Long-Term Results of Sterile Corneal Allograft Ring Segments Implantation in Keratoconus Treatment.

PURPOSE: To evaluate the efficacy and safety of sterile corneal allograft ring segments implantation for the treatment of keratoconus by analyzing long-term visual, refractive, and tomographic clinical outcomes. METHODS: This prospective study included 62 eyes of 49 patients with keratoconus who underwent corneal allograft ring segments implantation at Istanbul Medipol University Faculty of Medicine between February 2020 and August 2022. Surgical outcomes using the Istanbul nomogram were evaluated in patients preoperatively and postoperatively at 1 month, 6 months, 1 year, and 3 years. Outcomes measured were uncorrected distance visual acuity (UDVA), corrected distance visual acuity (CDVA), spherical equivalent (SE), spherical refraction (SR), cylindrical refraction (CR), topographic keratometric values, and corneal thickness at the thinnest point. RESULTS: Preoperative mean UDVA and CDVA (LogMAR) were 0.96 ± 0.50 and 0.72 ± 0.47, respectively, and increased to 0.41 ± 0.34 and 0.22 ± 0.19 at the last visit (P < 0.001). There was a significant decrease in SE, SR, and keratometric values postoperatively (P < 0.001). There was no difference in CR and thinnest corneal thickness values (P = 0.333 and 0.154, respectively). The stromal and epithelial thicknesses measured by anterior segment optical coherence tomography were stabilized at 6 months and 1 year, respectively. No major complications or side effects were observed intraoperatively or postoperatively. CONCLUSIONS: This study demonstrated that sterile corneal allograft ring segments implantation is a safe and feasible treatment for keratoconus, yielding notable long-term visual outcomes with minimal implant-related complications.

Prospective In Vitro Comparison of Kerasave and Optisol-GS Corneal Storage Solutions.

PURPOSE: The aim of this study was to compare the performance of Kerasave and Optisol-GS for hypothermic corneal storage for 14 days. METHODS: This study was a prospective laboratory investigation. Mate corneas were recovered into Kerasave or Optisol-GS (27 pairs) and stored at 2°C to 8°C for 14 days. Corneas were evaluated by trained eye bank technicians, and study parameters were compared between the initial and final evaluations. Endothelial cell density (ECD), hexagonality (HEX), and coefficient of variation (CV) were evaluated by specular microscopy, and central corneal thickness (CCT) was examined by optical coherence tomography after 1, 3, 7, and 14 days of storage. Corneal transparency was scored using slit lamp examination at days 1 and 14. RESULTS: Average ECD, HEX, and CV for the Kerasave (2653 ± 303 cells/mm 2 , 57 ± 4%, and 36 ± 3%) and Optisol-GS (2623 ± 306 cells/mm 2 , 57 ± 5%, and 36 ± 4%) groups were not significantly different at day 1. There was also no difference at any other study time points (all P > 0.05). ECD did not significantly change from day 1 to day 14 in either group ( P > 0.05), but a statistically significant change in HEX and CV was observed between day 1 and day 14 in both groups ( P < 0.01). Average CCT measured at day 1 for corneas stored in Kerasave was 622 ± 49 µm and those stored in Optisol-GS was 580 ± 35 µm ( P < 0.01). The difference in CCT measurements was not significantly different at day 14 (Kerasave: 674 ± 46 µm vs. Optisol-GS: 647 ± 58 µm, P > 0.05). Corneal transparency was not significantly different between the 2 groups at day 1 or day 14. CONCLUSIONS: The corneal quality and clinically relevant parameters including ECD, endothelial morphometry, and corneal transparency were not different in corneas stored in Kerasave or Optisol-GS for 14 days. The initial difference in CCT between the 2 groups decreased at day 14. These results demonstrated that Kerasave corneal storage solution preserves the corneal endothelium similarly to Optisol-GS.

Double-Scroll Formation by Fluid Column Manipulation in Preloaded DMEK Grafts Prepared From Younger and Older Donor Tissue.

