Storage Of Corneas Research Articles

Suppressing Pro-Apoptotic Proteins by siRNA in Corneal Endothelial Cells Protects against Cell Death.

Corneal endothelial cells (CE) are critical for the cornea's transparency. For severe corneal damage, corneal tissue transplantation is the most promising option for restoring vision. However, CE apoptotic cell death occurs during the storage of donor corneas for transplantation. This study used small interfering (si)RNA-mediated silencing of pro-apoptotic proteins as a novel strategy to protect CE against apoptosis. Therefore, the pro-apoptotic proteins Bax and Bak were silenced in the human corneal endothelial cell line (HCEC-12) by transfection with Accell™siRNA without any adverse effects on cell viability. When apoptosis was induced, e.g., etoposide, the caspase-3 activity and Annexin V-FITC/PI assay indicated a significantly reduced apoptosis rate in Bax+Bak-siRNA transfected HCECs compared to control (w/o siRNA). TUNEL assay in HCECs exposed also significantly lower cell death in Bax+Bak-siRNA (7.5%) compared to control (w/o siRNA: 32.8%). In ex vivo donor corneas, a significant reduction of TUNEL-positive CEs in Bax+Bak-siRNA corneas (8.1%) was detectable compared to control-treated corneas (w/o siRNA: 27.9%). In this study, we demonstrated that suppressing pro-apoptotic siRNA leads to inhibiting CE apoptosis. Gene therapy with siRNA may open a new translational approach for corneal tissue treatment in the eye bank before transplantation, leading to graft protection and prolonged graft survival.

Donor Cornea Storage Temperature: Investigation of the Time Period Immediately Before Cornea Transplantation Surgery

Purpose: Donor cornea storage temperature seems to affect tissue viability; however, the time period between removal from storage and transplant surgery has not been explored. We investigated the possible impact time at room temperature in the operating room may have on endothelial cell density (ECD) and corneal thickness (CT). Methods: In our simulation, transplant-ineligible corneas underwent 1 of 4 room temperature protocols. ECD (by specular microscopy) and CT (by anterior segment optical coherence tomography [OCT]), measured before and after protocols, were used to explore possible effects of time at room temperature. Control group tissue resided at room temperature only long enough for warming and subsequent ECD and CT measurements. Results: Corneas underwent one of the following protocols: 1) 17 (±1) minutes at room temperature (for baseline OCT measurements; n = 13) followed by 23 minutes at 35°C (to facilitate specular imaging, conducted for all 4 protocols), 2) 60 (±1) minutes (1 hour; n = 8) at room temperature, 3) 187 (±1) minutes (approximately 3 hours; n = 7) at room temperature, and 4) 416 (±1) minutes (approximately 7 hours; n = 7). Statistical analysis showed no significant difference in ECD change (P = 0.353) or CT change (P = 0.415) between protocol groups. Controlling for donor age and death-to-preservation time rendered no statistical significance. Conclusions: Time at room temperature did not seem to significantly influence ECD or CT measured soon after warming. Studies exploring ECD postoperatively and long-term graft survival are needed to assess the possible clinical impact of preoperative time at room temperature.

Comparative Proteomics Reveals Prolonged Corneal Preservation Impaired Ocular Surface Immunity Accompanied by Fibrosis in Human Stroma.

Cornea transplantation is one of the most commonly performed allotransplantations worldwide. Prolonged storage of donor corneas leads to decreased endothelial cell viability, severe stromal edema, and opacification, significantly compromising the success rate of corneal transplantation. Corneal stroma, which constitutes the majority of the cornea, plays a crucial role in maintaining its shape and transparency. In this study, we conducted proteomic analysis of corneal stroma preserved in Optisol-GS medium at 4 °C for 7 or 14 days to investigate molecular changes during storage. Among 1923 identified proteins, 1634 were quantifiable and 387 were significantly regulated with longer preservation. Compared to stroma preserved for 7 days, proteins involved in ocular surface immunomodulation were largely downregulated while proteins associated with extracellular matrix reorganization and fibrosis were upregulated in those preserved for 14 days. The increase in extracellular matrix structural proteins together with upregulation of growth factor signaling implies the occurrence of stromal fibrosis, which may compromise tissue clarity and cause vision impairments. This study is the first to provide insights into how storage duration affects corneal stroma from a proteomic perspective. Our findings may contribute to future research efforts aimed at developing long-term preservation techniques and improving the quality of preserved corneas, thus maximizing their clinical application.

