Aqueous Humor Samples Research Articles

Fuch's Endothelial Corneal Dystrophy in Cataract Patients Is Associated with Elevated Levels of Inflammatory Chemokines, but Not Growth Factors, in the Aqueous Humor.

The study investigated a profile of chemokines and growth factors in the aqueous humor (AH) of eyes with Fuch's endothelial corneal dystrophy (FECD) and cataracts in comparison with cataract patients as a control group. A total of 52 AH samples (26 FECD + cataract and 26 cataract/control) were collected before cataract surgery. None of the patients had any clinically apparent inflammation at the time of AH collection. The AH levels of MCP-1 (CCL2), MIP-1α (CCL3), MIP-1β(CCL4), RANTES (CCL5), eotaxin (CCL11), IP-10 (CXCL10), FGF basic, G-CSF, GM-CSF, PDGF-bb, and VEGF were compared between the groups. The analyses were performed using the Bio-Plex 200 System from Bio-Rad. Among the studied parameters, the AH levels of RANTES, eotaxin, and IP-10 significantly increased in the FECD + cataract eyes, compared with the cataract controls (p < 0.05). Elevated levels of the RANTES, Eotaxin, and IP-10 indicate more intense inflammation in the eyes of patients in the FECD + cataract group. Moreover, these factors exhibit potential as predictive biomarkers for early detection of FECD in cataract patients. The discovery of elevated concentrations of biochemical markers in a patient, who has not yet received a clinical diagnosis, may suggest the need for heightened observation of the other eye to monitor the potential development of FECD.

AHP DB: a reference database of proteins in the human aqueous humor.

The aqueous humor (AH) is a low-viscosity biofluid that continuously circulates from the posterior chamber to the anterior chamber of the eye. Recent advances in high-resolution mass-spectrometry workflows have facilitated the study of proteomic content in small-volume biofluids like AH, highlighting the potential clinical implications of the AH proteome. Nevertheless, in-depth investigations into the role of AH proteins in ocular diseases have encountered challenges due to limited accessibility to these workflows, difficulties in large-scale AH sample collection and the absence of a reference AH proteomic database. In response to these obstacles, and to promote further research on the involvement of AH proteins in ocular physiology and pathology, we have developed the web-based Aqueous Humor Proteomics Database (AHP DB). The current version of AHP DB contains proteomic data from 307 human AH samples, which were analyzed using liquid chromatography-tandem mass spectrometry (LC-MS/MS). The database offers comprehensive information on 1683 proteins identified in the AH samples. Furthermore, relevant clinical data are provided for each analyzed sample. Researchers also have the option to download these datasets individually for offline use, rendering it a valuable resource for the scientific community. Database URL: https://ahp.augusta.edu/.

Differences in aqueous humor protein profiles in patients with proliferative diabetic retinopathy before and after aflibercept treatment

PurposeTo investigate the changes in aqueous humor (AH) protein profiles before and after intravitreal aflibercept (IVA) treatment in patients with proliferative diabetic retinopathy (PDR).Methods5 PDR patients provided 10 samples of AH before and after IVA treatment (pre-group vs. post-group). Proteins were identified using liquid chromatography-tandem mass spectrometry. Then, bioinformatics was employed to investigate the functional significance of differentially expressed proteins (DEPs) and hub proteins.ResultsA total of 16 DEPs were identified, consisting of 8 downregulated proteins and 8 upregulated proteins. Bioinformatics analysis indicated that the most significantly enriched biological process was “blood coagulation, intrinsic pathway.” The most significantly enriched signaling pathway was “complement and coagulation cascades.” HBB, HPX, VEGFA, and CA1 were identified as hub proteins for IVA treatment.ConclusionsTogether with the downregulation of the intravitreal vascular endothelial growth factor level, IVA may also change the AH protein composition in PDR patients, with DEPs involved in the blood coagulation, intrinsic pathway, complement, and coagulation cascades. IVA treatment may protect against PDR by regulating HBB, HPX, VEGFA, and CA1 expression.

