Corneoscleral Rims Research Articles

Influence of dexamethasone-induced matrices on the TM transcriptome

Pathologic bidirectional interactions between the extracellular matrix (ECM) and cells within the human trabecular meshwork (hTM) contribute to ocular hypertension. An in vitro model is needed to study these cell-matrix interactions and their effect on outflow homeostasis. This study aimed to determine whether pathogenic ECM derived from dexamethasone (DEX)-treated hTM cultures induces clinically relevant glaucoma-like changes in healthy hTM cells at the transcriptional level. Corneoscleral rims from non-glaucoma donors were used to isolate primary hTM cells after validation according to the consensus recommendations for TM culture. Normal hTM cells (n = 5) were plated on a coverslip and treated with 100 nM DEX or ethanol for four weeks. These cultures were then decellularized, plated with primary hTM cells, and allowed to grow for another 72 h. RNA was extracted from these hTM cells for stranded total RNA-Seq. Sequencing libraries prepared using the Zymo-Seq RiboFree Total RNA library kit were pooled and sequenced using Illumina NovaSeq 6000. After quality control, sequence reads were aligned to the human genome build hg19. Differential expression (DE) analyses were performed using paired multi-factorial ANOVA. The expression of several DE genes associated with glaucoma (ANGPTL2, PDE7B, C22orf23, COL4A1, ADAM12, IFT122, SEMA6C) was validated using EvaGreen-based Droplet Digital PCR (ddPCR) assays. Gene ontology analyses of the DE genes were performed using the PANTHER and NDEx IQA databases, and functional analyses were performed with the DAVID Bioinformatics software. Using a cutoff of p-value <0.05 and fold change ≥2.0, our differential analysis identified 267 up- and 135 down-regulated genes in DEX-induced ECM-treated cells compared to the control. These differentially expressed genes were found to play a significant role in pathways such as cytokine and oxidative stress-induced inflammation, integrin signaling, matrix remodeling, and angiogenesis. These findings were further supported by previously performed proteomics studies using the same model. Using ddPCR, we validated the expression of seven genes associated with the risk of primary open-angle glaucoma. These results not only provide support for the pathogenic ECM model of steroid-induced glaucoma, but also demonstrate that the pathologic changes induced by this model are indeed found at the transcriptional level. These findings further demonstrate that matrix changes significantly influence cell expression profiles, which enable further understanding of the molecular mechanisms underlying glaucomatous changes in the TM. However, future studies with a larger and more diverse set of samples and longer time points are needed to confirm the utility of this model for mechanistic studies.

Investigating the Influence of Temperature and Supplementation Timing on Antifungal Efficacy in Storage Medium for Corneal Transplantation.

Although antifungal supplementation reduces the fungal load in the corneal storage medium, consensus is lacking on the influence of dosing and temperature. The study aims to evaluate the impact of eye bank warming protocol and timing of antifungal supplements on efficacy in Optisol-GS and tissue. Corneoscleral rims contaminated with Candida albicans (C.albicans) were incubated in Optisol-GS, either without antifungal agents or with the addition of amphotericinB or voriconazole at various concentrations (2 ×, 5 ×, 10 ×, and 20 × MIC), at different time points, and under various preservation temperatures (2-8°C versus 2h-room temperature exposure). Antifungal efficacy was evaluated by counting viable yeast colonies cultured from Optisol-GS samples. Tissue sterility was determined through direct tissue culture and histological examination of the contaminated rims after a 14-day incubation period. Room temperature exposure did not increase colony growth at the same multiple MIC of antifungal agents. Although antifungal addition reduced C.albicans growth in a concentration-dependent manner, yeast growth was still observed in all Optisol-GS samples with a single supplementation after a 14-day incubation. Only groups with additional antifungal supplementation on either day2 or day6 showed a 99% or greater reduction of C.albicans growth in Optisol-GS samples and yielded negative results in direct tissue culture. The eye bank warming protocol did not compromise antifungal efficacy. To sustain the required concentration and effectively reduce C.albicans growth in Optisol-GS and contaminated tissue, additional antifungal supplementation on either day2 or day6 was necessary during a 2-week preservation period.