Porcine Cornea Storage ExVivo Model as an Alternative to Human Donor Tissues for Investigations of Endothelial Layer Preservation.

Due to the growing shortage of human corneas for research, we developed a porcine cornea storage model with qualitative features comparable to human tissues. We established a decontamination procedure for porcine eye bulbs to ensure corneal storage at 31°C to 35°C for up to 28 days without contamination. We compared human and porcine corneas under hypothermic (2-8°C) or culture (31-35°C) conditions for central corneal thickness (CCT), corneal transparency, endothelial morphology, endothelial cell density (ECD), and a novel method to quantify whole endothelial mortality. We also examined portions of lamellar tissues consisting of Descemet's membrane and endothelial cells under the microscope after Alizarin red staining. Our decontamination procedure reduced corneal contamination from 94% (control corneas without decontamination) to 18% after 28 days of storage at 31°C to 35°C. ECD, CCT, transparency, and morphology were significantly higher in porcine corneas than in human corneas at day 0. Nevertheless, the qualitative parameters of porcine and human corneas showed comparable trends under both investigated storage conditions for up to 14 days. The presented corneal storage model provides a reliable alternative to human tissues for preliminary corneal investigations. The porcine cornea storage model can be used to investigate the efficacy and safety of new media, substances, or storage conditions. Furthermore, the method developed to assess the percentage of endothelial mortality is tissue conservative and can be used in eye banks to monitor endothelial mortality during storage of tissues intended for transplantation.

The Effect of Bioregulators Isolated from Blood Serum and Cornea of the Bovin’s Eye on the Condition of Tissue and Cells of the Rabbit Corneas during Cultivation and Storage

Objective: to study the condition of the cornea, as well as its epithelial and endothelial cells, while maintaining in vitro at various temperature conditions, under the influence of a number of factors, including bioregulators isolated from blood serum and cornea of the bovine, and epidermal growth factor.Methods. The study was carried out on rabbit corneas stored at temperatures of +4, –86 °C, as well as the cultivation of endothelial and epithelial cells isolated from the cornea after storage at these temperatures, followed by histological and immunohistochemical studies.Results. Storage of the cornea at +4 °C for 10 days leads to corneal edema and significantly reduces their transparency, both bioregulators partially prevent a decrease in the transparency of the cornea, while the endothelial layer lyses in groups with the addition of epidermal growth factor and corneal bioregulator; but remains in the cornea with the addition of a serum bioregulator. All three factors contribute to the preservation of the Bowman membrane. In the corneas stored at –86 °C on the 30th day, a preserved endothelial layer was observed, and the epithelium retained its multilayering in all groups with the addition of factors other than the control group. In the control samples, the epithelial layer partially exfoliated, the endothelial layer was almost completely lysed. Both bioregulators stimulated the proliferation of cells isolated from the native cornea and enhanced the action of the epidermal growth factor. Similar results were obtained on cells isolated from stored corneas for 2 weeks at –86 °C. In the case of combined use of the epidermal growth factor and bioregulators on the 30th day, the endothelial layer was mainly preserved, the Descemet’s membrane was not broken. In the control samples, the epithelium was mainly single-layered, partially exfoliated, and the endothelial layer was completely lysed.Conclusion. Storage of cornea during hypothermia (+4 °С) does not provide corneal viability for longer than 10 days. Storage under conditions of cryopreservation (–86 °C) ensures the viability of the cornea for 60 days. Adding bioregulators and an epidermal growth factor to the basic preservation medium allows one to obtain a structurally safe and viable cornea, while all cellular layers of the cornea, including the endothelial layer, are preserved and viable.