PURPOSE: The purpose of this study was to determine whether manipulation of preloaded single-scroll Descemet membrane endothelial keratoplasty (DMEK) grafts within the fluid column of an injector can safely and reliably result in formation of double-scroll DMEK grafts and whether there are differential effects on younger versus older donor tissue. METHODS: Pairs of DMEK grafts prepared from older (65-80 years) and younger (48-64 years) donors were preloaded into a Straiko modified Jones tube. One member of the pair was manipulated within the fluid column to form a double-scroll graft, and the other remained unmanipulated. Outcomes measured include success rate for double-scroll formation, endothelial cell loss (ECL), and relative scroll width. RESULTS: Older donor grafts formed double scrolls with a 100% success rate. ECL of older donor manipulated grafts was statistically higher than that of unmanipulated mate grafts (17.4% ± 3.5% vs. 13.0% ± 4.2%, P = 0.03), but was still within the acceptable range for transplant. Younger donor grafts were successfully manipulated into double scrolls with a 67% success rate, and there was no difference in the ECL of manipulated and unmanipulated grafts (15.5% ± 4.4% vs. 13.0% ± 4.5%, P = 0.24). For all grafts and conformations, there was a significant relationship between relative scroll width and ECL ( P < 0.01). CONCLUSIONS: Fluid column manipulation can be used reliably to form double-scroll DMEK grafts. For younger donor grafts, manipulation yields a double scroll without increasing ECL. For older donor grafts, manipulation results in a minimal, acceptable increase in ECL. Surgeons should weigh the advantage of an easily opened graft against the risk of increased ECL when considering this technique.

Allograft corneal ring segment for keratoconus management: Istanbul nomogram clinical results.

PURPOSE: To evaluate the clinical feasibility and visual outcomes of allograft corneal ring segment implantation for the treatment of keratoconus. METHODS: This case series, included forty-four eyes of 32 patients with a 6-month follow-up. All cases were treated according to the Istanbul nomogram. In the Istanbul Nomogram, corneal tunnels of 4 × 7.5 mm diameters are created at depth of 200 µm and implanted with sterile allograft corneal rings (KeraNaturalTM, Lions VisionGift, Portland, OR, USA) at the cone location. Uncorrected distance visual acuity (UDVA), corrected distance visual acuity (CDVA), spherical equivalent (SE) and keratometric values were compared preoperatively versus postoperatively. RESULTS: There was significant improvement in UDVA, CDVA, SE and topographic keratometric values. The mean preoperative CDVA (Snellen, decimal) increased from 0.29 ± 0.20, to 0.56 ± 0.26 (P < 0.001), at the last visit. There was no statistically significant difference between preoperative and postoperative thinnest pachymetry values (P = 0.509). No major complications or adverse event were observed during and after the operation. CONCLUSIONS: The results of this pilot study show that sterile allograft corneal ring segments may be safe, effective and enhance the visual performance of keratoconus patients. Larger clinical studies are needed to demonstrate the effectiveness and safety with long term follow-up.

Examination of a Modified Graft Preparation Technique to Induce Double-Scroll Formation and Promote the Use of Younger Descemet Membrane Endothelial Keratoplasty Donor Tissue.

PURPOSE: The purpose of this study was to determine whether controlled balanced salt solution (BSS) bursts during graft preparation can safely promote formation of a double-scrolled Descemet membrane endothelial keratoplasty (DMEK) graft in younger donor tissue. METHODS: DMEK grafts prepared from young donor tissue (average age, 55 years; range, 39-66 years) were floated in BSS to spontaneously form scrolls (N = 10 pairs). Controlled BSS bursts were used to promote double-scroll (DS) formation in 1 member of each pair. Grafts were stained, preloaded, and shipped before cell viability analysis. After appropriate training, a less experienced technician performed this technique on 10 additional corneas. Outcomes measured for both technicians include the success rate for obtaining a DS, scroll conformation after shipping, and endothelial cell loss (ECL). RESULTS: There was no difference in ECL between grafts subjected to additional manipulation compared with unmanipulated mate grafts (observer 1: 15.2% ± 3.3% vs. 15.2% ± 4.4%, P = 0.99; observer 2: 16.3% ± 2.9% vs. 15.9% ± 4.5%, P = 0.8). A technician experienced with this technique had a 90% success rate, whereas a less experienced technician had a 70% success rate. The mean ECL of the 10 grafts manipulated by the less experienced technician was not significantly different from results obtained from the experienced technician (observer 1: 18.5% ± 6.0% vs. 15.2% ± 3.3%, P = 0.15; observer 2: 18.1% ± 5.6% vs. 16.3% ± 2.9%, P = 0.34). Scrolls maintained their conformation during shipping events. CONCLUSIONS: Double-scroll graft formation using controlled BSS bursts is a reliable technique that can be performed without causing additional damage to DMEK grafts. This technique may make graft unscrolling easier and can promote the use of younger donor tissue for DMEK.

