Preparation of endothelial keratoplasty lamellae from donated whole eyes post vitreous humour aspiration at the central eye bank of Iran.

To compare the endothelial parameters and thickness profiles of endothelial keratoplasty (EK) lamellae from donated whole eyes post vitreous humour aspiration (VHA) with those prepared from their mate control eyes (without VHA). Between March 2019 and March 2020, a few steps were added when aspirating the vitreous humour and also before dissecting the corneal tissue with microkeratome. EK lamellae were prepared from whole eyes that underwent VHA and their corresponding endothelial and thickness profiles were compared with those prepared from their respective fellow control eyes. Post-operative data in terms of graft attachment and clarity and the rate of reoperation were also reviewed. 115 eyes that underwent VHA and 115 of their respective fellow eyes were enrolled. No significant difference was noted in the endothelial parameters between the two groups. Mean central thickness of the EK lamellae and increase of thickness towards the periphery were not significantly different between the groups. Both groups did not show a significant difference with respect to the anticipated dissection depth, post-operative graft clarity, graft attachment, and the rate of regraft. This study demonstrates that aspirating vitreous humour has no adverse effect on the endothelial and thickness profiles of the EK lamellae that are prepared from these donated whole eyes, once the specific steps outlined by the eye bank are adhered to when aspirating the vitreous humor before performing the microkeratome dissection. The success rate of the grafted lamellae was comparable between the study groups.

Comparative culture of human corneal endothelial cells following treatment with human platelet lysate/fibrin hydrogel versus Y-27632 ROCK inhibitor: in vitro and ex vivo study.

PURPOSE: The advancement of tissue engineering and cell therapy research has resulted in innovative therapeutic options for patients with corneal endothelial diseases. The aim of this study was to compare the potential effect of using human platelet lysate (HPL)/Fibrin hydrogel versus using a Y-27632 ROCK inhibitor, on the culture of human corneal endothelial cells (HCECs) under in vitro and ex vivo conditions. METHODS: HCECs were isolated from human donors and treated separately with HPL/Fibrin hydrogel, a Y-27632 ROCK inhibitor, and fetal bovine serum (FBS). MTT viability assay and cell counting were performed on the treated cells. Subsequently, we prepared ex vivo models of human corneal endothelial dysfunction and incubated them with DiI-labeled-HCECs. Specular and fluorescence microscopy were then performed on each of the ex vivo models. RESULTS: In comparison, similar viability results were achieved in the cells treated with HPL/Fibrin hydrogel versus those treated with the Y-27632 ROCK inhibitor, but both treatments showed higher viability than the control group (FBS). More importantly, based on the specular and fluorescence microscopic results, the HPL/Fibrin hydrogel and the Y-27632 ROCK inhibitor treatments showed similar inducible effects on the attachment of the cells to the Descemet membranes of the ex vivo models. CONCLUSION: HPL/Fibrin hydrogel and Y-27632 ROCK inhibitor have similar inducible effects on the viability and attachment of the HCECs. A definite advantage of treating cells with HPL/Fibrin hydrogel is that it serves as a xeno-free and biocompatible material which can have autologous applications in future usage by clinics.

Potential of a novel scaffold composed of human platelet lysate and fibrin for human corneal endothelial cells.

Cell-based therapies have been emerged to find innovative solutions for corneal endothelial dysfunction. The aim of this study is to investigate the suitability of a blended scaffold containing human platelet lysate (HPL) and fibrin not only for cultivating human corneal endothelial cells (HCECs) but also for serving as a scaffold for the respected cells. We isolated HCECs from human donors and encapsulated the cells with three concentrations of HPL/Fibrin scaffold, namely HPL/Fibrin 1, HPL/Fibrin 2 and HPL/Fibrin 3, by adding 28.9, 57.8 and 86.7 mg/dl of fibrinogen to HPL to obtain a final percentage of 10, 20 and 30 % of fibrinogen, respectively. SEM imaging and swelling test were done to characterize the scaffolds. Cell viability assay and cell counting were performed on the cells. HCECs were characterized by morphology and immunocytochemistry. SEM imaging on freeze-dried scaffolds showed higher porosity of HPL/Fibrin 1 and HPL/Fibrin 2 than HPL/Fibrin 3, but larger pores were observed only in HPL/Fibrin 1. Cellular attachment and morphology on HPL/Fibrin 1 were appropriate by SEM imaging. A higher swelling rate was observed in HPL/Fibrin 1. After 3 and 5 days, higher numbers of cells were observed specifically in HPL/Fibrin 1. A higher expression of Na+/K+-ATPase, ZO-1 and vimentin proteins was detected in the HPL/Fibrin 1-cultured HCECs as compared with control (no scaffold). HPL/Fibrin can be used as a suitable scaffold for HCECs while preserving the cells viability. Further investigations are necessitated to approve the beneficial effects of the suggested scaffold for delivering and transplantation of cultivated HCECs into the anterior chamber of the eye.