Cluster of Symptomatic Graft-to-Host Transmission of Herpes Simplex Virus Type 1 in an Endothelial Keratoplasty Setting

PurposeDescemet's membrane endothelial keratoplasty (DMEK) is becoming the gold standard to treat corneal endothelial dysfunctions worldwide. Compared with conventional penetrating keratoplasty, infectious complications after DMEK are ill defined. We describe the clinical picture of 2 DMEK recipients, operated on the same day and in the same clinic, who developed atypical herpes simplex virus type 1 (HSV-1) infection in the transplant recipient eye within days post-DMEK. Because recipients received cornea tissue from 2 different donors prepared by the same eye bank, the likelihood of a common HSV-1 source was determined.DesignCase series.ParticipantsTwo DMEK recipients who developed atypical intraocular HSV-1 disease shortly after surgery and surplus cornea specimens of 6 donors.MethodsSurplus cornea donor (pre-DMEK cornea remnants and conditioned cornea storage and transport media) and recipient samples (post-DMEK aqueous humor) were assayed for HSV-1 DNA and infectious virus by real-time polymerase chain reaction (RT-PCR) and cell culture, respectively. Target-enriched whole viral genome sequencing was performed on HSV-1 DNA–positive ocular specimens.Main Outcome MeasuresClinical picture of atypical intraocular HSV-1 infection post-DMEK and presence and homology of HSV-1 genomes between ocular specimens of DMEK donors and recipients.ResultsHerpes simplex virus type 1 DNA was detected in aqueous humor and donor cornea specimens of both DMEK cases, but not in the cornea remnants of 6 randomly selected donors processed by the same eye bank. Infectious HSV-1 was isolated from the cornea remnant and corresponding culture medium of 1 cornea donor. Notably, whole-genome sequencing of virus DNA-positive specimens demonstrated exceptionally high genetic similarity between HSV-1 strains in recipient and donor specimens of both DMEK cases.ConclusionsData indicate cross-contamination of cornea grafts during DMEK preparation with subsequent graft-to-host HSV-1 transmission that caused atypical sight-threatening herpetic eye disease shortly after DMEK. Ophthalmologists should be aware that HSV-1 transmission by DMEK is possible and can lead to atypical ocular disease, a condition that can easily be prevented by taking appropriate technical and clinical measures at both eye bank and surgical levels.

Переживаемость кератоцитов и эндотелиальных клеток заднего послойного трансплантата роговицы, культивированных в модифицированной консервационной среде

Purpose. To study the survival of keratocytes and endothelial cells of a human donor cornea storage in the standard and the new media which was specifically designed for optimized cornea hydration. Material and methods. 2D cell cultures of keratocytes and endothelial cells obtained from the Eye tissue bank were used for culture in improved storage media over a period of 14 and 7 days subsequently. To confirm phenotype characteristics, the cells were stained by the following markers: for keratocytes – Lumikan, Keratocan, and α-smooth muscle actin; for endothelial cells – ZO-1 and Na/K-ATPase. The onset of apoptosis in cell culture of keratocytes were detected with Cytochrome C, BAX, and Caspase 3 and 8. Viability of cell cultures after the cultivation was carried out using a commercial set of "Live and Dead". Morphology of the endothelial cells was assessed using an electron scanning microscope. Results. It was shown that the 2D keratocyte culture cultured in the improved storage media expressed specific markers: Lumican, Keratocan, and did not express α-smooth muscle actin. There were no markers of apoptosis in the cell culture of keratocytes after 14 days of cultivation. Corneal endothelium cultured in the improved storage media expresses ZO-1, Na/K-ATPase and presented hexagonal cell shape morphology according to electron microscopy. Conclusion. The improved storage media allow to preserve the unique phenotype of keratocytes, with a slight decrease in proliferative cells activity during 14 days. The media maintain a viable and functional corneal endothelium for at least seven days of cultivation. Key words: cell culture; corneal endothelium; keratocyte; posterior lamellar graft, corneal storage media.

Microtubule Stabilization Protects Hypothermia-Induced Damage to the Cytoskeleton and Barrier Integrity of the Corneal Endothelial Cells.