Prevalence, Anatomic Distribution, and Correlates of Chlamydia trachomatis and Neisseria gonorrhoeae Infections Among a Cohort of Men Who Have Sex With Men in Hanoi, Vietnam.

BACKGROUND: Neisseria gonorrhoeae (NG) and Chlamydia trachomatis (CT) disproportionately affect men who have sex with men (MSM). Data on the prevalence, anatomical distribution, and correlates of NG and CT infections among MSM in Vietnam are limited. METHODS: Between July 2017 and April 2019, MSM 16 years or older without HIV were enrolled into an observational cohort study. Baseline data, including sociodemographics, sexual behavior, and HIV status, were collected. Testing for NG and CT were performed on urine, rectal, and pharyngeal specimens. Multivariate logistic regression models identified factors associated with NG and CT infections at baseline. RESULTS: In total, 1489 participants underwent NG/CT testing. The median age was 22 years (interquartile range, 20-26 years). There were 424 (28.5%) NG or CT infections: 322 (21.6%) with CT and 173 (11.6%) with NG. Rectal infections were most common for CT (73.9%), whereas pharyngeal infections were most common for NG (70.5%). Independent risk factors for CT or NG infection included ≥2 sex partners in the prior month (adjusted odds ratio [aOR], 2.04; 95% confidence interval [CI], 1.44-2.91), condomless anal sex (aOR, 1.44; 95% CI, 1.12-1.86), and meeting sex partners online (aOR, 1.35; 95% CI, 1.03-1.76). Recent genitourinary or rectal symptoms were not associated with infections. CONCLUSIONS: The overall and extragenital prevalences of NG and CT infections were high within this sample of young MSM without HIV in Hanoi. Testing limited to urethral specimens would have missed nearly three-quarters of CT and NG infections, supporting the need for routine testing at multiple anatomic sites.

Modified procedure for Loading "Flat" DMEK Grafts Into an Injector.

PURPOSE: The aim of this study was to determine whether loading a Descemet membrane endothelial keratoplasty (DMEK) graft using a drop-in procedure results in more endothelial cell loss (ECL) than the standard suction procedure. METHODS: Pairs of donor corneas with equivalent preprocessing endothelium were prepared using the standard protocol of our eye bank. One member of each pair was loaded into an injector using the standard suction protocol. The mate graft was loaded using a drop-in protocol, in which the edge of the graft was gently grasped with a forceps, lifted to the edge of the injector, and dropped inside. Grafts were evaluated for ECL and examined for grab marks or other loading-associated damage. RESULTS: There was no difference in mean ECL of grafts prepared for DMEK using the standard protocol (20.6% ± 4.5%) compared with that of mate grafts prepared using the drop-in loading protocol (19.5% ± 4.8%, P = 0.59). There was no consistent pattern of damage in the drop-in-loaded grafts, as grab marks or other tissue damage associated with the drop-in loading protocol were not consistently identified by a trained corneal surgeon. CONCLUSIONS: ECL was not significantly different in grafts prepared using a drop-in loading procedure compared with grafts prepared using the standard suction protocol. The drop-in loading protocol may be particularly useful to surgeons who load their own grafts and eye bank processing technicians who encounter a "flat" DMEK graft that does not scroll or a loosely scrolled DMEK graft.

Toward electron-beam sterilization of a pre-assembled Boston keratoprosthesis.