Antimicrobial activity of cell-free supernatant derived from Ligilactobacillus animalis SWLA-1 in a novel ex vivo canine corneal infection model.

Canine bacterial keratitis is a corneal infection that causes various symptoms, including visual impairment, and necessitates eye removal in severe cases. Staphylococcus pseudintermedius is a pathogen that causes significant bacterial keratitis in canine patients. Moreover, multi-drug resistant Staphylococcus pseudintermedius (MDRSP) has been reported in both humans and animals. Regarding treatment failure against multi-drug resistant (MDR) pathogens with classic antibiotics, antimicrobial compounds derived from probiotics have been suggested as an alternative approach. Ligilactobacillus animalis SWLA-1 strain and its cell-free supernatant (CFS) have previously demonstrated potent antimicrobial activity against various MDR pathogenic bacteria. Based on this finding, we evaluated the anti-staphylococcal activity of CFS derived from Ligilactobacillus animalis SWLA-1 against MDRSP in a newly established ex vivo canine corneal infection model using fresh canine corneoscleral rims. Additionally, an in vitro cytotoxicity test using human keratocytes was performed. CFS significantly inhibited the growth of MDRSP in the novel ex vivo model and did not exhibit any significant toxicity against keratocytes in vitro. Based on these results, the antimicrobial compounds in CFS show potential as a novel approach for MDR staphylococcal keratitis treatment.

Use of discarded corneo-scleral rims to create cornea-like tissue.

Corneal disease is a major cause of blindness. Transplantation of cadaver-derived corneas (keratoplasty) is still the current therapy of choice; however, the global shortage of donor corneas continues to drive a search for alternatives. To this end, biosynthetic corneal substitutes have recently begun to gain importance. Here, we present a novel method for the generation of a cornea-like tissue (CLT), using corneo-scleral rims discarded after keratoplasty. Type I collagen was polymerized within the corneo-scleral rim, which functioned as a 'host' mould, directing the 'guest' collagen to polymerize into disc-shaped cornea-like material (CLM), displaying the shape, curvature, thickness, and transparency of normal cornea. This polymerization of collagen appears to derive from some morphogenetic influence exerted by the corneo-scleral rim. Once the CLM had formed naturally, we used collagen crosslinking to fortify it, and then introduced cells to generate a stratified epithelial layer to create cornea-like tissue (CLT) displaying characteristics of native cornea. Through the excision and reuse of rims, each rim turned out to be useful for the generation of multiple cornea-shaped CLTs. The approach effectively helps to shorten the gap between demand and supply of CLMs/CLTs for transplantation. We are exploring the surgical transplantation of this CLT into animal eyes, as keratoprostheses, as a precursor to future applications involving human eyes. It is possible to use either the CLM or CLT, for patients with varying corneal blinding diseases.

Risk Factors of Corneal Donor Contamination and Post-Keratoplasty Infectious Complication

Introduction &amp; Objectives : Infectious keratitis and endophthalmitis are rare yet devastating complications of corneal transplantation. Some studies have demonstrated increased incidence of infectious post-keratoplasty complications are related to contaminated corneal donor, although its risk factors remain controversial. This study provides concise review on possible risk factors of corneal donor contamination which related to post-keratoplasty infectious complications.&#x0D; Methods : This literature review was conducted by searching PubMed and Proquest for “corneal donor contamination” AND (“infectious complication” OR infection OR endophthalmitis) from the period of 2018-2023. Relevant articles were reviewed and selected for further understanding regarding various risk factors of corneal donor contamination leading to post-keratoplasty infectious complications and its preventive strategies.&#x0D; Results : The rate of corneal contamination varied between studies, ranging from 1,3 to 7,8%. The most common risks of contamination are donor’s diseases (e.g. cardiovascular, cerebrovascular accident) and prolonged time between death and donor retrieval. Longer preservation time (&gt;5 days), environmental situation (temperature and humidity), and prolonged treatment duration on intensive care unit (&gt;4 days) are also considered as its risk factors. Corneoscleral rim culture, addition of antifungal and antibacterial to corneal storage medium, and shortening of corneal processing time should be considered as probable preventive strategies.&#x0D; Conclusion : Donor diseases, prolonged donor retrieval time from death, and longer donor preservation and hospitalization duration, are associated with higher rates of corneal donor contamination. Rapid retrieval and donor handling time, together with corneoscleral rims culture, at least for fungus regarding its better predictive values, should be taken into consideration by corneal donors providing eye banks.