Purpose: To determine the impact of hypothermia on the barrier function of donor corneal endothelium, thereby enhancing the success of corneal transplantation. Methods: Primary cultures of porcine endothelial cells were subjected to hypothermia (15 h; 4°C). The impact on microtubule assembly, peri-junctional actomyosin ring (PAMR), and ZO-1 was assessed by immunocytochemistry with and without pretreatment with a microtubule-stabilizing agent (Epothilone B; EpoB; 100 nM) and a p38 MAP kinase inhibitor (SB-203580; 20 μM). In addition, EpoB-loaded PLGA nanoparticles (ENPs) prepared by nanoprecipitation technique and coated with poly-L-lysine (PLL-ENPs) were administered one-time for sustained intracellular delivery of EpoB. Results: Exposure to hypothermia led to microtubule disassembly concomitant with the destruction of PAMR and the displacement of ZO-1 at the cellular periphery, suggesting a loss in barrier integrity. These adverse effects were attenuated by pretreatment with EpoB or SB-203580. PLL-ENPs possessed a zeta potential of ∼26 mV and a size of ∼110 nm. Drug loading and entrapment efficiency were 5% (w/w) and ∼87%, respectively, and PLL-ENPs showed a biphasic release in vitro: burst phase (1 day), followed by a sustained phase (∼4 weeks). Pretreatment with PLL-ENPs (0.4 mg/mL) for 24 h stabilized the microtubules and opposed the hypothermia-induced damage to PAMR and the redistribution of ZO-1. Conclusions: Hypothermia induces microtubule disassembly via activation of p38 MAP kinase and subsequently breaks down the barrier function of the endothelium. Sustained intracellular delivery of EpoB using nanoparticles has the potential to overcome endothelial barrier failure during prolonged cold storage of donor cornea.

A case series of donor tissue-transmitted keratitis and endophthalmitis following Descemet stripping automated endothelial keratoplasty

Donor-transmitted infection is one of the most dreaded complications after cornea transplant. We report four cases of fulminant graft infection with endophthalmitis following descemet stripping automated endothelial keratoplasty (DSAEK); all of these resulted from donor contamination by multidrug-resistant gram-negative bacteria (Pseudomonas in two cases, Proteus and Enterobacter in the other two). All donor corneas were retrieved in hospital premises, either ward or mortuary suggesting a possible nosocomial origin of these microorganisms. All four patients required surgical intervention for control of infection and had dismal clinical outcomes. Furthermore, in three cases, mate cornea also showed similar contamination, and additional two of these recipients also developed graft infection indicating an infection rate as high as 86%. This case series highlights an important issue of fulminant nosocomial donor cornea-related infections, possible despite standard precautions, with emergence of multidrug-resistant virulent pathogens. Adequate training of eye bank and health-care personnel and ensuing stringent protocols for retrieval and storage of corneas may help in reducing such sight-threatening adverse reactions and also consequent further ramifications.

Comparison of porcine corneal decellularization methods and importance of preserving corneal limbus through decellularization.

The aim of this study is to compare the three previously applied, conventional porcine corneal decellularization methods and to demonstrate the importance of preserving the corneal limbus through decellularization. Fresh, wild-type (with or without) limbus porcine corneas were decellularized using three different methods, including (i) sodium dodecyl sulfate (SDS), (ii) hypertonic saline (HS), and (iii) N2 gas (NG). Post-treatment evaluation was carried out using histological, residual nuclear material, and ultrastructural analyses. Glycerol was used to help reduce the adverse effects of decellularization. The corneas were preserved for two weeks in cornea storage medium. All three decellularization methods reduced the number of keratocytes at different rates in the stromal tissue. However, all methods, except SDS, resulted in the retention of large numbers of cells and cell fragments. The SDS method (0.1% SDS, 48h) resulted in almost 100% decellularization in corneas without limbus. Low decellularization capacity of the NG method (<50%) could make it unfavorable. Although HS method had a more balanced damage-decellularization ratio, its decellularization capacity was lower than SDS method. Preservation of the corneoscleral limbus could partially prevent structural damage and edema, but it would reduce the decellularization capacity. Our results suggest that SDS is a very powerful decellularization method, but it damages the cornea irreversibly. Preserving the corneoscleral limbus reduces the efficiency of decellularization, but also reduces the damage.

Eye bank and theatre factors for positive microbiological culture of corneoscleral rim and cornea storage medium in the real-world.