PURPOSE: To evaluate the effects of electron-beam (E-beam) irradiation on the human cornea and the potential for E-beam sterilization of Boston keratoprosthesis (BK) devices when pre-assembled with a donor cornea prior to sterilization. METHODS: Human donor corneas and corneas pre-assembled in BK devices were immersed in recombinant human serum albumin (rHSA) media and E-beam irradiated at 25 kGy. Mechanical (tensile strength and modulus, and compression modulus), chemical, optical, structural, and degradation properties of the corneal tissue after irradiation and after 6 months of preservation were evaluated. RESULTS: The mechanical evaluation showed that E-beam irradiation enhanced the tensile and compression moduli of human donor corneas, with no impact on their tensile strength. By chemical and mechanical analysis, E-beam irradiation caused a minor degree of crosslinking between collagen fibrils. No ultrastructural changes due to E-beam irradiation were observed. E-beam irradiation slightly increased the stability of the cornea against collagenase-induced degradation and had no impact on glucose diffusion. The optical evaluation showed transparency of the cornea was maintained. E-beam irradiated corneal tissues and BK-cornea pre-assembled devices were stable for 6 months after room-temperature preservation. CONCLUSIONS: E-beam irradiation generated no detrimental effects on the corneal tissues or BK-cornea pre-assembled devices and improved native properties of the corneal tissue, enabling prolonged preservation at room temperature. The pre-assembly of BK in a donor cornea, followed by E-beam irradiation, offers the potential for an off-the-shelf, ready to implant keratoprosthesis device.

Primary Graft Failure, Infection, and Endothelial Cell Density in Corneal Transplants With Increased Death-to-Preservation Time.

PURPOSE: To ascertain whether death-to-preservation time (DPT) is associated with donor endothelial cell density (ECD), primary graft failure (PGF), and infection. METHODS: Donor corneas aged older than 10 years with ECD 2000 to 4500 cells/mm2 were procured between 2011 and 2018 by a single eye bank. Donor corneas were analyzed retrospectively for the main outcome measures of PGF, infection, and ECD. Means and proportions of study parameters were compared between corneas with long and short DPT, defined as greater or less than 14 hours, respectively, excluding corneas with a history of intraocular surgery or diabetes. Multivariate analyses were performed using logistic regression, adjusting for donor age at time of death, history of diabetes mellitus, and history of cataract surgery. RESULTS: Among 12,015 corneas, those with long DPT had a statistically but not clinically significant higher ECD than that of corneas with short DPT (2754 vs. 2724 cells/mm2, P < 0.01). There was no difference in PGF and infections in corneas with long versus short DPT (0.28% vs. 0.26%, P = 0.86; 0.43% vs. 0.29%, P = 0.51, respectively). CONCLUSIONS: Longer DPT is not associated with a clinically meaningful reduction in donor ECD, PGF, or infection.

Electron Beam Sterilization of Poly(Methyl Methacrylate)-Physicochemical and Biological Aspects.

Electron beam (E-beam) irradiation is an attractive and efficient method for sterilizing clinically implantable medical devices made of natural and/or synthetic materials such as poly(methyl methacrylate) (PMMA). As ionizing irradiation can affect the physicochemical properties of PMMA, understanding the consequences of E-beam sterilization on the intrinsic properties of PMMA is vital for clinical implementation. A detailed assessment of the chemical, optical, mechanical, morphological, and biological properties of medical-grade PMMA after E-beam sterilization at 25 and 50 kiloGray (kGy) is reported. Fourier transform infrared spectroscopy, thermogravimetric analysis, and differential scanning calorimetry studies indicate that E-beam irradiation has minimal effect on the chemical properties of the PMMA at these doses. While 25 kGy irradiation does not alter the mechanical and optical properties of the PMMA, 50 kGy reduces the flexural strength and transparency by 10% and 2%, respectively. Atomic force microscopy demonstrates that E-beam irradiation reduces the surface roughness of PMMA in a dose dependent manner. Live-Dead, AlamarBlue, immunocytochemistry, and complement activation studies show that E-beam irradiation up to 50 kGy has no adverse effect on the biocompatibility of the PMMA. These findings suggest that E-beam irradiation at 25 kGy may be a safe and efficient alternative for PMMA sterilization.

