Composition of the gut microbiome, role of diet, lifestyle, and antioxidant therapies in diabetes mellitus and diabetic retinopathy.

The gut microbiome is a complex ecosystem in the gastrointestinal tract composed of trillions of bacteria, viruses, fungi, and protozoa. Disruption of this delicate ecosystem, formally called "dysbiosis", has been linked to a variety of metabolic and inflammatory pathologies. Several studies have focused on abnormal microbiome composition and correlated these findings with the development of type 2 diabetes mellitus (T2DM) and diabetic retinopathy (DR). However, given the complexity of this ecosystem, the current studies are narrow in design and present variable findings. Composition of the gut microbiome in patients with DR significantly differs from patients with diabetes without retinopathy as well as from healthy controls. Additionally, the gut microbiome has been shown to modify effects of medication, diet, exercise, and antioxidant use on the development and progression of DR. In this paper, we present a comprehensive review of literature on the effect of oxidative stress, antioxidant therapies, and dysbiosis on DR.

MMP inhibition as a novel strategy for extracellular matrix preservation during whole liver decellularization.

As the only reliable treatment option for end-stage liver diseases, conventional liver transplantation confronts major supply limitations. Accordingly, the decellularization of discarded livers to produce bioscaffolds that support recellularization with progenitor/stem cells has emerged as a promising translational medicine approach. The success of this approach will substantially be determined by the extent of extracellular matrix (ECM) preservation during the decellularization process. Here, we assumed that the matrix metalloproteinase (MMP) inhibition could reduce the ECM damage during the whole liver decellularization of an animal model using a perfusion-based system. We demonstrated that the application of doxycycline as an MMP inhibitor led to significantly higher preservation of collagen, glycosaminoglycans, and hepatic growth factor (HGF) contents, as well as mechanical and structural features, including tensile strength, fiber integrity, and porosity. Notably, produced bioscaffolds were biocompatible and efficiently supported cell viability and proliferation in vitro. We also indicated that produced bioscaffolds efficiently supported HepG2 cell function upon seeding onto liver ECM discs using albumin and urea assay. Additionally, MMP inhibitor pretreated decellularized livers were more durable in contact with collagenase digestion compared to control bioscaffolds in vitro. Using zymography, we confirmed the underlying mechanism that results in these promising effects is through the inhibition of MMP2 and MMP9. Overall, we demonstrated a novel method based on MMP inhibition to ameliorate the ECM structure and composition preservation during liver decellularization as a critical step in fabricating transplantable bioengineered livers.

Age-related macular degeneration and neurodegenerative disorders: Shared pathways in complex interactions.

Age-related macular degeneration (AMD) is a leading cause of irreversible blindness in the elderly, and neurodegenerative disorders such as Alzheimer disease and Parkinson disease are debilitating conditions that affect millions worldwide. Despite the different clinical manifestations of these diseases, growing evidence suggests that they share common pathways in their pathogenesis including inflammation, oxidative stress, and impaired autophagy. In this review, we explore the complex interactions between AMD and neurodegenerative disorders, focusing on their shared mechanisms and potential therapeutic targets. We also discuss the current opportunities and challenges for developing effective treatments that can target these pathways to prevent or slow down disease progression in AMD. Some of the promising strategies that we explore include modulating the immune response, reducing oxidative stress, enhancing autophagy and lysosomal function, and targeting specific protein aggregates or pathways. Ultimately, a better understanding of the shared pathways between AMD and neurodegenerative disorders may pave the way for novel and more efficacious treatments.

Exosomal Cargo: Pro-angiogeneic, anti-inflammatory, and regenerative effects in ischemic and non-ischemic heart diseases - A comprehensive review.

Heart diseases are the primary cause of mortality and morbidity worldwide which inflict a heavy social and economic burden. Among heart diseases, most deaths are due to myocardial infarction (MI) or heart attack, which occurs when a decrement in blood flow to the heart causes injury to cardiac tissue. Despite several available diagnostic, therapeutic, and prognostic approaches, heart disease remains a significant concern. Exosomes are a kind of small extracellular vesicles released by different types of cells that play a part in intercellular communication by transferring bioactive molecules important in regenerative medicine. Many studies have reported the diagnostic, therapeutic, and prognostic role of exosomes in various heart diseases. Herein, we reviewed the roles of exosomes as new emerging agents in various types of heart diseases, including ischemic heart disease, cardiomyopathy, arrhythmia, and valvular disease, focusing on pathogenesis, therapeutic, diagnostic, and prognostic roles in different areas. We have also mentioned different routes of exosome delivery to target tissues, the effects of preconditioning and modification on exosome's capability, exosome production in compliance with good manufacturing practice (GMP), and their ongoing clinical applications in various medical contexts to shed light on possible clinical translation.