To evaluate microbiological culture rate and positivity of corneoscleral rim and cornea storage media as well as possible risk factors for contamination with real-world data. Data of consecutive cornea donors implanted in the reference centre from January 2013 to January 2018 were reviewed. Information about cornea characteristics (donor demographic data, endothelial cell density, type of cornea conservation, days of storage, and precut vs full-thickness tissue), and microbiological culture information (corneoscleral rim vs storage sample, positive result) were statistically analysed. During the study period, 1369 corneas (737 donors) were implanted. Cultures were performed in 76.8% (n = 1052) of them and were positive in 3.2% of cases, mainly bacteria (84.4%). Corneas preserved in hypothermia represented 61.8% of all positive microbiology results (p < 0.001). Other analysed risk factors did not reach statistically significant association with microbiological positivity. None of the 34 cases with positive microbiological cultures reported ocular infection for the recipients in at least 6 months' follow-up. Microbiological tests rate in real-world practice are high despite not being compulsory. Organotypic cultured corneas showed a statistically less positivity in corneoscleral and storage medium than hypothermic ones, resulting in another advantage of this kind of cornea storage. Although precut corneas are thought to present less microbiological positivity, a statistically significant association was not found in the present study.

Evaluation of various preservation media for storage of donor corneas.

Purpose:To compare the physical and microbiological characteristics of McCarey-Kaufman (MK), Cornisol, and Optisol-GS media and evaluate the outcomes of keratoplasty performed using corneas stored in these three media.Methods:The study involved 60 donor corneas which were distributed in 3 groups: MK, Cornisol, and Optisol-GS. Corneas in these groups were further analyzed based on the type of keratoplasty performed (full thickness versus endothelial keratoplasty). At baseline, the endothelial cell density and death to preservation time of donor corneas were recorded. Following keratoplasty, patients were evaluated on day 1, at 1 month, 3 months, and 6 months follow-up. Outcomes were assessed in terms of corrected distance visual acuity (CDVA), endothelial cell density, percentage endothelial cell loss, and corneal thickness. The storage media were also assessed for their physical quality and their microbiological characteristics.Results:Physical characteristics of all three media were found to be within normal limits. Mean CDVA was comparable among the 3 groups at 6-month follow-up. The absolute endothelial cell count values were significantly lower for corneas stored in MK medium (1873.7 ± 261.1 cells/mm2) compared to the Cornisol (2085.0 ± 230.3 cells/mm2) and Optisol-GS media [(2180.3 ± 217.2 cells/mm2) (P = <0.001)]. Corneas stored in Optisol-GS medium were significantly thinner at 1-month follow-up with no significant difference at 6 months (P = 0.66).Conclusion:Optisol-GS and Cornisol media were found to preserve endothelial cell density better and stabilize corneal thickness earlier as compared to the MK medium. However, the functional outcomes were comparable among the three groups.

The impact of herpes simplex virus on the cornea engraftment

According to the recent WHO data, 39 million people in the world are blind. In developing countries cornea diseases are the second most common cause of blindness. Cornea transplantation remains the only radical method to regain lost vision for many blind people around the world. However, according to literature reports, cadaveric donor corneas pose a potential risk of herpes virus transmission to the recipient during penetrating keratoplasty. It is known that herpes simplex virus-1 persisting in the donor cornea can adversely affect graft survival up to causing the graft failure reaction. The latent herpes simplex virus may be reactivated by a number of factors, most of them occurring with penetrating keratoplasty. One of these factors is immunosuppressive therapy, an essential element of the pharmacological graft protection. Antiviral agents are strongly recommended in order to inhibit the replicating herpes simplex virus in the cornea graft. The most common antiviral agents are interferons with their inducers and acyclic nucleosides. Viral decontamination during cornea storage would prevent the donor-to-recipient transmission of herpes simplex virus in relation to keratoplasty.

Commentary: Long-term solutions to revive eye banking in India in COVID-19 era - Needs more than long-term corneal preservation.

Commentary: Long-term solutions to revive eye banking in India in COVID-19 era - Needs more than long-term corneal preservation.

COVID-19 pandemic from an ophthalmology point of view.