Ink Retention and Endothelial Cell Viability After the Application of an Orientation Stamp Over an Air Bubble During Descemet Membrane Endothelial Keratoplasty Graft Preparation.

PURPOSE: To investigate stamp visibility and endothelial cell loss (ECL) after the application of an orientation mark to Descemet membrane endothelial keratoplasty (DMEK) grafts supported by an air bubble. METHODS: Eighteen DMEK grafts were prepared at an eye bank using a technique where an orientation mark was applied to the stromal surface of a DMEK graft that was supported by a small air bubble placed at the edge of the 2 endothelial surfaces of the graft. Grafts were evaluated at 2 and 5 days for stamp visibility and at 5 days with calcein-AM staining for ECL. Nine grafts underwent cross-country shipping, and the ECL of shipped and nonshipped grafts was compared using unpaired t test. RESULTS: All 18 DMEK grafts exhibited a single, solid, readily visible orientation mark 2 and 5 days after preparation with a mean ECL of 13.5% ± 4.9%. Shipping conditions had no effect on stain retention or ECL. CONCLUSIONS: The application of an orientation stamp to a DMEK graft over an air bubble in an eye bank setting results in a single, solid orientation mark that is readily visible within the period in which most eye bank-prepared tissue is used. This technique produces no further ECL compared with the methods where the orientation stamp is applied through a stromal window. Eye bank technicians and surgeons can be confident that this modified preparation technique results in transplant-quality DMEK grafts with the additional benefit of conserving the stromal cap for use in other anterior lamellar procedures, thereby making efficient use of donor tissue.

Tissue Loss, Processing Time, and Primary Graft Failure in Eye Bank-Prepared Descemet Membrane Endothelial Keratoplasty Grafts Before and After Prestripped to Preloaded Graft Transition.

PURPOSE: To examine tissue loss rates, processing time, and primary graft failure (PGF) of "prestripped-only" Descemet membrane endothelial keratoplasty (DMEK) grafts at a single eye bank and how these parameters changed after the introduction of steps to preload tissue among experienced processors. METHODS: Tissue loss and processing time during DMEK graft preparation as well as PGF were analyzed retrospectively at a single eye bank between 2012 and 2018. Outcomes were assessed in consecutive grafts before and after the introduction of preloading to the eye bank's standard operating procedure. RESULTS: A total of 1326 grafts were analyzed, composed of the first 663 preloaded DMEK grafts and, for comparison, the 663 DMEK grafts processed immediately before starting the preloaded service. Mean processing time increased from 17.0 ± 3.9 minutes to 26.0 ± 5.4 minutes with the advent of preloading (P < 0.01). Initially, average processing time increased dramatically, with a maximum processing time of 51 minutes, before regressing to the average. No significant difference in the rate of tissue wastage was observed before versus after the implementation of preloaded DMEK (1.2% vs. 1.7%, P = 0.48). PGF occurred in 7 grafts before the preloaded service and 10 grafts after starting the service (1.6% vs. 2.3%, P = 0.47). CONCLUSIONS: Preloading does not affect tissue wastage for experienced technicians or the PGF rate but increases processing time. Eye banks that are considering adding preloading to their standard operating procedure may need to account for longer processing times in their daily operations.

A cytoskeletal protein complex is essential for division of intracellular amastigotes of Leishmania mexicana.

Previous studies in Leishmania mexicana have identified the cytoskeletal protein KHARON as being important for both flagellar trafficking of the glucose transporter GT1 and for successful cytokinesis and survival of infectious amastigote forms inside mammalian macrophages. KHARON is located in three distinct regions of the cytoskeleton: the base of the flagellum, the subpellicular microtubules, and the mitotic spindle. To deconvolve the different functions for KHARON, we have identified two partner proteins, KHAP1 and KHAP2, which associate with KHARON. KHAP1 is located only in the subpellicular microtubules, whereas KHAP2 is located at the subpellicular microtubules and the base of the flagellum. Both KHAP1 and KHAP2 null mutants are unable to execute cytokinesis but are able to traffic GT1 to the flagellum. These results confirm that KHARON assembles into distinct functional complexes and that the subpellicular complex is essential for cytokinesis and viability of disease-causing amastigotes but not for flagellar membrane trafficking.