Conditional deletion of CD25 in the corneal epithelium reveals sex differences in barrier disruption.

PURPOSE: IL-2 promotes activation, clonal expansion, and deletion of T cells. IL-2 signals through its heterotrimeric receptor (IL-2R) consisting of the CD25, CD122 and CD132 chains. CD25 knockout (KO) mice develop Sjögren Syndrome-like disease. This study investigates whether corneal CD25/IL-2 signaling is critical for ocular health. METHODS: Eyes from C57BL/6 mice were collected and prepared for immunostaining or in-situ hybridization. Bulk RNA sequencing was performed on the corneal epithelium from wild-type and CD25KO mice. We generated a conditional corneal-specific deletion of CD25 in the corneal epithelium (CD25Δ/ΔCEpi). Corneal barrier function was evaluated based on the uptake of a fluorescent dye. Mice were subjected to unilateral corneal debridement, followed by epithelial closure over time. RESULTS: In C57BL/6 mice, CD25 mRNA was expressed in ocular tissues. Protein expression of CD25, CD122, and CD132 was confirmed in the corneal epithelium. Delayed corneal re-epithelization was seen in female but not male CD25KO mice. There were 771 differentially expressed genes in the corneal epithelium of CD25KO compared to wild-type mice. While barrier function is disrupted in CD25Δ/ΔCEpi mice, re-epithelialization rates are not delayed. CONCLUSIONS: All three chains of the IL-2R are expressed in the corneal epithelium. Our results indicate for the first time, deleting CD25 systemically in all tissues in the mouse and deleting CD25 locally in just the corneal epithelium compromises corneal epithelial barrier function, leading to dry eye disease in female mice. Future studies are needed to delineate the pathways used by IL-2 signaling to influence cornea homeostasis.

Dry eye disease and blinking behaviors: A narrative review of methodologies for measuring blink dynamics and inducing blink response.

Dry eye disease (DED) is a leading cause of ophthalmology clinical encounters with prevalence ranging from 8.7% to 64% in various populations. Blinking is an endogenous process to refresh the tear film, clear debris and maintain quality vision. Altered blinking behavior is a common feature of DED and is implicated in the pathology of the disease. However, lack of a comprehensive review on the relationship between altered blinking behavior and DED is notable in the literature. Blinking behavior may be an effect of DED due to an unstable tear film sensitizing a motor response or be its cause due to destabilization of the tear film in conditions such as benign essential blepharospasm. In this comprehensive review, we summarize the current models and theories of tear film dynamics and blinking behavior to better understand their connection to DED and introduce contemporary technologies and measurement tools used in the evaluation and induction of blinking behavior. We also describe future directions of research to better understand the relationship between DED and blinking and explore therapies that address the abnormal blinking component of DED.

The effect of decellularization protocols on characterizations of thermoresponsive and light-curable corneal extracellular matrix hydrogels.