Preparation and outcome of swollen versus dextran deswollen grafts in descemet membrane endothelial keratoplasty (DMEK)

Aims/Purpose: To evaluate the influence of organ culture medium containing dextran on graft characteristics during preparation as well as the outcome of subsequent Descemet Membrane Endothelial Keratoplasty (DMEK).Methods: 31 corneoscleral rims of the Eye Bank of the University Hospital of Cologne with a donor age between 52 and 74 years were examined between 2018 and 2022 for their intraoperative preparation conditions and the outcome after performing 31 DMEKs. For this purpose, corneoscleral rims were randomized into two organ‐cultured groups, one immersed in culture medium with dextran (group A, n = 16) and the other in medium without dextran (group B, n = 15). The intraoperative graft preparation conditions were noted by using a standardized scoresheet, including splitting and stripping behaviour, central and peripheral attachment as well as graft fragility. The post‐operative outcome parameters were defined as Best Spectacle‐Corrected Visual Acuity (BSCVA), central and peripheral corneal thickness (CCT and PCT), endothelial cell count (ECC) and re‐bubbling rate.Results: In both groups, all 31 DM‐endothelium grafts were usable for DMEK. There was no significant difference in the intraoperative preparation conditions of swollen versus dextran deswollen DM‐endothelium grafts in respect to stripping (p = 0.722) and to Descemet membrane splitting behaviour (p = 0.013). The fragility of the grafts showed no significant difference (p = 0.433), as well as the peripheral scars indicating a pseudophakic donor (p = 1.000) and the tendency for central tearing (p = 0.484) during preparation. No significant difference in the outcome 12 months after DMEK in conditions of swollen versus dextran deswollen DM‐endothelium grafts regrading CCT (p = 0.189), re‐bubbling (p = 0.704), PCT (p = 0.063), could be observed. BSCVA was significantly better in group B 12 months after DMEK with p = 0.032 (group A 0.08 ± 0.38: group B 0.03 ± 0.13).Conclusions: This pilot study does not show relevant and significant differences between swollen versus dextran deswollen grafts in DMEK transplantation. Future prospective studies with higher patient numbers are needed. Our study suggests also non‐deswollen donors as suitable for DMEK surgery.

Gene expression in calcium triggered healthy primary limabal epithelial cells, in vitro

Aims/Purpose: Calcium supplementation can induce differentiation of several cell types. The tear film has an important role in maintenance of the ocular surface homeostasis with an ionic calcium concentration of 0.4 mM‐1.0 mM. The effect of calcium concentration on healthy primary limbal epithelial cells (LECs) has not been analysed, yet. Our purpose was to investigate the effect of calcium supplementation on gene expression of primary limbal epithelial cells (LECs) of healthy donor corneas, in vitro.Methods: Primary LECs were isolated from corneoscleral rims of healthy donors (n = 6) and were cultured in serum free low calcium (0.06 mM Ca+2) Keratocyte growth medium 3 (KGM3). The primary LECs were then supplemented by 0.3 mM‐2.0 mM Ca+2 concentrations for 72 h. Control LEC cultures were supplemented by low calcium (0.06 mM Ca+2) KGM3 medium. After cell harvesting, RNA and protein were extracted from treated and control LECs to perform qPCR and western blot analysis.Results: The calcium supplementation had no significant effect on PAX6 mRNA expression after 72 h. The differentiation marker and adhesion protein DSG1 and the cornea specific keratin K3 expression did not change significantly through Ca+2 supplementation (p ≥ 0.5). The retinoic acid signalling component fatty acid binding protein FABP5 expression (p ≥ 0.5) the wound healing and vascularization marker CAV1 gene expression (p ≥ 0.5) did not change significantly through Ca+2 supplementation, after 72 h.Conclusions: Calcium supplementation for 72 h did not change PAX6 expression and did not trigger differentiation of healthy primary limbal epithelial cells. We plan to analyse the effect of longer incubation time in the future in order to trigger a full differentiated status of limbal epithelial cells, using calcium supplementation.