Eccentrically Punched Descemet Membrane Endothelial Keratoplasty Grafts Expand the Descemet Membrane Endothelial Keratoplasty Donor Pool: One Eye Bank's Reappraisal of the Central Clear Zone Convention.

PURPOSE: To ascertain whether offsetting the Descemet membrane endothelial keratoplasty (DMEK) punch can expand the donor pool in conjunction with prepunched and preloaded services by recapturing the corneas otherwise excluded by the conventional central clear zone criteria. METHODS: In this retrospective review of corneas recovered and processed for DMEK by a single eye bank between March 2017 and October 2018, corneas failing to meet the conventional central clear zone requirement during initial evaluation (defined as an area in the central cornea where an 7.5- to 8.0-mm diameter graft can be obtained free of previous surgical scars, Descemet tears, or confined areas of endothelial defects) were further evaluated for offset punching. Corneas with a central endothelial cell density of at least 2000 cells/mm at the initial screening (average of 3 specular images assessed with the center dot method) that had a clear zone of 7.5- to 8.0-mm diameter where a graft could be obtained were designated as eligible for offset punching for either prepunched or preloaded DMEK. RESULTS: A total of 2607 corneas were found to be suitable for DMEK using the conventional central clear zone criteria. An additional 62 corneas were deemed DMEK suitable by offsetting the punch, yielding a 2.4% increase in the availability of DMEK suitable corneas. CONCLUSIONS: Offsetting the DMEK punch can recapture corneas otherwise excluded from the DMEK donor pool because of a failure to meet the conventional central clear zone criteria, and by our estimation may help eye banks meet the growing demand for DMEK tissue while maximizing the transplant potential of every cornea.

Efficacy of Amphotericin B in Corneal Preservation Media After Extended Frozen Storage.

PURPOSE: To investigate the antimycotic activity of amphotericin B deoxycholate that has been previously frozen for 28 days before supplementation of Optisol-GS. METHODS: Triplicate Optisol-GS samples were inoculated with 10 colony-forming units (CFU) of Candida albicans. Each set of triplicate cultures was supplemented with 2.5 µg/mL of amphotericin B that was either freshly resuspended and never frozen, frozen overnight at -20°C and thawed, or frozen at -20°C for 4 weeks and thawed. The cultures were stored at 4°C, with aliquots taken at 0, 6, 24, and 72 hours for quantification. The efficacy of each preparation of amphotericin B in reducing C. albicans growth was assessed at these time points. RESULTS: Six hours after antifungal supplementation, there was a 1.33 log10 CFU reduction with freshly resuspended amphotericin B, compared with a 1.31 log10 reduction with amphotericin B that was frozen overnight (P = 0.20) and a 1.18 log10 reduction with amphotericin B that was frozen for 4 weeks (P = 0.05). After 72 hours, there was a 2.72 log10 CFU reduction with freshly resuspended amphotericin B, a 2.64 log10 CFU reduction with amphotericin B that was frozen overnight (P = 0.45), and a 2.18 log10 CFU reduction with amphotericin B that was frozen for 4 weeks (P = 0.05). CONCLUSIONS: Previously frozen amphotericin B remains highly effective against C. albicans. Optisol-GS supplemented with 2.5 µg/mL amphotericin B that was frozen for 4 weeks at -20°C resulted in >90% CFU reduction by 6 hours and >99% reduction by 72 hours.

Donor DMEK Tissue Characteristics: Association With Rebubble Rate and 6-Month Endothelial Cell Loss.