To compare the effects of two decellularization protocols on the characteristics of fabricated COrnea Matrix (COMatrix) hydrogels. Porcine corneas were decellularized with Detergent (De) or Freeze-Thaw (FT)-based protocols. DNA remnant, tissue composition and α-Gal epitope content were measured. The effect of α-galactosidase on α-Gal epitope residue was assessed. Thermoresponsive and light-curable (LC) hydrogels were fabricated from decellularized corneas and characterized with turbidimetric, light-transmission and rheological experiments. The cytocompatibility and cell-mediated contraction of the fabricated COMatrices were assessed. Both protocols reduced the DNA content to < 0.1 µg/mg (native, > 0.5 µg/mg), and preserved the collagens and glycosaminoglycans. The α-Gal epitope remnant decreased by > 50% following both decellularization methods. We observed more than 90% attenuation in α-Gal epitope after treatment with α-galactosidase. The thermogelation half-time of thermoresponsive COMatrices derived from De-Based protocol (De-COMatrix) was 18 min, similar to that of FT-COMatrix (21 min). The rheological characterizations revealed significantly higher shear moduli of thermoresponsive FT-COMatrix (300.8 ± 22.5 Pa) versus De-COMatrix 178.7 ± 31.3 Pa, p < 0.01); while, this significant difference in shear moduli was preserved after fabrication of FT-LC-COMatrix and De-LC-COMatrix (18.3 ± 1.7 vs 2.8 ± 2.6 kPa, respectively, p < 0.0001). All thermoresponsive and light-curable hydrogels have similar light-transmission to human corneas. Lastly, the obtained products from both decellularization methods showed excellent in vitro cytocompatibility. We found that FT-LC-COMatrix was the only fabricated hydrogel with no significant cell-mediated contraction while seeded with corneal mesenchymal stem cells (p < 0.0001). The significant effect of decellularization protocols on biomechanical properties of hydrogels derived from porcine corneal ECM should be considered for further applications.

Induction of Innate Inflammatory Pathways in the Corneal Epithelium in the Desiccating Stress Dry Eye Model.

Purpose: To determine whether 24-hour exposure to the desiccating stress (DS) dry eye model induces NF-kB and NLRP3 inflammasome pathways in the mouse cornea epithelium. Methods: Six- to 8-week-old C57BL/6J mice were housed under normal humidity (nonstressed) or subjected to DS from a drafty, low-humidity environment combined with subcutaneous scopolamine four times/day for one day to suppress tear production (DS1). Cornea whole mounts were prepared for immunofluorescent staining, or the corneal epithelium was scraped for NF-kB p-p65 ELISA, Western blot, or real-time PCR to detect NF-kB and inflammasome pathway proteins and gene transcripts, respectively. Results: NF-kB phospho-p65 protein, nuclear NF-kB p-p65, and expression of the NF-kB inducible cytokines (IL-12a, IL-12b, and lymphotoxin b [Ltb]) and chemokine (CCL-2) genes were significantly increased in DS1 compared to nonstressed control. NLRP3 protein and RNA transcripts significantly increased in DS1. NLRP3 and Caspase-1 immunostaining increased in the cornea epithelium at DS1. At DS1 there was no change in IL-18 and a decrease in IL-1ß mRNA transcripts; however, levels of bound and total IL-18 protein increased at DS1, and the level of mature IL-1ß increased from DS1 to DS5. Conclusions: These findings indicate innate NF-kB and NLRP3 inflammasome inflammatory pathways are induced in the corneal epithelium within one day in the DS dry eye model. NF-kB activation was associated with increased expression of inflammatory mediators involved in dry eye. Induction of these pathways is accompanied by increased bound/total IL-18 and mature IL-1ß.

Artificial intelligence in retinal image analysis: Development, advances, and challenges.

Modern advances in diagnostic technologies offer the potential for unprecedented insight into ophthalmic conditions relating to the retina. We discuss the current landscape of artificial intelligence in retina with respect to screening, diagnosis, and monitoring of retinal pathologies such as diabetic retinopathy, diabetic macular edema, central serous chorioretinopathy, and age-related macular degeneration. We review the methods used in these models and evaluate their performance in both research and clinical contexts and discuss potential future directions for investigation, use of multiple imaging modalities in artificial intelligence algorithms, and challenges in the application of artificial intelligence in retinal pathologies.

Optical coherence tomography angiography measurements in Parkinson's disease: A systematic review and meta-analysis.

Optical coherence tomography angiography (OCT-A) is an ocular imaging technology that has emerged as a non-invasive tool to evaluate retinal microvascular changes in neurodegenerative diseases including Parkinson's disease (PD) and Alzheimer's disease. While several studies have reported on the presence of pathologic retinal microvascular alterations in PD, the utility of OCT-A as a biomarker for PD evaluation is still unclear. A systematic review and meta-analysis were performed to explore the current evidence for the role of OCT-A in PD published up until June 2022. PubMed, Scopus, and Web of Science databases were used to systematically identify relevant papers and a meta-analysis was conducted using Stata16 software according to the level of heterogeneity applying a random- or fixed-effect model. Thirteen studies of 925 eyes in the PD group and 1501 eyes in the control group assessing OCT-A findings in PD patients were included. The meta-analyses revealed that the foveal region of PD patients had a significantly lower vessel density in the superficial capillary plexus (SCP) compared to healthy controls but that there were no significant differences in the foveal avascular zone, the SCP in whole, parafoveal, and perifoveal regions, and deep capillary plexus. OCT-A metrics may act as a potential biomarker for a more accurate and early PD diagnosis. Still, the OCT-A algorithms and interchangeability between OCT-A devices require further standardization to draw clinical conclusions regarding their utility.