Performance outcomes from a DMEK peeling and preparation wet lab

ObjectiveTo evaluate the Descemet membrane endothelial keratoplasty (DMEK) preparation performance of trainee surgeons in an ex vivo human donor cornea DMEK wet lab simulation setting.MethodsHuman donor corneoscleral rims unsuitable for...

Impact of cornea recovery training and certification on ophthalmology residents

BackgroundIn Asian countries, such as Taiwan, social taboos regarding organ and tissue donation decreases the prevalence of organ and tissue transplants. This also applies to cornea recovery, which is a skill that requires precision and practice to perform well. In Taiwan, to ensure the maintenance of high-quality corneas, a comprehensive training program and certified examination has been implemented. This study aims to investigate the impact of these programs and examinations on cornea recovery. MethodsResearchers evaluated the efficiency of the training and certified examination process by comparing the corneoscleral rim width, Descemet's membrane folds, endothelial layer stress lines, and endothelial cell density performed by ophthalmology residents in 2018 and 2019. ResultsAfter training and certification, the Descemet's membrane folds rate decreased from 14.3 % to 2.0 % and endothelial layer stress lines rate decreased from 22.5 % to 5.0 %. The endothelial cell density of donor grafts significantly improved from 2681.9 cells/mm2 to 2869.7 cells/mm2 (p < 0.001). ConclusionsThis study used objective data to evaluate cornea recovery quality after training and certification. The training and certified examination significantly improved the surgical skills of ophthalmology residents and could be applied in other tissue or organ recovery procedures to maintain and improve quality.

Prevalence of Herpes Simplex and Varicella-Zoster Virus DNA in Corneal Grafts Is Higher than Expected.

(1) To determine the prevalence of herpes simplex virus type 1 (HSV-1) and type 2 (HSV-2), varicella-zoster virus (VZV), and cytomegalovirus (CMV) DNA in donor corneas; (2) To evaluate the clinical outcome of the grafts with viral DNA and to compare donors with and without viral DNA. We analyzed data from all donors and recipients who underwent penetrating keratoplasty (PK) or Descemet membrane endothelial keratoplasty (DMEK) between September 2022 and March 2023. Donor corneoscleral rims and excised recipients' corneal buttons were tested for the presence of HSV-1, HSV-2, VZV, and CMV DNA by polymerase chain reaction (PCR). The results were known 2-3 days after the surgery. We closely followed up on patients whose grafts tested positive for viral DNA. We compared the medical histories of donors with and without viral DNA. We included 85 corneas from 67 donors. Seven (8.2%) donor corneas tested positive for HSV-1 (n = 3) or VZV (n = 4) DNA. We did not detect any HSV-2 or CMV DNA. In the postoperative follow-up of patients with positive PCR, a graft failure was observed in one and infections in two eyes. Re-operation was necessary in three of these cases (42.9%). Patients without herpes DNA in the donor cornea needed reoperation in 7.7% of the cases. Cultural duration, the cause of the donor's death, and the death-to-explantation interval did not differ significantly between donors with and without viral DNA. Additionally, 3 of the 7 (42.9%) donors with positive PCR were in a septic status at the time of death, compared to 21 of the 78 (26.9%) donors with negative PCR (p = 0.52). The prevalence of herpes DNA in the donor corneas was 8.2% and thus higher than previously reported. We did not notice any evidence for a donor-to-host transmission, but a higher rate of postoperative complications in recipients of the grafts with viral DNA. The donors with and without herpetic DNA did not differ significantly regarding systemic diagnoses or cultural conditions, but sepsis was more frequent in the group with viral DNA.