Coronavirus disease 2019 (COVID-19) is caused by a highly contagious RNA virus termed as severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Ophthalmologists are at high-risk due to their proximity and short working distance at the time of slit-lamp examination. Eye care professionals can be caught unaware because conjunctivitis may be one of the first signs of COVID-19 at presentation, even precluding the emergence of additional symptoms such as dry cough and anosmia. Breath and eye shields as well as N95 masks, should be worn while examining patients with fever, breathlessness, or any history of international travel or travel from any hotspot besides maintaining hand hygiene. All elective surgeries need to be deferred. Adults or children with sudden-onset painful or painless visual loss, or sudden-onset squint, or sudden-onset floaters or severe lid oedema need a referral for urgent care. Patients should be told to discontinue contact lens wear if they have any symptoms of COVID-19. Cornea retrieval should be avoided in confirmed cases and suspects, and long-term preservation medium for storage of corneas should be encouraged. Retinal screening is unnecessary for coronavirus patients taking chloroquine or hydroxychloroquine as the probability of toxic damage to the retina is less due to short-duration of drug therapy. Tele-ophthalmology and artificial intelligence should be preferred for increasing doctor-patient interaction.

A new approach to extend the storage of donor corneas after 28days of corneal culture in an Italian eye bank.

This study aimed to evaluate the possibility to extend the storage of unused organ-cultured donor corneas. After 28 days of corneal culture in TISSUE-C (AL.CHI.MI.A. S.R.L., Italy) and 5-day storage in transport/deswelling medium CARRY-C (AL.CHI.MI.A. S.R.L., Italy), 25 corneas that were deemed suitable for transplantation were transferred in fresh TISSUE-C at 31°C for additional 7days and then in fresh CARRY-C at room temperature for 24h. Tissues were assessed for endothelial cell density (ECD), endothelial mortality and morphology after the standard and the extended corneal storage. In addition, the effect of donor age < 85years and ≥ 85years on corneal characteristics was assessed. After the extended storage, 6 out of 25 tested corneas (24%) showed ECD values below the acceptance limit (< 2000cells/mm2). 19 corneas (76%) were still suitable for transplantation and showed a 5.9% loss in ECD, which was not statistically significant (P = 0.0949) compared to standard storage period. The two donor age groups did not show statistically significant differences in any tested parameter, although a trend for lower ECD and higher mortality in Descemet's folds after standard storage was observed in the ≥ 85 age donor group. Thus, the attempt of the current study to provide new sight-restorative options for unused tissues and increasing the availability of corneas in case of shortage gave encouraging results. Although a higher vulnerability of corneas from very old donors could not be statistically demonstrated in the present study, higher sample size could be required for prolonging the shelf life of these tissues.

Lower risk of graft dislocation applying organ cultured corneas compared with cold short-term cultured corneas during DSAEK surgery.

To investigate factors that may affect graft dislocation after uneventful Descemet stripping endothelial keratoplasty (DSAEK), with a special focus on different donor cornea storage conditions. All DSAEK surgeries performed during 6 years in our hospital were retrospectively investigated. Patients with previous intraocular surgeries other than cataract extraction were excluded. Donor data (age, gender, death-to-preservation time, cornea storage conditions, graft storage time, and estimated thickness of the graft) and patient information (age, gender, and indication for surgery) were noted. A total of 26 dislocations (6.6%) out of 394 DSAEK surgeries were found. The dislocation rate was considerably reduced between 2011 and 2016, due to more experienced surgeons. There was a significantly higher dislocation rate with the use of donor corneas stored in cold storage solution (12.7%) compared with organ culture solution (2.5%) (P < 0.001). During the study period, donor corneas stored in cold storage solution and organ culture solution were applied in 158 (40%) and 236 (60%) cases, respectively. There were no differences in recipient age and gender between patients who received corneas from the two storage systems, indicating a random distribution of grafts. In a multivariate analysis, only operation year (reduced rate over the years) and cold storage solution were statistically significantly associated with graft dislocation. This study suggests that the type of storage solution may have a decisive role in graft dislocation in DSAEK surgery.

Procurement, storage and utilization trends of eye banks in India.