Quantitative assessment of botulinum toxin injection on blink rate in blepharospasm.

PURPOSE: To objectively measure the blink rate in patients with blepharospasm managed by botulinum toxin type-A injections. METHODS: In this prospective, non-interventional case series, the complete blink rates of subjects were measured before incobotulinumtoxina injection and at follow-up within 4 weeks using slow-motion video-taping. Additionally, subjects graded the frequency of blinking, the severity of light-sensitivity, and the severity and frequency of dry eye symptoms on a categorical visual analog scale. The results are reported as median (range). RESULTS: Ten subjects were enrolled, with nine females. The total duration of treatment was 70 (5-116) months with total of 27.5 (2-51) injections. The subjects were grouped as short-time (<52w) or long-time (>52w) treatments. The median age, follow-up time, and injected doses were 73.5 (49-81) years, 21 (14-28) days, and 38 (8-47) units, respectively, with no significant difference between groups. The total complete blinks per minute before incobotulinumtoxina injection was 39 (23-64) which decreased to 18.5 (1-60) at follow-up (p = 0.004). The average change in complete blink rate was -67.4 ± 23.7% in long-time and -45.2 ± 31.2% in short-time groups (mean ± SD, p = 0.01). The total self-graded frequency of blinking and light-sensitivity decreased significantly at follow-up (p = 0.004, p = 0.047, respectively). Similar patterns of subject reported grades were seen in both groups. CONCLUSION: Videotaping is a low-cost method for objective measurement of blink rate in blepharospasm patients after incobotulinumtoxina injection. There was a significant reduction in blink rate after incobotulinumtoxina injections with higher percentage of change in the long-time treatment group. Incobotulinumtoxina injection also significantly improves subjective photophobia.

A narrative review on the association of high intraocular pressure and glaucoma in patients with retinal vein occlusion.

Background and Objective: Retinal vein occlusion (RVO) is a major cause of vision loss and elevated intraocular pressure (IOP), high ocular perfusion pressure, and glaucoma are known ophthalmic risk factors for RVO. The aim of this paper is to provide the update on the association and management of high IOP/glaucoma and RVO. Methods: A literature review was performed in PubMed and Medline until May 2022 utilizing specific keywords and cross-matched reference lists. Key Content and Findings: The association of RVO with high IOP/glaucoma may be attributed to retinal ganglion cell loss due to retinal ischemia in high IOP and glaucoma. As new modalities showed, decreased optic disc perfusion, reduced density of blood vessels in the optic nerve head of glaucoma patients, changes in the peripapillary microvascular parameters, and decreased retinal nerve fiber layer (RNFL) thickness of the optic nerve head of eyes with RVO suggest a common pathway between RVO and glaucoma. Literature suggests the close follow up for glaucoma development among patients with non-arteriovenous (AV) crossing (optic cup or optic nerve sited) RVO in fellow eye and management of elevated IOP among RVO cases treated with anti-vascular endothelial growth factor (VEGF) antibodies/corticosteroids and those with preexisting primary open angle glaucoma (POAG). Conclusions: Determining potential patient responses to treatment and considering therapeutic options are challenging among patients with RVO and glaucoma. However, IOP lowering managements in preventing IOP spikes in patients with preexisting glaucoma and early treatment of macular edema in eyes with RVO is recommended.

New, potent, small molecule agonists of tyrosine kinase receptors attenuate dry eye disease.