Manufacturing of human corneal endothelial grafts

PurposeHuman corneal endothelial cells (HCECs) play a significant role in maintaining visual function. However, these cells are notorious for their limited proliferative capacity in vivo. Current treatment of corneal endothelial dysfunction resorts to corneal transplantation. Herein we describe an ex vivo engineering method to manufacture HCEC grafts suitable for transplantation through reprogramming into neural crest progenitors. MethodsHCECs were isolated by collagenase A from stripped Descemet membrane of cadaveric corneoscleral rims, and induced reprogramming via knockdown with p120 and Kaiso siRNAs on collagen IV-coated atelocollagen. Engineered HCEC grafts were released after assessing their identity, potency, viability, purity and sterility. Phase contrast was used for monitoring cell shape, graft size, and cell density. Immunostaining was used to determine the normal HCEC phenotype with expression of N-cadherin, ZO-1, ATPase, acetyl-α-tubulin, γ-tubulin, p75NTR, α-catenin, β-catenin, and F-actin. Stability of manufactured HCEC graft was evaluated after transit and storage for up to 3 weeks. The pump function of HCEC grafts was measured by lactate efflux. ResultsOne HCEC graft suitable for corneal transplantation was generated from 1/8th of the donor corneoscleral rim with normal hexagonal cell shape, density, and phenotype. The manufactured grafts were stable for up to 3 weeks at 37 °C or up to 1 week at 22 °C in MESCM medium and after transcontinental shipping at room temperature by retaining normal morphology (hexagonal, >2000 cells/mm2, >8 mm diameter), phenotype, and pump function. ConclusionsThis regenerative strategy through knockdown with p120 and Kaiso siRNAs can be used to manufacture HCEC grafts with normal phenotype, morphology and pump function following prolonged storage and shipping.

Expansion and characterization of human limbus-derived stromal/mesenchymal stem cells in xeno-free medium for therapeutic applications

BackgroundMesenchymal stem cells (MSCs) have been proven to prevent and clear corneal scarring and limbal stem cell deficiency. However, using animal-derived serum in a culture medium raises the ethical and regulatory bar. This study aims to expand and characterize human limbus-derived stromal/mesenchymal stem cells (hLMSCs) for the first time in vitro in the xeno-free medium.MethodsLimbal tissue was obtained from therapeutic grade corneoscleral rims and subjected to explant culture till tertiary passage in media with and without serum (STEM MACS XF; SM), to obtain pure hLMSCs. Population doubling time, cell proliferation, expression of phenotypic markers, tri-lineage differentiation, colony-forming potential and gene expression analysis were carried out to assess the retention of phenotypic and genotypic characteristics of hLMSCs.ResultsThe serum-free medium supported the growth of hLMSCs, retaining similar morphology but a significantly lower doubling time of 23 h (*p < 0.01) compared to the control medium. FACS analysis demonstrated ≥ 90% hLMSCs were positive for CD90+, CD73+, CD105+, and ≤ 6% were positive for CD45−, CD34−andHLA-DR−. Immunofluorescence analysis confirmed similar expression of Pax6+, COLIV+, ABCG2+, ABCB5+, VIM+, CD90+, CD105+, CD73+, HLA-DR−andCD45−, αSMA− in both the media. Tri-lineage differentiation potential and gene expression of hLMSCs were retained similarly to that of the control medium.ConclusionThe findings of this study demonstrate successful isolation, characterization and culture optimization of hLMSCs for the first time in vitro in a serum-free environment. This will help in the future pre-clinical and clinical applications of MSCs in translational research.

Modulation of mucin secretion using combined polyethylene glycol-propylene glycol topical formulation in a hyperosmotic stress-based explant model.