Potential Effect of Human Platelet Lysate on in vitro Expansion of Human Corneal Endothelial Cells Compared with Y-27632 ROCK Inhibitor.

PURPOSE: Corneal endothelial cell (CEC) therapy can be used as a promising therapeutic option for patients with various corneal endothelial dysfunctions. In this study, we compared the proliferative effect of human platelet lysate (HPL), as a xeno-free medium supplement, with Y-27632 Rho/rho-associated protein kinase (ROCK) inhibitor, as a well-known proliferative and adhesive agent for CECs, and fetal bovine serum (FBS) as the control, in the culture medium of human corneal endothelial cells (HCECs). METHODS: We isolated HCECs from human donors and treated the cells as three different treatment groups including 20% HPL only, 10 µM Y-27632 ROCK inhibitor, combination of 20% HPL and 10 µM Y-27632 ROCK inhibitor, and 20% FBS as the control group. ELISA cell proliferation assay and cell counting was performed on the treated cells. Finally, HCECs were characterized by morphology and immunocytochemistry (ICC). RESULTS: There was no significant proliferative effect of HPL on cell proliferation compared with the cells treated with Y-27632 ROCK inhibitor or the combination of HPL and Y-27632 ROCK inhibitor, but all the respected treatments had significant inducible effect on cell proliferation as compared with FBS-treated cells. The cells grown in all three treatment groups exhibited CEC morphology. Also, there was a higher expression of Na+/K+-ATPase and ZO-1, as CEC characteristic markers, in the culture of HCECs treated with HPL as compared with FBS. CONCLUSION: HPL offers a xeno - free and affordable medium supplement for CEC expansion that can be used in clinical applications.

Sinasol versus Optisol-GS for cold preservation of human cornea: a prospective ex vivo and clinical study.

To compare ex vivo results of donor corneas maintained in Sinasol with those stored in Optisol-GS and reporting clinical outcomes of grafted Sinasol-versus Optisol-GS-stored corneas. In phase I, paired donor corneas were maintained in Sinasol or Optisol-GS. Afterward, the corneas were subjected to slit-lamp biomicroscopic and specular microscopic examinations on days 1 and 7, and then to trypan blue staining on day 7. The same examinations were performed on the corneas that were kept in Sinasol or Optisol-GS for 14 days. In phase II, the post-operative reports of 72 consecutive corneal transplantations were recorded using Sinasol- or Optisol-GS-preserved corneas. In phase I, 128 corneas from 64 donors and 59 corneas from 33 donors were investigated for 7 and 14 days, respectively. The EC indices were comparable between the groups at the measurement periods. The EC losses over 7 and 14 days were 3.7% and 19.9% in Sinasol against 4.6% and 20.8% in Optisol-GS. Although fair quality corneas were more common in Optisol-GS group after 7 (P = 0.04) and 14 days (P = 0.034), changes of stromal edema, Descemet's fold, and other quality ratings during 14 days were not different between the groups. In phase II, all the transplanted corneas were postoperatively clear with no adverse reactions. The overall results indicate that Sinasol is a safe, effective, and affordable intermediate cold storage medium for preservation of corneas.

Preparation of endothelial keratoplasty lenticules with Gebauer SLc Original versus Moria CBm Carriazo-Barraquer and Moria One-Use Plus microkeratomes.