Influence of Endogenous Substances on Site-II to Site-I Displacement of Diclofenac Bound to Albumin in the Aqueous Humor of Patients with Cataract.

Diclofenac instillation is useful in preventing intraoperative miosis and macular edema caused by postoperative inflammation in cataract surgery; however, optimum efficacy is not attained when the instilled diclofenac strongly binds to albumin in patients' aqueous humor. Therefore, a method that inhibits diclofenac binding and increases the concentration of its free fraction is needed. We conducted a basic study regarding the effects of inhibitors on the binding of instilled diclofenac to albumin and endogenous substances in aqueous humor. Aqueous humor samples from 16 patients were pooled together for analysis. The free fraction of diclofenac was measured using ultrafiltration methods in various experiments with pooled and mimic aqueous humor. Free fraction of diclofenac, a site II drug, in pooled aqueous humor was 0.363 ± 0.013. The binding of diclofenac in the presence of phenylbutazone (PB), a site I inhibitor, was significantly inhibited (free fraction = 0.496 ± 0.013); however, no significant inhibition by ibuprofen, a site II inhibitor, (free fraction = 0.379 ± 0.004), was observed. The unexpected result was due to free fatty acids (FFAs; palmitic acid (PA)) and L-tryptophan (Trp). The inhibition of diclofenac binding by PB in the mimic aqueous humor containing these endogenous substances revealed significant binding inhibition in the presence of PA and Trp. Diclofenac is strongly rebound from site II to site I in the presence of FFAs and Trp in the aqueous humor because FFAs and Trp induce a conformational change in albumin. Therefore, PB significantly inhibits the binding of diclofenac to albumin.

Low expression of TGF-β2 and matrilin2 in human aqueous humour with acute primary angle closure.

Primary angle-closure glaucoma (PACG) is the leading cause of irreversible blindness in the world. Angle closure induced by pupil block and secondary iris synechia is the fundamental pathology of the PACG. The molecular mechanisms of angle closure have not yet been clearly illustrated. This study was designed to investigate the protein difference in the aqueous humour and explore new biomarker of the PACG. Aqueous humour (AH) was collected from patients with acute primary angle closure (APAC) and cataract (n = 10 in APAC group) and patients with cataract only (n = 10 in control group). Samples were pooled and measured using label-free proteome technology. Then, the differentially expressed proteins (DEPs) were verified by ELISA using independent AH samples (n = 20 each group). More than 400 proteins were revealed in both groups through proteomics. Comparing the two groups, there were 91DEPs. These proteins participate in biological activities such as inflammation, fibrosis, nerve growth and degeneration and metabolism. We found that the expression of transforming growth factor-β2 and matrilin2 was downregulated in the APAC group. The two proteins are related to inflammation and extracellular matrix formation, which might be involved in angle closure. This study characterized DEPs in AH of the APAC and found a downregulated protein matrilin2.

Expression of GNAQ, BAP1, SF3B1, and EIF1AX Proteins in the Aqueous Humor of Eyes Affected by Uveal Melanoma.

The purpose of this study was to quantify specific aqueous humor (AH) proteins in eyes affected by posterior uveal melanoma (UM). Thirty-six eyes affected by primary UM were included. Tumor thickness and largest basal diameter were specific clinical characteristics. Tumors were staged with the American Joint Commission on Cancer Eighth Edition (AJCC) classification. During the brachytherapy (Iodine-125) surgical procedure, both the AH sample collection and the 25-gauge transscleral fine needle aspiration biopsy (FNAB) were performed. AH samples were analyzed by immunoprecipitation and SDS PAGE techniques to quantify GNAQ, BAP1, SF3B1, and EIF1AX proteins. Cytologic material underwent fluorescence in situ hybridization for chromosome 3. The AH of 36 healthy eyes was used as the control group. Cluster analysis of groups was also performed. Compared with the control group, significantly higher protein levels of: GNAQ (P = 0.02), BAP1 (P = 0.01), and SF3B1 (P = 0.02) were detected in eyes with UM. Cluster analysis of UM group revealed 2 clusters, one showing higher expression of GNAQ and BAP1 protein and one of EIF1AX protein. Moreover, the 2 clusters corresponded with the chromosome 3 status of UM. Specific and selected proteins may be detected in the AH of eyes affected by UM. These findings confirm the possibilities provided by AH analysis in UM.