Ocular surface discomfort and dry eye disease are caused by a dysfunctional tear film. The efficacy of lubricating eye drops on the human eye is known, but the compositions may show differential effects on rescuing the tear film. Mucins form a critical layer of the tear film, a reduction of which may be causative for ocular surface conditions. Therefore, it is essential to develop relevant human-derived models to test mucin production. Human corneoscleral rims were obtained from a healthy donor (n = 8) post-corneal keratoplasty and cultured in DMEM/F12 media. Hyperosmolar stress mimicking dry eye disease was induced by exposing the corneoscleral rim tissues to +200 mOsml NaCl-containing media. The corneoscleral rims were treated with polyethylene glycol-propylene glycol (PEG-PG)-based topical formulation. Gene expression analysis was performed for NFAT5, MUC5AC, and MUC16. Secreted mucins were measured by enzyme-linked immunosorbent assay (ELISA) (Elabscience, Houston, TX, USA) for MUC5AC and MUC16. The corneoscleral rims responded to hyperosmolar stress by upregulating NFAT5, a marker for increased osmolarity, as observed in the case of dry eye disease. The expression of MUC5AC and MUC16 was reduced upon an increase in hyperosmotic stress. The corneoscleral rim tissues showed induction of MUC5AC and MUC16 expression upon treatment with PEG-PG topical formulation but did not show significant changes in the presence of hyperosmolar treatments. Our findings showed that PEG-PG-based topical formulation slightly alleviated hyperosmolar stress-induced decrease in MUC5AC and MUC16 gene expression that is encountered in DED.

The Prevalence of Positive Donor Corneoscleral Rim Culture and its Association with Ocular Infection After Transplantation.

The aim of the study was to determine the prevalence of positive corneoscleral donor rim cultures and to report keratitis and endophthalmitis after keratoplasty. Eye bank records and medical records of patients who underwent keratoplasty between September 1, 2015, and December 31, 2019, were retrospectively reviewed. Patients who had routine donor-rim culture taken during surgery and followed up for at least 1 year in the post-operative period were included in the study. A total of 826 keratoplasty procedures were performed. A total of 120 (14.5%) cases had a positive donor corneoscleral rim culture. Positive bacterial cultures were obtained from 108 (13.7%) of the donors. Bacterial keratitis was observed in one patient (0.83% of recipients) who had a positive bacterial culture. Positive fungal cultures were obtained from 12 (1.45%) donors, of whom one (8.33% of recipients) developed fungal keratitis. Endophthalmitis was observed in one patient whose culture result was negative. Both bacterial and fungal culture results were similar in penetrating and lamellar surgical procedures. Although the donor corneoscleral rims have a high positive culture result, the rate of bacterial keratitis and endophthalmitis is low, the risk of infection is high in patients with a fungal positive donor rim. Closer follow-up of patients with fungal positive donor corneo-scleral rim result and initiation of aggressive antifungal treatment when infection occurs will be beneficial.

Maximize Donor Cornea Use in a Hepatitis B Endemic Area via Serology Matching.

This study aims to investigate the rationality of the allocation guidelines in a hepatitis B endemic area that uses corneas from hepatitis B donors. Under Taiwan's current guidelines, corneas donated from hepatitis B surface antigen (HBsAg)(+) donors can be allocated to HBsAg(+) or hepatitis B surface antibody recipients. From January 1, 2015, to December 31, 2019, corneas donated to National Taiwan University Hospital were divided into HBsAg(+), HBsAg(-)/hepatitis B core antibody (anti-HBc)(+), and HBsAg(-)/anti-HBc(-) groups. Hepatitis B virus (HBV) DNA extracted from corneoscleral rims was quantified by polymerase chain reaction and correlated with donor serum HBsAg, anti-HBc, and HBV DNA. Recipients of HBV DNA(+) grafts were called back for serology and serum HBV DNA tests. The corneoscleral HBV DNA of 170 corneas (113 donors) was quantified, of which 45 corneas were from 28 HBsAg(+) donors, 87 were from 57 HBsAg(-)/anti-HBc(+) donors, and 38 were from 28 HBsAg(-)/anti-HBc(-) donors, and HBV DNA was detected in 80.0%, 9.2%, and 0% of the corneoscleral rims in each group. Donor anti-HBc(+) provided the highest sensitivity (1.00) and negative predictive value (1.00) for corneoscleral HBV DNA. Twenty-eight of 40 recipients (70%) using HBV DNA(+) grafts were called back, and none developed hepatitis in follow-up periods ranging from 6 to 55.5 mo. Donor anti-HBc should be tested routinely with HBsAg. Allocating corneas from HBsAg(+) or anti-HBc(+) donors to HBsAg(+) or hepatitis B surface antibody recipients maximizes cornea usage from hepatitis B donors without compromising transplant safety in a hepatitis B endemic setting.