Purpose: To investigate endothelial keratoplasty lenticules prepared from fresh whole eyes via Gebauer SLc Original (SLc) versus Moria CBm Carriazo-Barraquer (CBm), and those prepared from corneoscleral buttons via SLc versus Moria One-Use Plus (OUP) in terms of eye bank preparation criteria. Methods: Fresh whole eyes-dissected endothelial keratoplasty lenticules with SLc were compared with CBm in terms of thickness profile measurements, over/under dissection values, endothelial cell loss, and postoperative graft failures. A similar comparison was made between corneoscleral buttons-dissected endothelial keratoplasty lenticules with SLc and OUP. Results: Means of central thicknesses and increase of thickness toward periphery were not significantly different between 33 fresh whole eyes-dissected endothelial keratoplasty lenticules with SLc and 33 fresh whole eyes-dissected ones with CBm. There was no significant difference between 19 corneoscleral buttons-dissected endothelial keratoplasty lenticules with SLc and 19 corneoscleral buttons-dissected ones with OUP in terms of mean central thickness and post-cut endothelial cell loss. However, in the corneoscleral buttons-dissected endothelial keratoplasty lenticules, a mean increase of thickness was significantly different from central to two pericentral locations with OUP (P = 0.001) and from central to two peripheral parts with SLc (P = 0.011). Both CBm and OUP systems showed deeper dissection depths than head descriptions as compared to SLc (P < 0.001). Conclusion: Unlike fresh whole eyes-dissected endothelial keratoplasty lenticules with SLc or CBm, thickness profiles of corneoscleral buttons-dissected endothelial keratoplasty lenticules with both SLc and OUP systems showed a partial asymmetric increase of thickness toward the periphery. A high agreement was observed between endothelial keratoplasty lenticules thicknesses and SLc nomograms.

Proposed risk factors for failure of transplanted eye bank-prepared descemet stripping automated endothelial keratoplasty tissues.

To investigate the possible risk factors for failure of transplanted eye bank-prepared Descemet stripping automated endothelial keratoplasty (DSAEK) tissues. In a retrospective study between March 2011 and March 2019, all the failed DSAEK cases (131 cases) reported to the Central Eye Bank of Iran were compared with a surgeon-matched successful DSAEK group (control, 126 cases) in terms of the donor, DSAEK tissue, and recipient characteristics. Univariate analysis indicated that the DSAEK tissue preparation from excised corneoscleral tissues (OR 2.17; P = 0.026) and donor conjunctival hyperemia (OR 1.62; P = 0.042) were more common in the re-DSAEK group than in the controls. Other donor and recipient characteristics as well as other DSAEK tissue criteria were not significantly different between the re-DSAEK and control groups. DSAEK lenticules in both re-DSAEK and control groups demonstrated symmetric contours. Subgroup analyses in the re-DSAEK group revealed a higher rate of surgeons' low to moderate experience in comparison to those of high experience (P < 0.001). Additionally, failed DSAEK was observed more commonly in donors aged over 50 years than among those under 30 years (P < 0.001). Our study showed that DSAEK tissues prepared from excised corneoscleral tissues and from donors with conjunctival hyperemia were associated with higher rates of re-DSAEK due to failed DSAEK. An increasing trend of re-DSAEK was observed with shorter surgeons' experience and more advanced donor's age.

Trephine- and dye-free technique for eye bank preparation of pre-stripped Descemet membrane endothelial keratoplasty tissue.

We describe a rapid and easy technique for the preparation of pre-stripped Descemet membrane (DM) endothelial keratoplasty (DMEK) tissue in the eye bank, without the use of dye or trephining by the eye bank technician. After drawing 2 radial marks 90° apart on the scleral rim of the cornea, a circumferential incision was made internal to the trabecular meshwork and the edge of the DM opposite to the marked area was scored and gently peeled towards the marked area. Finally, the peeled DM was gently unrolled and attached to the posterior stromal bed via absorbing excess liquid from the periphery. The prepared DMEK tissue while reattached on the posterior aspect of the cornea was subjected to slit-lamp biomicroscopic and specular microscopic examinations. In case of proper preparation of DMEK tissue and suitability of endothelial cells for DMEK surgery, the tissue was distributed for transplantation. At the time of surgery, trypan blue staining was performed by the cornea surgeon before DM trephination. Duration of DMEK tissue preparation, uneventful preparation, post-preparation endothelial cell loss, surgeon's satisfaction, and intra- and post-operative attachment of the tissue were investigated variables in this study. Over a 2-year period, 28 DMEK tissues were successfully prepared from 30 corneas using our new technique. The median duration of DMEK preparation was 100 s. The area of endothelial cell loss was 1.8% ± 1.2% centrally and 6.2% ± 1.8% in areas close to the forceps contacts. Seven out of the 28 (25%) DMEK tissues were ruined by surgeons during surgery and the overall rate of re-bubbling in 19 transplanted DMEK tissues was 16%. The presented method, which uses no trephining or vital dye, has simplified and expedited the preparation process of pre-stripped DMEK tissue in the eye banks.