A study of the relationship between cytokine levels and the response to anti-VEGF therapy in polypoid choroidal vasculopathy with different choroidal thicknesses.

Polypoidal choroidal vasculopathy (PCV) is an irreversible retinal choroidal disease. Individuals with PCV exhibit diverse baseline characteristics, including systemic characteristics, ocular traits, metabolic factor levels, and different responses to intravitreal anti-VEGF therapy. This study aims to investigate the pathogenesis of PCV by analyzing the systemic characteristics, ocular traits, and cytokine levels at baseline within a cohort of patients who exhibit different responses to anti-VEGF treatment. We conducted a retrospective analysis involving 80 eyes diagnosed with PCV. Patients were categorized into two groups based on responses to suboptimal intravitreal ranibizumab injection therapy: those with suboptimal responses and optimal responses. Aqueous humor samples were collected from the experimental eyes, and cytokine expression levels were assessed using cytometric bead array analysis. All subjects were further stratified into two groups according to the median choroidal thickness. Subsequently, logistic regression analysis and the ROC curve were employed to examine the relationship between cytokine expression levels, choroidal thickness, and anti-VEGF response. The results revealed that compared to the group of optimal anti-VEGF response, the choroid in the suboptimal response group exhibited a significantly greater thickness. Additionally, compared to the suboptimal anti-VEGF response group, the expression levels of VEGF and VCAM-1 were markedly lower observed in the optimal anti-VEGF response group, while TNF-α showed the opposite trend. Logistic regression analysis indicated that VEGF, VCAM-1, and TNF-α in the aqueous humor were independent risk factors for a suboptimal anti-VEGF response. After adjusting other risk factors, the risk of suboptimal anti-VEGF response decreased to 0.998-fold, 0.997-fold, and 1.294-fold. The AUC values for VEGF, VCAM-1, and TNF-α were determined to be 0.805, 0.846, and 0.897, respectively. Furthermore, the risk of VEGF, VCAM-1, and TNF-α were significantly associated with an increased risk of suboptimal anti-VEGF response after correction for risk factors in the thick choroid group. Our study demonstrated that PCV exhibits systemic and ocular characteristics variations based on different anti-VEGF responses. The levels of cytokines in aqueous humor were found to have a significant correlation with the anti-VEGF response in PCV. VEGF, VCAM-1, and TNF-α are potential targets for assessing treatment response in thick choroidal PCV.

Preoperative Intravitreal Conbercept Injection Reduced Both Angiogenic and Inflammatory Cytokines in Patients With Proliferative Diabetic Retinopathy.

Aims: To investigate the impact of intravitreal injection of conbercept, a recombinant fusion protein with decoy receptors for the vascular endothelial growth factor (VEGF) family, on intraocular concentrations of angiogenic and inflammatory mediators in patients with proliferative diabetic retinopathy (PDR), analyzed its potential impact on surgical outcomes. Methods: Forty eyes from 40 patients with PDR were included in this prospective study. Patients received intravitreal injection of conbercept followed by vitrectomy or phacovitrectomy in 1 week. Aqueous humor samples were collected before and 1 week after the conbercept injection. The concentrations of angiogenic and inflammatory cytokines and chemokines were measured by flow cytometry. Follow-up clinical data were collected and analyzed. Results: Intravitreal conbercept injection significantly decreased aqueous concentrations of VEGF (325.5 (baseline) versus 22.3 pg/mL (postinjection), p < 0.0001), PlGF (39.5 versus 24.5 pg/mL, p < 0.0001), and PDGF-A (54.1 versus 47.0 pg/mL, p = 0.0016), while no impact on bFGF levels. For inflammatory mediators, the concentration of TNF-α (0.79 versus 0.45 pg/mL, p = 0.0004) and IL-8 (180.6 versus 86 pg/mL, p < 0.0001) were decreased, while IL-6 (184.1 versus 333.7 pg/mL, p = 0.0003) and IL-10 (1.1 versus 1.5 pg/mL, p = 0.0032) were increased. No significant changes in IFN-γ or MCP-1 were detected. Three months after surgery, the mean best-corrected visual acuity improved from a baseline of 1.8 ± 0.1 logMAR to 0.7 ± 0.1 logMAR (p < 0.0001), with 36 eyes (90%) achieving an improvement of visual function. Conclusions: Intravitreal conbercept injection presents dual effects of antiangiogenesis and anti-inflammation and can be served as an adjuvant treatment to vitrectomy for PDR patients.