Purpose:To study the trends in collection, storage and utilization of donor corneas in eye banks in India.Methods:The data was collected from 12 eye banks in India that collected more than 1000 corneas per year. The retrospective analysis of the parameters like characteristics of the donor and the host, storage media used, number of eyes collected, number of eyes utilized, causes of non-utilization of the tissue and the procedures performed was done.Results:A total of 20,564 eyes were collected by the 12 eye banks during the year 2013–2014. Voluntary eye donation (VED), and hospital cornea retrieval program (HCRP) contributed to 59.6% and 40.4% of tissue procurement respectively. Whole globe enucleation (52.3%) was more commonly performed as compared to in-situ excision of the donor corneas. The most commonly used storage media at all eye banks was McCarey-Kaufman (MK) media (83.3%). The utilization rate of the donor eyes was 50.5%. The most frequent indication for corneal transplantation was infection (active infection - 33.13%, healed infection - 10.78%) followed by Pseudophakic bullous keratopathy (PBK) (13.57%). Full thickness keratoplasty (optical penetrating keratoplasty - 47.23%, therapeutic penetrating keratoplasty - 31.74%) was performed most often followed by endothelial keratoplasty (12.41%) in the developing country.Conclusion:VED still contributes to majority of the donor tissue retrieval in India. The majority of the eye banks still utilize whole globe enucleation technique and store tissues in MK media. Trends from previous years showed a change towards HCRP, in-situ excision technique and preservation in the long-term storage media.

Outcome of Human Donor Corneas Stored for More Than 4 Weeks.

Organ culture is the most common method for preservation of human donor corneas within Europe, and the maximum accepted storage period is typically set to 4 weeks. However, little is known about the fate of prolonged (4+ wks) organ culture storage of donor corneas. The aim of this study was to evaluate the clinical outcomes of corneal grafts preserved from 4 to 7 weeks compared with corneal grafts preserved up to 4 weeks in organ culture medium. A registry study was performed, identifying 1252 corneal transplantations [Descemet stripping automated endothelial keratoplasty (DSAEK) and penetrating keratoplasty (PKP)] between February 2008 and November 2015 at Aarhus University Hospital, Denmark. Data were extracted from The Danish Cornea Bank Registry and The Swedish Cornea Registry, which also includes Danish patients. The outcomes evaluated, 2 years after transplantation, were graft function, rebubbling rates in DSAEK surgeries, rejection rates, and endothelial cell density (ECD). A total of 808 donor corneas were stored for ≤29 days, with a mean preservation time being 20 ± 6 days (mean ± SD, range: 6-29 days). A total of 444 donor corneas were stored for >29 days, with a mean preservation time being 38 ± 7 days (mean ± SD, range: 30-72 days). In both DSAEK (n = 835) and PKP (n = 417), no statistically significant difference was observed in the frequency of graft failure (DSAEK, P = 0.9 and PKP, P = 0.74), rebubbling rates (DSAEK P = 0.07), rejection rates (DSAEK, P = 0.08 and PKP, P = 0.95), or ECD (DSAEK, P = 0.56 and PKP, P = 0.44) between donor corneas cultured for ≤29 days and donor corneas cultured for >29 days. Corneas stored in organ culture for more than 4 weeks are equally suitable for DSAEK and PKP transplantation as corneas stored in organ culture for up to 4 weeks.

Storage of Porcine Cornea in an Innovative Bioreactor.

To quantify cell survival and tissue structure preservation of porcine cornea stored in a bioreactor (BR) that recreates a transcorneal pressure gradient equivalent to intraocular pressure (IOP) and renews the medium. A BR comprising endothelial and epithelial chambers was machined in a biocompatible material. The porcine cornea, securely held, separated the chambers. Medium flow and pressure inside the endothelial chamber were maintained by a peristaltic pump. In the epithelial chamber, the corneal surface was alternatively exposed to air and a specific medium. Two transparent windows allowed thickness measurement by optical coherence tomography without opening the BR. Porcine corneas stored in the BR-on (pressure 20 mm Hg, flow 5 μL/min, temperature 31°C) were compared with (1) BR-off (no pressure or flow); (2) organ culture; and (3) Petri dish with agar on the endothelial side. Epithelial and limbal structure and differentiation, corneal transparency and thickness, and endothelial viability were compared after 7 days of storage and with fresh corneas. Corneas stored in the BR-on were thinner and more transparent than those stored with the other methods. The BR-on preserved a stratified and differentiated (K3/K12+) corneal epithelium and undifferentiated basal limbal cells with stemness markers (K3/K12-; ABCB5, K14, p75+), as well as endothelial integrity. By recreating equivalent IOP and medium renewal, the BR obtained unprecedented storage quality of porcine corneas and preserved their main epithelial, limbal, and endothelial characteristics.