Gonioscopy and Angle-Based Glaucoma Surgical Training on Human Eyes in the Wet Lab.

Human corneoscleral rims provide a low cost model and resource to enhance angle based glaucoma surgical training skills. To understand the feasibility, utility, and efficacy of a low-cost model for performing anterior chamber angle gonioscopy and micro-surgery on human corneoscleral tissues in wet lab on resident skill enhancement. Post-keratoplasty residual corneoscleral rims and/or the expired donor corneoscleral buttons were included for the purpose of this study. Initially, inverted rims were used to demonstrate the basic angle anatomy on a slit lamp bio-microscope. Central hazy cornea was then replaced with an artificial cornea and an artificial anterior chamber to simulate indirect gonioscopy, direct gonioscopy, goniotomy, and other angle-based surgeries. Direct gonioscopy and goniotomy exercises (as a means to simulate intraoperative gonioscopy and other angle-based surgeries respectively) were evaluated quantitatively, and the rest qualitatively. A total of 65 residents were included in the exercise. The mean age was 26.69±1.74 years. Amongst these, 55 ophthalmology residents performed the basic examination, that is, slit lamp mounted angle anatomy examination (n=55) and artificial chamber mounted indirect gonioscopic examination through artificial cornea (n=55). In the second exercise, 10 glaucoma fellows performed the direct gonioscopic and the goniotomy exercises on artificial chamber. The fellows demonstrated a statistically significant improvement in interpretation and surgical skill enhancement concurrent with reduction in time taken with increasing number of attempts ( P <0.05). In the third simulation exercise, micro-stent placement within the Schlemm's canal, subconjunctival space, and the suprachoroidal space was re-produced successfully on multiple occasions. The residual human corneoscleral rims proved to be an important resource for teaching both the basic examination skills and those required for transitioning to advanced intraoperative gonioscopy and goniotomy skills. These low cost, easy to adapt models can be incorporated within the curriculum for improved glaucoma training of both the residents and the glaucoma fellows.

Incidence and Outcomes of Transplant of Infected Donor Corneal Tissues in a Tertiary Hospital in Saudi Arabia.

Purpose: The aim of this study is to assess donor rim culture results and predict ocular infections after corneal transplantation to determine the relationship between positive corneoscleral rim cultures and post-keratoplasty infection.Design: This is a retrospective study.Methods: The microbiology results of positive donor rim culture and clinical outcomes of all contaminated grafts at Dhahran Eye Specialist Hospital (DESH) in the Eastern Region of Saudi Arabia between October 2016 and February 2020 were reviewed.Results: A total of 684 corneal transplantation procedures were performed at DESH during this period. Routine donor rim cultures were done for all cases with six positive cases (0.88%). There were five positive fungal donor rim cultures (0.73%), and one corneal rim was positive for both bacterial and fungal cultures (0.15%). Among these six patients, two patients (33.33%) who received graft positive with candida developed an infection. Both cases required further interventions but were unsuccessful and eventually ended up with re-grafting.Conclusion: In the sample collected, positive fungal contamination was more frequent than bacterial contamination, and the incidence of bacterial infection is relatively low compared to that of fungal infection due to contaminated grafts. Despite the limitations of our study, we support routine corneoscleral rim culture and to start prophylactic antifungal treatment in response to positive donor fungal rim culture.