The impact of lutein-loaded poly(lactic-co-glycolic acid) nanoparticles following topical application: An in vitro and in vivo study.

Antioxidant therapies are of interest in the prevention and management of ocular disorders such as cataracts. Although an active area of interest, topical therapy with antioxidants for the treatment of cataracts is complicated by multiple ocular anatomical barriers, product stability, and solubility. Entrapment and delivery of antioxidants with poly(lactic-co-glycolic acid) nanoparticles is a possible solution to these challenges, however, little is known regarding their effects in vitro or in vivo. Our first aim was to investigate the impact of blank and lutein loaded PLGA nanoparticles on viability and development of reactive oxygen species in lens epithelial cells in vitro. Photo-oxidative stress was induced by ultraviolet light exposure with cell viability and reactive oxygen species monitored. Next, an in vivo, selenite model was utilized to induce cataract formation in rodents. Eyes were treated topically with both free lutein and lutein loaded nanoparticles (LNP) at varying concentrations. Eyes were monitored for the development of anterior segment changes and cataract formation. The ability of nanodelivered lutein to reach the anterior segment of the eye was evaluated by liquid chromatography coupled to mass spectrometry of aqueous humor samples and liquid chromatography coupled to tandem mass spectrometry (targeted LC-MS/MS) of lenses. LNP had a minimal impact on the viability of lens epithelial cells during the short exposure timeframe (24 h) and at concentrations < 0.2 μg LNP/μl. A significant reduction in the development of reactive oxygen species was also noted. Animals treated with LNPs at an equivalent lutein concentration of 1,278 μg /mL showed the greatest reduction in cataract scores. Lutein delivery to the anterior segment was confirmed through evaluation of aqueous humor and lens sample evaluation. Topical treatment was not associated with the development of secondary keratitis or anterior uveitis when applied once daily for one week. LNPs may be an effective in the treatment of cataracts.

Cytomegalovirus detected by qPCR in iris and ciliary body of immunocompetent corneal donors

BackgroundCytomegalovirus (CMV) can cause a wide panel of ocular infections. The involvement of CMV as a cause of anterior uveitis in the immunocompetent patient is recent and remains poorly understood. ObjectiveTo investigate the presence of CMV in anterior uveal tissues of immunocompetent corneal donors. Study DesignWe collected aqueous humor, iris, and ciliary body from both eyes of 25 donors died at the Limoges University Hospital between January 2020 and July 2021. CMV serology was determined for all patients from post-mortem blood sample. Ocular tissues were split in 2 fragments for qPCR and 2 for histological analysis. CMV genomes copies were quantified by Multiplex qPCR after DNA extraction. Results16 of 25 patients (64%) displayed positive CMV serology, with a median age of 67 years. Viremia was positive in 3 of 16 (19%) CMV-positive patients. No CMV DNA copies were found from the aqueous humor samples. CMV DNA was detected in iris and ciliary body of 28 of 32 eyes of seropositive donors, and 5 of 18 eyes of seronegative donors. The median viral copy number [IQR] was 2.41 × 102 [8.91 × 101 - 1.01 × 103] copies/1 × 106 cells in the CMV-positive group and 0.00 [0.00 - 3.54 × 102] copies/1 × 106 cells in the CMV-negative group (p<0.001). Histology and immunohistochemistry did not reveal any CMV lesions from any sample. ConclusionCMV DNA was found in iris and ciliary body of immunocompetent seropositive patients, but also, although less frequently, from seronegative donors. These results highlight mechanisms of infection, latency and reactivation of CMV in ocular tissues.