Nerve growth factor (NGF), brain-derived neurotrophic factor (BDNF), and neurotrophin3 (NT-3) bind to tyrosine kinase (Trk) receptors, TrkA, TrkB, and TrkC, respectively. This study investigated the efficacy of novel molecule agonists of Trk receptors in an in vivo model of dry eye disease (DED). Small molecule TrkC agonist (C1) and a pan-Trk agonist (pan) were synthesized for this. C57BL/6J mice were subjected to desiccating stress (DS) and received bilateral eye drops of C1, pan, or vehicle (2x/day). Dry eye signs, inflammation and expression of corneal barrier function, and conjunctival goblet cell (GC) densities were measured as part of the DED phenotype. Corneal epithelial lysates were collected for either western blot or RNA extraction. Extracted total RNAs were used for NanoString analyses. Immunofluorescent staining was performed on whole-mount corneas using anti-TNFAIP3 and anti-EP4 antibodies. Compared to vehicle, mice subjected to desiccating stress and treated with agonists pan and C1 showed improved corneal barrier function, while C1 also increased GC density. NanoString analyses revealed upregulation of specific mRNA transcripts (Ptger4, Tnfaip3, Il1a and Ptger4, Tlr3, Osal1) in pan- and C1-treated corneas compared to vehicle-treated corneas. Western blots showed that pan and C1 decreased vehicle-induced NFkB nuclear translocation after DS for one day and increased EP4 and TNFAIP3 protein levels after 5 days of DS in corneal epithelium lysates. We conclude that small-molecule agonists of Trk receptors improve DED by decreasing NFkB activation and increasing protein expression of anti-inflammatory molecules TNFAIP3 and EP4. Surprisingly, the most efficacious small molecule agonists were not TrkA selective but TrkC and panTrk, suggesting that wider exploration of TrkB and C and pan Trk agonists are warranted in efforts to treat DED.

A Light-Curable and Tunable Extracellular Matrix Hydrogel for In Situ Suture-Free Corneal Repair.

Corneal injuries are a major cause of blindness worldwide. To restore corneal integrity and clarity, there is a need for regenerative bio-integrating materials for in-situ repair and replacement of corneal tissue. Here, we introduce Light-curable COrnea Matrix (LC-COMatrix), a tunable material derived from decellularized porcine cornea extracellular matrix containing un-denatured collagen and sulfated glycosaminoglycans. It is a functionalized hydrogel with proper swelling behavior, biodegradation, and viscosity that can be cross-linked in situ with visible light, providing significantly enhanced biomechanical strength, stability, and adhesiveness. Cross-linked LC-COMatrix strongly adheres to human corneas ex vivo and effectively closes full-thickness corneal perforations with tissue loss. Likewise, in vivo, LC-COMatrix seals large corneal perforations, replaces partial-corneal stromal defects and bio-integrates into the tissue in rabbit models. LC-COMatrix is a natural ready-to-apply bio-integrating adhesive that is representative of native corneal matrix with potential applications in corneal and ocular surgeries.

Single-cell transcriptional profiling of murine conjunctival immune cells reveals distinct populations expressing homeostatic and regulatory genes.

Immune cells in the exposed conjunctiva mucosa defend against environmental and microbial stresses. Expression profiling by single-cell RNA sequencing was performed to identify conjunctival immune cell populations expressing homeostatic and regulatory genes. Fourteen distinct clusters were identified, including myeloid cells (neutrophils, monocytes, macrophages), dendritic cells (DC), and lymphoid cells (B, T, γδT, ILC2, and NK) lineages. Novel neutrophil [lipocalin (Lcn2) high and low), and MHCIIlo macrophage (MP) clusters were identified. More than half of the cells map to myeloid and dendritic cell populations with differential expression profiles that include genes with homeostatic and regulatory functions: Serpinb2 (MHCIIlo macrophage), Apoe (monocyte), Cd209a (macrophage), Cst3 (cDC1), and IL4i1 in migratory DC (mDC). ILC2 expresses the goblet cell trophic factor IL-13. Suppressed inflammatory and activated anti-inflammatory/regulatory pathways were observed in certain myeloid and DC populations. Confocal immunolocalization of identity markers showed mDC (CCR7, FASCIN1) located on or within the conjunctival epithelium. Monocyte, macrophage, cDC1 and IL-13/IL-5+ ILC2 were located below the conjunctival epithelium and goblet cells. This study found distinct immune cell populations in the conjunctiva and identified cells expressing genes with known homeostatic and immunoregulatory functions.

IL-17 Producing Lymphocytes Cause Dry Eye and Corneal Disease With Aging in RXRα Mutant Mouse.