Analysis in Results of Microbiologic Exam Related to Donor Corneas

Purpose: This study analyzed the microorganisms identified in donor corneas and their clinical significance.Methods: The medical records of 94 patients (114 eyes) who underwent keratoplasty and microbiological tests of the donor corneas from October 2008 to December 2020 at our hospital were reviewed retrospectively. During keratoplasty, we conducted microbiological tests of the corneoscleral rim of the donor cornea and preserving solution Optisol™-GS (Bausch &amp; Lomb, Rochester, NY, USA), and examined the antibiotic susceptibility of bacterial isolates from the cultures.Results: Some isolates of domestic donor corneas revealed bacteria, but none of the imported corneas did. Gram-negative bacilli were detected from the corneoscleral rim in three eyes (2.6%): two cases of Acinetobacter baumannii/haemolyticus and one case of Pseudomonas aeruginosa. In one case (0.9%), Acinetobacter baumannii/haemolyticus was identified from preserving solution, with no bacteria found in the corneoscleral rim. Antibiotic susceptibility tests showed multi-drug resistance, except to colistin. In all cases where bacteria were detected, there was no keratitis or endophthalmitis after corneal transplantation.Conclusions: Although rare, bacteria can be identified from donor corneas or the preserving solution used in corneal transplantation. Therefore, caution is needed in all processes dealing with donor corneas. Empirical antibiotics that have sufficient antimicrobial activity to suppress multi-drug resistant bacteria should be selected in corneal transplantation.

Endophthalmitis secondary to donor to host infection in posterior lamellar keratoplasties: A systematic review

Donor to host transmission of infectious agents is a well-recognized entity, more commonly related to Endothelial Keratoplasty (EK) than to Penetrating Keratoplasty (PK), that involves complications ranging from interface keratitis to endophthalmitis. A systematic review of the literature was conducted to identify the published articles until December 2020 reporting cases of endophthalmitis secondary to corneal graft contamination in posterior lamellar keratoplasties (DSAEK and DMEK) along with donor characteristics, microbiological profile, treatment and outcomes. Each identified article was assessed to meet donor to host infection criteria, defined as a post-procedural infection in which the same microbiological agent was identified in both the donor corneoscleral rim or preservation medium and receptor ocular tissue. From 23 research articles, eight reports of endophthalmitis in nine patients following DSAEK or DMEK secondary to donor to host infection fulfilled the inclusion criteria. The majority were male and the median age was 72.0 (45.0-81.0) years old. Indications of surgery were primarily pseudophakic bullous keratopathy and Fuchs dystrophy. A fungal pathogen was identified in eight of nine patients. All the cases underwent surgical management with lenticule removal or endothelial plaque aspiration. The final corrected distance visual acuity (CDVA) in all cases was 20/200 or better. Endophthalmitis after an EK procedure is a rare complication whose outcome depends on the aggressive and precocious treatment. Identification of early signs of interface keratitis and lenticule removal seems mandatory in patients undergoing DSAEK or DMEK to prevent further involvement of the globe.

A unique pre-endothelial layer at the posterior peripheral cornea: ultrastructural study

This study was conducted to investigate the ultrastructure of a unique structures at the anterior side of the endothelium of the posterior peripheral cornea and compare their inner fibers to those of the limbus and sclera. The unique structures at the anterior side of endothelium was referred as a pre-endothelial (PENL) structures in the present manuscript. Ten anonymous-donor human corneoscleral rims (leftover after corneal transplants) were processed for electron microscopy. Semi-thin sections were examined using an Olympus BX53 microscope, and ultrathin sections were studied using a JOEL 1400 transmission electron microscope. A unique PENL structures was identified at the posterior peripheral cornea at a radial distance of approximately 70–638 µm, from the endpoint of Descemet’s membrane. The PENL thinned out gradually and disappeared in the center. The contained an electron-dense sheath with periodic structures (narrow-spacing fibers), wide-spacing fibers, and numerous microfibrils. Typical elastic fibers were present in the sclera and limbus but were not observed in the PENL. This study revealed the existence of a new acellular PENL, containing unique fibrillar structures that were unseen in the corneal stroma. From the evidence describe in this paper we therefore suggest that PENL is a distinct morphological structure present at the corneal periphery.