MiRNA profiling in the aqueous humour in patients with Fuch's endothelial corneal dystrophy

Aims: The aim of the study was to determine and compare the expression profiles of miRNA molecules in the aqueous humour (AH) of patients with Fuch's endothelial corneal dystrophy (FECD) and patients with normal corneal endothelium undergoing cataract surgery.Methods: The study consisted of 20 Caucasian patients (10 FECD and 10 control patients) who underwent comprehensive ophthalmology examination, thus routine corneal transplant or cataract surgery. 100 μL of AH were obtained from the anterior chamber at the beginning of the surgery with special care to avoid contamination with blood tears. The total RNA was isolated from the AH samples using an miRNeasy Serum/Plasma kit according to the manufacturer's instructions. The RNA concentration was determined by using NanoDrop2000c spectrophotometer. In addition, RNA analysis was performed using Agilent Bioanalyzer2100 and PicoRNA Kit according to the manufacturer's protocol. A microarray system (GeneChip miRNA 4.0 Array chip, Affymetrix) was used to determine the miRNA expression profiles. The RNA preparation and hybridization were performed according to the manufacturer's protocol, with one modification to extend the hybridization time to 42 h. The gene chips were scanned with an Affymetrix GeneChip Scanner 3000.Results: We found significant overexpression of hsa‐miR‐4532, hsa‐miR‐6800‐5p, hsa‐miR‐3621, hsa‐miR‐4270, hsa‐miR‐548ac, hsa‐miR‐328‐5p, hsa‐miR‐6771 ‐5p, hsa‐miR‐6858‐5p, hsa‐miR‐548a‐3p, hsa‐miR‐6069, hsa‐miR‐6716‐5p, hsa‐miR‐7151‐3p, hsa‐miR‐5189‐5p, hsa ‐miR‐4279, hsa‐miR‐1233‐5p, hsa‐miR‐455‐3p in patients with FECD compared to those patients with normal corneal endothelium. Reduced expression of hsa‐miR‐3613‐3p, hsa‐miR‐1298‐3p, hsa‐miR‐16‐2‐3p, hsa‐miR‐4668‐5p was observed in patients with FECD in addition to the control group.Conclusions: The analysis of miRNA expression profiles and their application as potential biomarkers will facilitate and accelerate the diagnosis of FECD, as well as enable the development of individualized molecularly targeted therapy.

A clinical metagenomic study of biopsies from Mexican endophthalmitis patients reveals the presence of complex bacterial communities and a diversity of resistance genes.

Infectious endophthalmitis is a severe ophthalmic emergency. This infection can be caused by bacteria and fungi. For efficient treatment, the administration of antimicrobial drugs to which the microbes are susceptible is essential. The aim of this study was to identify micro-organisms in biopsies of Mexican endophthalmitis patients using metagenomic next-generation sequencing and determine which antibiotic resistance genes were present in the biopsy samples. In this prospective case study, 19 endophthalmitis patients were recruited. Samples of vitreous or aqueous humour were extracted for DNA extraction for metagenomic next-generation sequencing. Analysis of the sequencing results revealed the presence of a wide variety of bacteria in the biopsies. Resistome analysis showed that homologues of antibiotic resistance genes were present in several biopsy samples. Genes possibly conferring resistance to ceftazidime and vancomycin were detected in addition to various genes encoding efflux pumps. Our findings contrast with the widespread opinion that only one or a few bacterial strains are present in the infected tissues of endophthalmitis patients. These diverse communities might host many of the resistance genes that were detected, which can further complicate the infections.