PURPOSE: To determine whether specific donor characteristics influence postoperative rebubble rate and 6-month endothelial cell loss (ECL) in Descemet membrane endothelial keratoplasty (DMEK). METHODS: A retrospective analysis of a consecutive series of 857 DMEK surgeries using eye bank-prepared donor tissue was performed between September 2013 and April 2018. DMEK graft characteristics including donor age, preoperative endothelial cell density (ECD), preservation time, death-to-preservation time, and donor diabetes status were analyzed for correlation with rebubble rate and 6-month postoperative ECL. Subgroup analyses of donor age, preoperative ECD, preservation time, death-to-preservation time, preparation-to-surgery time, and diabetes severity were also performed. Statistically significant relationships between donor characteristics and rebubble rate or 6-month postoperative ECL were determined using Pearson correlation, one-way analysis of variance, t test, and χ analysis. RESULTS: The overall rate of rebubble after 857 surgeries performed by 7 surgeons during the study period was 12.6%. There was no significant relationship between postoperative rebubble rate and donor age, preoperative ECD, preservation time, death-to-preservation time, preparation-to-surgery time, or donor diabetes status. The subgroup analysis of these characteristics also yielded no significant relationship with rebubble rate. There was also no significant relationship between 6-month postoperative ECL and analyzed donor factors. CONCLUSIONS: Donor characteristics such as higher donor age, lower preoperative ECD (<2500), longer preservation time, and donor diabetes did not increase the rebubble rate or the 6-month ECL after DMEK. These results indicate that common surgeon preferences for donor tissues that are younger, fresher, with higher cell count, and without diabetes do not translate into superior postsurgical outcomes.

Efficacy and Safety of Various Amphotericin B Concentrations on Candida albicans in Cold Storage Conditions.

PURPOSE: To determine the concentration of amphotericin B that would be both effective against Candida albicans contamination and safe for corneal endothelial cells (CECs) in cold storage conditions. METHODS: Triplicate media cultures were inoculated with 10 colony-forming units (CFUs)/mL of C. albicans (American Type Culture Collection 10231), supplemented with amphotericin B (0-20 µg/mL), stored in cold conditions (2°C-8°C) for 72 hours, and analyzed quantitatively for CFUs. C. albicans concentration in each sample was determined initially and after 6, 24, 48, and 72 hours of storage. CEC mitochondrial function (oxygen consumption rate), apoptosis, and necrosis were examined in donor corneas after 7 days of amphotericin B exposure and compared with untreated controls. CEC viability was also examined by calcein-AM staining and Fiji segmentation after 72 hours or 2 weeks of amphotericin B exposure to mimic potential eye bank practices. RESULTS: Amphotericin B concentrations of 1.25, 2.5, and 5.0 µg/mL resulted in 0.47, 1.11, and 1.21 log10 CFU reduction after only 6 hours of cold storage and continued to decrease to 3.50, 3.86, and 4.49 log10 reductions after 72 hours, respectively. By contrast, amphotericin B 0.255 µg/mL showed only 1.01 log10 CFU reduction after 72 hours of incubation. CEC mitochondrial function and viability did not differ in donor corneas exposed to amphotericin B ≤2.59 µg/mL compared with the controls. CONCLUSIONS: Optimal efficacy of amphotericin B against C. albicans is achieved in cold storage conditions at concentrations ≥1.25 µg/mL, and 2.5 µg/mL reduces Candida contamination by >90% after 6 hours of cold storage without sacrificing CEC health.

Surgeon Preference for Endothelial Keratoplasty Techniques.

PURPOSE: To examine barriers and facilitators to adoption of Descemet membrane endothelial keratoplasty (DMEK) for treatment of endothelial dysfunction. METHODS: Anonymous electronic surveys were emailed to surgeons ordering corneal transplant tissue in the past 18 months from 2 eye banks in the United States (US). RESULTS: Of 365 surgeons emailed, 118 (32%) completed the survey. Most respondents were located at an urban site (59%), were in private practice (64%), and had been in practice for >10 years (53%). Participants performing DMEK (n = 70) were more likely to have been in practice for ≤10 years than those not performing DMEK (56% vs. 35%, respectively, P = 0.03). Among respondents not performing DMEK (n = 48, 41%), the most frequently reported barriers were technical difficulty (n = 30, 63%), a lack of experience (n = 29, 60%), low surgical volume (n = 20, 42%), and risk of postoperative complications (n = 19, 40%). Most (n = 46, 96%) respondents not performing DMEK were interested in doing so, with the following reported as helpful resources: wet laboratory courses (n = 35, 73%), eye bank-prepared tissue: preloaded/prestripped (n = 32, 67%), back-up tissue in case of inadvertent graft damage (n = 29, 60%), higher surgical volume to support the learning curve (n = 28, 58%), and surgical mentorship (n = 22, 46%). Only a minority wanted more data to highlight superiority over other EK procedures (n = 12, 25%). CONCLUSIONS: There is substantial interest in learning to perform DMEK among the surgical community. Barriers identified in this survey may be addressed to help surgeons increase DMEK adoption.