Single versus double pass technique for preparation of ultrathin Descemet's stripping automated endothelial keratoplasty tissues from donated whole eyes.

This study was conducted to analyze the preoperative thickness profile and endothelial rating of ultrathin Descemet's stripping automated endothelial keratoplasty (UT-DSAEK) tissues prepared with a single versus double microkeratome pass from donated whole eyes and corresponding eye bank postoperative results. Microkeratome-assisted UT-DSAEK tissues were prepared from freshly donated whole eyes with single-pass (SP) and double-pass (DP) technique in the Central Eye Bank of Iran. Preoperative thickness profiles and endothelial cell densities of UT-DSAEK tissues were obtained from optical coherence tomography and specular microscopy, respectively, and compared between groups. Corneal perforation rates during the eye bank preparation and postoperative reports of transplanted UT-DSAEK tissues were also compared. Over a 15-month period, 342 UT-DSAEK tissues were prepared: 248 via SP and 94 with DP technique. Mean donor corneal central thickness was 610 ± 58 µm with SP and 790 ± 100 µm with DP technique. Mean central thickness of UT-DSAEK tissues was not statistically different between the groups (84.8 ± 11.0 µm with SP and 85.1 ± 10.5 µm with DP technique, P = 0.857). Mean increase of UT-DSAEK thickness from central to pericentral and peripheral cornea was not significantly different with both techniques. Mean differences between thicknesses of 2 pericentral locations and between those of 2 peripheral locations were not statistically different in the study groups. Corneal perforation of 1.6 and 1.1% occurred in SP and DP groups, respectively. Failed graft was reported 6 months postoperatively in 4 (1.6%) cases with SP and in 1 (1.1%) case with DP technique. Preoperative thickness profiles of UT-DSAEK tissues prepared from donated whole eyes via SP technique were not significantly different from those prepared with DP, showing a symmetric increase of thickness towards peripheral locations.

Preparation and Thickness Profile of Endothelial Keratoplasty Lenticules from Donated Whole Eyes with Previous Photorefractive Keratectomy.

PURPOSE: To describe the preparation and thickness profiles of endothelial keratoplasty lenticules harvested from eyes with previous photorefractive keratectomy (PRK). METHODS: Donor whole eyes that underwent PRK were subjected to microkeratome-assisted dissection for Descemet stripping automated endothelial keratoplasty. Specular microscopy and Visante optical coherence tomography were performed on precut corneas. Endothelial cell indices and thickness profiles of endothelial keratoplasty lenticules were statistically analyzed. Postoperative reports for transplanted lenticules were recorded. RESULTS: Over a 6-month period, 2,929 whole eyes from 1,471 donors were screened for PRK. Twenty-five (0.85%) eyes from 14 donors were diagnosed with disciform haziness due to prior PRK and were used uneventfully for preparation of endothelial keratoplasty lenticules. Mean endothelial cell count was 3164.6 ± 311.0/mm2 and mean central posterior lenticule thickness was 128 ± 34 µm. Posterior lenticules revealed an increase in thickness from the central to peripheral cornea (mean increase of 26.2 µm at pericentral and 90.4 µm at peripheral locations). Mean increase in thickness was statistically different between two peripheral locations (74.5 µm vs. 108.1 µm, P = 0.047). Postoperative reports of transplanted lenticules revealed no posterior flap detachment or loss of clarity at least three months after the surgery. CONCLUSION: PRK donor whole eyes are potential sources for preparation of microkeratome-assisted thin endothelial keratoplasty lenticules with a high endothelial cell count. Although an asymmetric and significant increase in thickness was present at the peripheral cornea, neither attachment nor clarity of transplanted lenticules was affected by variations in thickness of precut corneas.

Measurements of donor endothelial keratoplasty lenticules prepared from fresh donated whole eyes by using ultrasound and optical coherence tomography.