Ocular pharmacokinetics of a new preservative‐free bimatoprost 0.01% ophthalmic gel

Aims/Purpose: The aim of this study was to compare the aqueous humour (AH) and iris‐ciliary body (ICB) concentrations of bimatoprost and its main metabolite bimatoprost free acid after topical instillation in rabbits of a new preservative‐free bimatoprost 0.01% ophthalmic gel (PFB 0.01% gel) versus two commercially available preserved bimatoprost ophthalmic solutions ‐ PB 0.01% (200 ppm benzalkonium chloride [BAK], Lumigan® 0.01%) and PB 0.03% (50 ppm BAK, Lumigan® 0.03%).Methods: Female Dutch Belted rabbits (n = 45) were divided into three treatment groups (30 eyes per group): PFB 0.01% gel, PB 0.01% and PB 0.03% solutions. Animals received a single topical instillation (25 μL) into both eyes. AH and ICB samples were collected at 0.5 h, 1 h, 2 h, 4 h and 8 h post instillation. These samples were analysed by rapid resolution liquid chromatography tandem mass spectrometry for bimatoprost and bimatoprost free acid concentrations.Results: Following a single topical instillation, Tmax of bimatoprost and bimatoprost free acid in both ocular matrices was 1 h across the 3 treatments. Cmax of bimatoprost/bimatoprost free acid was 50.2 ng/mL, 26.3 ng/mL and 59.9 ng/mL of AH respectively following PFB 0.01%, PB 0.01% and PB 0.03% treatments. Maximal concentrations and area under the curve in AH and ICB of bimatoprost/bimatoprost free acid were greater (from 2‐ to 2.6‐fold) with PFB 0.01% gel compared to PB 0.01% and similar (from 0.9‐ to 0.95‐fold) to PB 0.03%.Conclusions: Topical instillation of PFB 0.01% gel produced significantly higher bimatoprost/bimatoprost free acid concentrations in the AH and ICB of rabbits than PB 0.01% (despite a high BAK concentration) and was similar to PB 0.03%. The preservative‐free gel formulation improves corneal penetration and bioavailability of bimatoprost compared to a preserved solution at the same concentration.

A connection between levels of soluble Fas and Fas ligand in the aqueous humor and the parameters of structural and functional damage of glaucoma patients

Introduction/Objective. Fas ligant (FasL) induces apoptosis when interacting with the Fas-receptor. The aim was to determine the concentration of soluble Fas (sFas) and FasL (sFasL) in the aqueous humor of open-angle glaucoma patients, and establish a connection between these markers of apoptosis and the parameters of structural and functional glaucoma damage. Methods. This study examined 88 aqueous humor samples; 35 primary open-angle glaucoma with elevated intraocular pressure (POAG-HTG) patients, 24 open angle pseudoexfoliative glaucoma patients (XFG) and 29 patients with senile cataract (CAT). The concentration of sFas and sFasL was determined by commercial ELISA tests in the aqueous humor. Results. The conducted study showed that aqueous humor sFas concentration was the highest in XFG (720.14 ? 167.39 pg/ml), and slightly lower in POAG-HTG (713.43 ? 162.69 pg/ml), than in cataract patients (632.46 ? 217.11 pg/ml), without statistical significance. There was a significant negative correlation of sFas concentration and RNFL Inf thickness in POAG-HTG (p &lt; 0.05). The concentration of sFasL was the lowest in POAG-HTG (9.28 ? 0.551 pg/ml), higher in XFG (9.45 ? 0.61 pg/ml; p = 0.0566), and the highest in the cataract group (9.48 ? 0.73 pg/ml). A negative correlation of sFasL and MD in the POAG-HTG, and a negative correlation with RNFL Sup in the XFG were significant. Conclusion. sFasL has an active role in the regulation of the inflammatory process in glaucoma. sFas and sFasL, as markers of apoptosis, are associated with the parameters of structural, RNFL thinning, and functional glaucoma damage, namely visual field defects.