Purpose: To investigate IL-17 related mechanisms for developing dry eye disease in the Pinkie mouse strain with a loss of function RXRα mutation. Methods: Measures of dry eye disease were assessed in the cornea and conjunctiva. Expression profiling was performed by single-cell RNA sequencing (scRNA-seq) to compare gene expression in conjunctival immune cells. Conjunctival immune cells were immunophenotyped by flow cytometry and confocal microscopy. The activity of RXRα ligand 9-cis retinoic acid (RA) was evaluated in cultured monocytes and γδ T cells. Results: Compared to wild type (WT) C57BL/6, Pinkie has increased signs of dry eye disease, including decreased tear volume, corneal barrier disruption, corneal/conjunctival cornification and goblet cell loss, and corneal vascularization, opacification, and ulceration with aging. ScRNA-seq of conjunctival immune cells identified γδ T cells as the predominant IL-17 expressing population in both strains and there is a 4-fold increased percentage of γδ T cells in Pinkie. Compared to WT, IL-17a, and IL-17f significantly increased in Pinkie with conventional T cells and γδ T cells as the major producers. Flow cytometry revealed an increased number of IL-17+ γδ T cells in Pinkie. Tear concentration of the IL-17 inducer IL-23 is significantly higher in Pinkie. 9-cis RA treatment suppresses stimulated IL-17 production by γδ T and stimulatory activity of monocyte supernatant on γδ T cell IL-17 production. Compared to WT bone marrow chimeras, Pinkie chimeras have increased IL-17+ γδ T cells in the conjunctiva after desiccating stress and anti-IL-17 treatment suppresses dry eye induced corneal MMP-9 production/activity and conjunctival goblet cell loss. Conclusion: These findings indicate that RXRα suppresses generation of dry eye disease-inducing IL-17 producing lymphocytes s in the conjunctiva and identifies RXRα as a potential therapeutic target in dry eye.

Current and Emerging Therapies for Limbal Stem Cell Deficiency.

The corneal epithelium serves to protect the underlying cornea from the external environment and is essential for corneal transparency and optimal visual function. Regeneration of this epithelium is dependent on a population of stem cells residing in the basal layer of the limbus, the junction between the cornea and the sclera. The limbus provides the limbal epithelial stem cells (LESCs) with an optimal microenvironment, the limbal niche, which strictly regulates their proliferation and differentiation. Disturbances to the LESCs and/or their niche can lead to the pathologic condition known as limbal stem cell deficiency (LSCD) whereby the corneal epithelium is not generated effectively. This has deleterious effects on the corneal and visual function, due to impaired healing and secondary corneal opacification. In this concise review, we summarize the characteristics of LESCs and their niche, and present the current and future perspectives in the management of LSCD with an emphasis on restoring the function of the limbal niche.

Developing a pro-angiogenic placenta derived amniochorionic scaffold with two exposed basement membranes as substrates for cultivating endothelial cells.

Decellularized and de-epithelialized placenta membranes have widely been used as scaffolds and grafts in tissue engineering and regenerative medicine. Exceptional pro-angiogenic and biomechanical properties and low immunogenicity have made the amniochorionic membrane a unique substrate which provides an enriched niche for cellular growth. Herein, an optimized combination of enzymatic solutions (based on streptokinase) with mechanical scrapping is used to remove the amniotic epithelium and chorion trophoblastic layer, which resulted in exposing the basement membranes of both sides without their separation and subsequent damages to the in-between spongy layer. Biomechanical and biodegradability properties, endothelial proliferation capacity, and in vivo pro-angiogenic capabilities of the substrate were also evaluated. Histological staining, immunohistochemistry (IHC) staining for collagen IV, and scanning electron microscope demonstrated that the underlying amniotic and chorionic basement membranes remained intact while the epithelial and trophoblastic layers were entirely removed without considerable damage to basement membranes. The biomechanical evaluation showed that the scaffold is suturable. Proliferation assay, real-time polymerase chain reaction for endothelial adhesion molecules, and IHC demonstrated that both side basement membranes could support the growth of endothelial cells without altering endothelial characteristics. The dorsal skinfold chamber animal model indicated that both side basement membranes could promote angiogenesis. This bi-sided substrate with two exposed surfaces for cultivating various cells would have potential applications in the skin, cardiac, vascularized composite allografts, and microvascular tissue engineering.