This study was conducted to analyze the profile and thickness of endothelial keratoplasty lenticules prepared from fresh donated whole eyes with Visante optical coherence tomography (V-OCT) compared to measurements obtained from ultrasound pachymetry (USP) at the Central Eye Bank of Iran. Microkeratome-assisted precut corneas were prepared for Descemet stripping automated endothelial keratoplasty by using standard eye bank protocol. Central posterior lenticule thickness (CPLT) on fresh whole eye, before excising corneoscleral disc and transferring to Optisol-GS, was measured by USP. V-OCT was used to measure central, paracentral, and midperipheral thicknesses of lenticules after transferring the tissue to Optisol-GS. Chi Square and Bonferroni tests were respectively used to uncover the differences between the USP and V-OCT measurements and also the thickness profile of lenticules. Postoperative reports for the entire transplanted lenticules were recorded. Accordingly, on evaluation of 312 enrolled precut corneas, CPLT measurements by V-OCT versus USP were statistically different (mean: 136 µm vs 165 µm, respectively; P = 0.008). Thickness profile of the posterior lenticules revealed increased thickness from the central to the peripheral parts of the cornea (mean increase of 16 µm at the pericentral and 64.2 µm at the peripheral locations, respectively); however, the increase in the thickness was relatively symmetrical. Postoperative reports of transplanted lenticules were unremarkable, since there were no posterior flap detachments. In essence, V-OCT measurements of microkeratome-assisted precut lenticules prepared from fresh donated whole eyes averaged 29 µm thinner than USP measurements and revealed a significant but symmetric increase of thickness towards the peripheral parts of the corneas. However, the variation in the thickness profile did not affect the attachment or the clarity of transplanted precut lenticlues.

Trends in Indications and Techniques of Corneal Transplantation in Iran from 2006 to 2013; an 8-year Review.

PURPOSE: To report changing trends in indications and techniques of corneal transplantation in Iran. METHODS: We reviewed eye bank records of 47,129 patients who had undergone corneal transplantation between 2006 and 2013 at different eye centers throughout Iran. RESULTS: The most common indication for corneal transplantation was keratoconus (KCN, n = 21,350 eyes, 45.3%), followed by bullous keratopathy (BK, n = 8,566 eyes, 18.2%), corneal opacity and scar (COS, n = 7,158 eyes, 15.2%), graft failure (n = 3,252 eyes, 6.9%), corneal dystrophies (n = 2,553 eyes, 5.4%), and infectious keratitis (n = 2,238 eyes, 4.7%). Over the study period, there was a significant increase in the frequency of BK (P = 0.001) and graft failure (P = 0.025), and a significant decrease in the relative frequency of COS (P = 0.012). The prevalence of KCN (P = 0.172), infectious keratitis (P = 0.107), and corneal dystrophies (P = 0.836) remained unchanged. The most common technique of corneal transplantation was penetrating keratoplasty (PKP, n = 33,476 eyes, 71.0%), followed by deep anterior lamellar keratoplasty (DALK, n = 8,363 eyes, 17.7%), Descemet's stripping automated endothelial keratoplasty (DSAEK, n = 3,516 eyes, 7.5%), tectonic (n = 1752, 3.7%), and keratolimbal allograft (KLAL, n = 19 eyes, 0.1%). Regarding the shift in surgical techniques, a significant increase was observed in DSAEK (P < 0.001), whereas PKP was significantly decreased (P = 0.005) over the 8-year period. No significant change was seen in the rates of DALK (P = 0.354), tectonic graft (P > 0.999) and KLAL (P = 0.151). CONCLUSION: KCN was the most common indication and PKP was the most prevalent technique used for corneal transplantation. Significant changes in surgical techniques were observed over the past 8 years; DSAEK demonstrated an increasing trend while PKP showed a decrease.

Comparing quantitative and qualitative indices of the donated corneas maintained in Optisol-GS with those kept in Eusol-C.