Clinical pattern and outcomes of acute retinal necrosis in a referral Center in Tunisia, North Africa: A 17‐year retrospective study

Aims/Purpose: To describe the pattern of acute retinal necrosis (ARN) in a referral centre in Tunisia, North Africa.Methods: The charts of 22 patients diagnosed with ARN from January 2006 to January 2023 were retrospectively reviewed.Results: The mean age at presentation was 39.2 (range, 12–70) years, sex ratio was 1.75. The mean diagnostic delay was 17.9 days (range 1–60). The mean visual acuity at presentation was 20/400. All patients presented with a mild to moderate anterior uveitis (100%) with granulomatous keratic precipitates in 18 eyes (81%) and ocular hypertension in 4 eyes (18%). Necrotizing retinitis involved &gt;50% of the retina in 36.5%of eyes. Polymerase chain reaction on aqueous humour sample identified varicella zoster virus in 10 eyes (45%), herpes simplex virus‐1 in 4 eyes (18%) and was negative in 8 eyes (22%). All patients received intravenous acyclovir, followed by oral antivirals. Intravitreal injections of ganciclovir were administered in 12 eyes (54.5%). Follow‐up ranged from 1 month to 48 months. The mean final visual acuity was 20/200. Complications included retinal detachment (RD) (31.8%), macular oedema (18%), macular atrophy (22%), epimacular membrane (22%), cataract (27%), optic nerve atrophy (18%) and central retinal artery occlusion (4%). No patient developed bilateral ARN. Prophylactic laser was performed in 7 eyes (32%). RD was treated with pars plana vitrectomy and silicone oil injection in 7 eyes.Conclusions: ARN syndrome in Tunisia is most commonly caused by VZV.

Clinicopathologic Analyses of Failed Endothelial Keratoplasty (the Clinical Cases)

Purpose: To evaluate the clinicopathologic features of failed endothelial keratoplasty. Methods. In this study 11 patients (11 eyes) with recurrence of bullous keratopathy (BK) were included. Group 1 consisted of 4 patients who underwent repeat Descemet Stripping Automated Endothelial Keratoplasty (DSAEK), group 2 included 7 patients who underwent penetrating keratoplasty (PK) after failed Descemet Membrane Endothelial Keratoplasty (DMEK). Preoperative anterior segment optical coherence tomography (OCT), RTVue-100, Optovue, USA, was performed. Intraoperatively aqueous humour (AqH) samples were collected for multiplex cytokine analysis. During keratoplasty failed grafts/corneal buttons were obtained and then investigated histologically (hematoxylin/eosin staining, primary antibodies to pancytokeratin, vimentin, collagen III). Results. Recurrence of BK in all clinical cases manifests by the increase of inflammatory factors in AqH, corneal edema, neovascularization and remodeling to fibrosis. Glaucoma may induce DMEK/ DSAEK graft failure due to chronic local inflammation. In the clinical cases recurrence of BK was caused by peripheral and central graft detachment, rebubbling, graft upside-down orientation and donor corneal genetic disorders. Conclusions. Chronic local inflammation (including corneal morphological changes) in patients with BK recurrence is an indication for reoperation. The modification of keratoplasty — DMEK/DSAEK/PK — is determined according to slit-lamp and OCT images of the cornea. High-level risk of immune reaction (especially in cases of three and more times repeated keratolasty) is the reason for systemic corticosteroid and, sometimes, cytostatic therapy.

Isolation of three different sizes of exosomes in an Asian population with different retinal diseases before and after treatment: preliminary results