Patent Blue V as an Alternative Stain for DMEK Grafts: Safety, Stain Retention, and Feasibility.

PURPOSE: To determine whether Patent Blue V (PB) can be used as an alternative dye for staining Descemet membrane endothelial keratoplasty (DMEK) grafts. METHODS: DMEK grafts from donor corneas were stained with 2.5% PB for 4 minutes (n = 2), 2.5% PB for 10 minutes (n = 2), or 0.06% trypan blue (TB) for 4 minutes (n = 1). The stain intensity of the grafts was compared initially and at different time points over a 15-minute period of balanced salt solution wash using a grading scale based on a serial dilution series of PB. Three additional grafts were stained with 2.5% PB for 10 minutes and used in mock surgeries to assess stain retention after injection and manipulation in a donor eye model for 25 minutes. To assess the safety of PB, DMEK grafts were prepared from 10 pairs of corneas where one cornea was stained with 0.06% TB for 4 minutes and the mate cornea was stained with 2.5% PB for 10 minutes. The grafts were preloaded and stored for 5 days in Optisol-GS at 2°C to 8°C, and endothelial cell loss was quantified using Calcein-AM staining and FIJI segmentation by 2 masked readers. RESULTS: PB stain intensities were lighter than TB-stained grafts. Grafts stained with 2.5% PB for 10 minutes retained sufficient color for the 25-minute duration of mock surgery. The average endothelial cell loss for PB versus TB grafts was 21.6% ± 5.3% versus 20.2% ± 5.4% (P = 0.202) as determined by reader 1 and 23.6% ± 4.8% versus 21.2% ± 5.8% (P = 0.092) as determined by reader 2. CONCLUSIONS: PB is a viable alternative dye in DMEK grafts for applications where TB may not be available or approved for use.

Electron Beam Irradiated Corneal Versus Gamma-Irradiated Scleral Patch Graft Erosion Rates in Glaucoma Drainage Device Surgery.

INTRODUCTION: Patch graft erosion and implant exposure is a known complication of glaucoma drainage device (GDD) surgery. Recently, electron beam (e-beam) irradiated corneal tissue ha s become available; however, limited data exist on the rates of erosion for e-beam irradiated corneal grafts compared to traditional scleral grafts after GDD surgery. METHODS: This retrospective study examines the records of 253 eyes from 225 adult subjects who underwent GDD surgery with either e-beam irradiated corneal or scleral grafts at the Casey Eye Institute by five surgeons between April 22, 2014 and October 11, 2017. Surgical procedures and the occurrence of graft erosion were determined using billing codes and verified by manual review of electronic health records. RESULTS: The average age at the time of surgery was 61.3 ± 17.5 years (n = 200) and 60.8 ± 16.8 years (n = 53) for the e-beam irradiated cornea and sclera groups, respectively. The average follow-up time post-surgery was 416 ± 345 days and 495 ± 343 days for the e-beam irradiated cornea and sclera groups, respectively. There were no statistically significant differences in sex, age, follow-up time, and glaucoma diagnosis between the groups; however, the e-beam irradiated cornea group was statistically more likely to have an Ahmed implant as compared to the sclera group. No erosion events were noted in either group. CONCLUSION: e-Beam irradiated corneal grafts were used 3.8 times more frequently relative to scleral grafts, yet there were no cases of graft erosion in either group during the follow-up period.