This study was conducted to compare the quantitative and qualitative indices of the donated corneas maintained in either Optisol-GS or Eusol-C storage media. In an ante-grade single blind study, two corneas from each donor with a death to preservation time of less than 30 h and with a minimum of "an apparent good cornea rating" were maintained in corneal storage media; randomly one in Optisol-GS and the other in Eusol-C. Slit-lamp biomicroscopic and specular microscopic examinations were performed on days 1 and 7. The results of the qualitative and quantitative indices and the final cornea rating were recorded. Statistical analyses were performed to evaluate any differences between the two media. 180 corneas from 90 donors with an age range of 29.3 ± 22.4 years were allocated in two groups: 90 corneas in Optisol-GS and the other 90 in Eusol-C. Five corneas in Optisol-GS and four corneas in Eusol-C were excluded from the study due to lack of specular images. There was no significant change of endothelial rating from day 1 to day 7 between two media (P = 0.175). As the maintenance time of the donated corneas increased, no significant difference was noted between the two groups in terms of endothelial indices, stromal edema, and Descemet's folding. This study shows no superiority between Eusol-C and Optisol-GS in short term preservation of donated corneas.

Preparation of pre-cut corneas from fresh donated whole globes for Descemet's stripping automated keratoplasty: 3-year results at the Central Eye Bank of Iran.

To describe the technique and the results of the preparation of pre-cut corneas for Descemet's stripping automated endothelial keratoplasty (DSAEK) during a 3-year period at the Central Eye Bank of Iran (CEBI). The method of preparation of pre-cut corneas from donated whole globes at the CEBI is described and the frequency and percentage of pre-cut corneas prepared for DSAEK, between April 2009 and March 2012, are specified. Moreover, post-operative reports are reviewed for any complaints about using pre-cut tissues for DSAEK. Out of the 1,518 donated whole globes appropriate for DSAEK, 1,478 (97.4 %) pre-cut corneas were successfully prepared. The method of preparation failed in 40 (2.6 %) cases. Based on the eye bank post-operative reports, thickness of pre-cut tissues for DSAEK was deemed unacceptable in only 6 (0.4 %) cases prior to surgery; five of these were too thick and one was too thin. Preparation of pre-cut corneas, for DSAEK from donated whole globes, in the CEBI is a safe and easy method, with very good preservation of endothelial cells after the preparation of the pre-cut corneas and reduced risks from corneal manipulation.

Screening of donated whole globes for photorefractive keratectomy.

PURPOSE: To introduce a screening method for diagnosis of photorefractive keratectomy (PRK) in donated whole globes. METHODS: Donated whole globes with either no or an unknown history of refractive surgery on family interview were evaluated in the Central Eye Bank of Iran. After disinfection of globes by immersion in 3% povidone iodine for 3 minutes, followed by irrigation with 0.9% normal saline, gross inspection and slit-lamp biomicroscopic examination of the corneas were performed. Corneas with a disciform, round-shaped hazy area within the central 8 mm of the cornea, suggestive of previous PRK (case), and corneas without this criterion (control) were excised and sent for histopathology. Then, sensitivity and specificity of the screening method were calculated by analyzing the histopathology results of case and control corneas. RESULTS: Twelve thousand five hundred fifty-four donated whole globes from 6832 donors were examined. Twenty case corneas from 10 donors and 54 controls from 27 donors were evaluated by histopathology. Age and sex of the donor population in both case and control groups were comparable with the entire donor population. PRK was confirmed by histopathology in all 20 case corneas but not in any of the control corneas. Both the sensitivity and specificity of this screening method were 100%. CONCLUSIONS: Gross inspection of donated whole globes after exposure to povidone iodine followed by slit-lamp biomicroscopic examination, with particular attention to finding a midperipheral-to-midperipheral disciform hazy area within the central 8 mm of cornea, is a safe, simple, and inexpensive screening method, with high sensitivity and specificity for diagnosing PRK.

Specular microscopic features of corneal endothelial vacuolation.

PURPOSE: To introduce a specular microscopic reference image for endothelial vacuolation in donated corneas. METHODS: Two corneas from a donor with diffuse, round to oval dark areas at the endothelial level on slit lamp biomicroscopy and one normal-appearing donor cornea underwent specular microscopy, histopathologic evaluation and transmission electron microscopy. RESULTS: Specular microscopy of the two corneas with abnormal-looking endothelium revealed large numbers of dark, round to oval structures within the endothelium in favor of endothelial vacuolation. Light microscopy disclosed variable sized cyst-like structures within the cytoplasm. Transmission electron microscopy showed electron-lucent and relatively large-sized intracytoplasmic vacuoles. These features were not observed in the endothelium of the normal cornea. CONCLUSION: The specular microscopic features of endothelial vacuolation in donated corneas were confirmed by light microscopy and transmission electron microscopy, therefore the specular image may be proposed as a reference to eye banks.