ABSTRACT Exosomes are membranous structures measuring between 40–120 nm that are secreted by various cells of the human body into the body fluid system. Exosomes contain proteins, mRNA, miRNA, and signaling molecules, and physiologically they assist in the intercellular transport of proteins and RNA molecules. In this study, we used an immunoaffinity filter paper platform combined with scanning electron microscopy and microfluidic systems to detect the size of exosomes within the aqueous humor. Eight aqueous humor samples showed three distinct sizes of exosomes that were significantly different on scanning electron microscopy(P < 0.01). We further used nanoparticle tracking analysis to assess the size distribution of exosomes within the aqueous humor. We found significantly different distributions of exosomes between patients with three different ocular diseases and patients with normal cataracts as controls. An obvious peak of exomeres(size around 35 nm)was found in the patients with central retinal vein occlusion and vitreous hemorrhage. Flare-ups of large exosomes(size 90–120 nm)were found in the patients with the inflammatory ocular disease pars planitis. No obvious peaks in exomeres or large exosomes were found in the control group. There was a high association between the distribution of exosomes and the pathogenesis of ocular diseases. After intravitreal anti-vascular endothelial growth factor treatment, the aqueous humor from the patients with neovascular diseases showed a significant reduction in exosomes in nanoparticle tracking analysis. These findings suggest that at least three distinct sizes of exosomes exist in the aqueous humor:(1)exomeres:<35 nm;(2)small exosomes:60–80 nm; and (3)large exosomes:90–120 nm. Different sizes of exosomes may have different implications in normal or diseased eyes.

Insights into myopic choroidal neovascularization based on quantitative proteomics analysis of the aqueous humor

BackgroundPrevious studies on the biomarkers of pathologic myopia choroidal neovascularization (pmCNV) development merely detected limited types of proteins and provide a meagre illustration of the underlying pathways. Hence, a landscape of protein changes in the aqueous humor (AH) of pmCNV patients is lacking. Here, to explore the potential mechanisms and biomarkers of pmCNV, we analyzed the clinical data and protein profile among atrophic (A) lesions, tractional lesions (T) and neovascular (N) lesions in myopic patients based on the ATN grading system for myopic maculopathy (MM).ResultsAfter investigating demographic data of our patients, a correlation was found between A and N lesions (R = 0.5753, P < 0.0001). Accordingly, groups were divided into patients without MM, patients with myopic atrophic maculopathy (MAM), and patients with pmCNV (N2a lesion). In proteomics analysis, the increased protein level of GFAP and complement-associated molecules in AH samples of the 3 groups also indicated that MAM and pmCNV shared similar characteristics. The GO enrichment and KEGG pathway analysis were performed, which mapped that differential expressed proteins mainly engaged in JAK-STAT pathway between the pmCNV group and two controls. Furthermore, we identified several potential biomarkers for pmCNV, including FCN3, GFAP, EGFR, SFRP3, PPP2R1A, SLIT2, and CD248.ConclusionsAtrophic lesions under pathologic myopic conditions demonstrated similarities to neovascularization development. Potential biomarkers including GFAP were associated with the pathogenesis of pmCNV. In summary, our study provides new insights for further research on pmCNV development.

Evaluating the long-term biological stability of cytokine biomarkers in ocular fluid samples

PurposeThe quality of biological fluid samples is vital for optimal preanalytical procedures and a requirement for effective translational biomarker research. This study aims to determine the effects of storage duration...

Cytokines in aqueous humor of patients with congenital cataract during delayed sequential bilateral cataract surgery

BackgroundTo explore the changes of cytokines expression in aqueous humor (AH) of eyes of patients with congenital cataract (CC) who underwent delayed sequential bilateral cataract surgery (DSBCS).Methods28 patients with CC underwent DSBCS. AH samples were collected from each eye before surgery. The contents of cytokines in AH were detected by Luminex xMAP Technology.ResultsThere was no significant difference in the expression of IL-8, IP-10, MCP-1 and PDGFAA in the AH of the first and second eyes (P = 0.35, 0.39, 0.17, respectively). The level of IL-8 in the first-eye AH was negatively correlated with age (ρ=– 0.519, P = 0.008). IP-10 and MCP-1 in the second-eye AH were negatively correlated with age (ρ=– 0.483, P = 0.009; ρ=– 0.445, P = 0.018,respectively).ConclusionThe first-eye surgery in patients with CC may not cause the change of cytokines in the contralateral eye. The expression of IL-8, IP-10 and MCP-1 in the AH was negatively correlated with the age of patients.Trial registrationThe study was registered at www.chictr.org.cn on March 22, 2022 and the clinical trial number is ChiCTR2200057927.