmacological reversal of mature scar and remains dominated by preclinical studies and small clinical datasets. Translation will require stage- and phenotype-matched models, objective structural and optical endpoints, biologically informed patient stratification, and clear separation of prevention, suppression of active remodelling, regenerative stabilisation, and true scar reversal.