Nanohydroxyapatite Coating Attenuates Fibrotic and Immune Responses to Promote Keratoprosthesis Biointegration in Advanced Ocular Surface Disorders.

Keratoprosthesis (KPro) implantation is frequently the only recourse for patients with severe corneal disease. However, problems arise due to inadequate biointegration of the KPro, particularly the PMMA optical cylinder, such as tissue detachment, tissue melting, or eye-threatening infection in the interface. Here, using the AuroKPro as a model prosthesis, a surface functionalization approach─coating the optical cylinder with nanohydroxyapatite (nHAp)─was trialed in rabbit eyes with and without a proceeding chemical injury. In chemically injured eyes, which simulated total limbal epithelial stem cell deficiency, clear benefits were conferred by the coating. The total modified Hackett-McDonald score and area of tissue apposition differences 12 weeks after implantation were 5.0 and 22.5%, respectively. Mechanical push-in tests revealed that 31.8% greater work was required to detach the tissues. These differences were less marked in uninjured eyes, which showed total score and tissue apposition differences of 2.5 and 11.5%, respectively, and a work difference of 23.5%. The improved biointegration could be contributed by the attenuated expression of fibronectin (p = 0.036), collagen 3A1 (p = 0.033), and α-smooth muscle actin (p = 0.045)─proteins typically upregulated during nonadherent fibrous capsule envelopment of bioinert material─adjacent to the optical cylinders. The coating also appeared to induce a less immunogenic milieu in the ocular surface tissue, evidenced by the markedly lower expression of tear proteins associated with immune and stimulus responses. Collectively, the level of these tear proteins in eyes with coated prostheses was 1.1 ± 13.0% of naïve eyes: substantially lower than with noncoated KPros (246.5 ± 79.3% of naïve, p = 0.038). Together, our results indicated that nHAp coating may reduce the risk of prosthesis failure in severely injured eyes, which are representative of the cohort of KPro patients.

Corneal Guttae After Descemet Membrane Endothelial Keratoplasty.

PURPOSE: The aim of this study was to report on the occurrence of corneal guttae after Descemet membrane endothelial keratoplasty (DMEK). METHODS: In this retrospective case series, 13 eyes of 13 patients who underwent DMEK at 2 tertiary referral centers between 2007 and 2021 (average available follow-up 73 ± 52 months, range 18-174 months) and showed corneal guttae during postoperative examinations were included. Eye bank images were retrospectively reviewed. RESULTS: Occurrence of guttae was observed by specular microscopy in 13 eyes. In 11 cases, presence of guttae was confirmed by confocal microscopy and in 1 case by histology. Five eyes showed an increase in guttae density during the postoperative course. Surgery indications were Fuchs endothelial corneal dystrophy (n = 11), pseudophakic bullous keratopathy (n = 1), and DMEK graft failure after allograft rejection (n = 1); the latter eye had shown no signs of guttae after primary DMEK. Two eyes with guttae required a repeat DMEK due to graft failure. At the last available follow-up, all 11 remaining eyes had clear corneas and 10 eyes had a best-corrected visual acuity of ≥0.9 (decimal). During donor cornea processing in the eye bank, no guttae were observed on the donor tissue. CONCLUSIONS: Corneal guttae can occur after DMEK including in eyes operated for indications other than Fuchs endothelial corneal dystrophy and most likely guttae were present on the donor graft but were not detectable by routine slit-lamp and light microscopy evaluation in the eye bank. Postoperative guttae density varies among patients and especially small isolated guttae do not seem to affect clinical outcomes.

Bowman Layer Onlay Grafting as a Minimally Invasive Treatment for the Most Challenging Cases in Keratoconus.

PURPOSE: To analyze the clinical outcomes after Bowman layer (BL) onlay grafting for the treatment of progressive, advanced keratoconus. DESIGN: Prospective, interventional case series. METHODS: Twenty-one eyes underwent BL onlay grafting. After removing the epithelium, a single or double BL graft was "stretched" onto the corneal surface, allowed to dry-in, and a soft bandage lens was placed until the graft was re-epithelialized. Best spectacle- and/or best contact lens-corrected visual acuity (BSCVA/BCLVA), corneal tomography, and postoperative complication rates were analyzed for the total group and 2 subgroups (group 1: preoperative maximum keratometry [Kmax] <69 diopters [D; n = 7); group 2: preoperative Kmax ≥69 D [n = 14]). Follow-up ranged from 6 to 36 months (mean 21 ± 11 months). RESULTS: All 21 surgeries were uneventful. Overall, Kmax changed from 76 ± 12 D preoperatively to 72 ± 9 D at 6 to 36 months postoperatively (P = .015). Kmax decreased by 6 D in group 2 (P = .002) but did not change in group 1. Average BSCVA remained stable for group 1 and improved from preoperatively 0.8 ± 0.4 to 0.4 ± 0.2 logarithm of the minimum angle of resolution postoperatively in group 2 (P = .032); BCLVA remained stable (P > .05). Within the first postoperative weeks, 2 eyes required BL graft repositioning after inadvertent bandage lens removal and 4 eyes underwent BL retransplantation for incomplete re-epithelialization. One eye underwent BL regrafting 12 months postoperatively after traumatic corneal erosion. All eyes showed a completely re-epithelialized graft at the last available follow-up. CONCLUSIONS: BL onlay grafting is a completely extraocular, minimally invasive surgical technique, providing up to -6 D of corneal flattening in eyes with advanced progressive keratoconus, allowing for continued (scleral) contact lens wear and therefore preserving the BCLVA.

P19-A118†Bowman layer onlay: preparation and transplantation (BLOT).

PURPOSE: With the introduction of Bowman layer onlay transplantation (BLOT), the need for BL transplants increases.In this study, the clinical outcomes of BLOT are described and the results of three different BL graft preparation methods are evaluated: manually (m-BL), femtosecond laser-assisted (fs-BL), and femtosecond laser-assisted followed by excimer laser (fs/ex-BL). METHOD: Twenty-one eyes with advanced progressive keratoconus underwent BLOT with m-BL. Best spectacle- and/or best contact lens-corrected visual acuity (BSCVA/BCLVA), corneal tomography, and complications were recorded. Follow-up ranged from 6-36 months with a mean follow-up time of 21±12 months.To evaluate BL preparation methods, Descemet membrane-denuded donor corneas (n=41) were used (n=2 for m-BL, n=18 for fs-BL and n=21 for fs/ex-BL). For fs-BL, corneas were placed on an artificial anterior chamber and different depth cuts were performed with decreasing decrements starting from 30 µm (diameter 9.0 mm). For fs/ex-BL, a superficial flap of 80 µm was created by the femtosecond laser (FEMTO-LDV Z8, Ziemer). Followed by residual stroma ablation by excimer laser (Schwind Amaris 750S) with increasing increments. Grafts were analyzed visually, and graft thickness regularity was evaluated by histological analysis and Transmission Electron Microscopy (TEM). RESULTS: All twenty-one surgeries could be performed without intraoperative complications. Average maximum keratometry changed from 75.8±12D preoperatively to 72.2±9D at the last available follow-up (n=21, P<0.05), and BSCVA/BCLVA improved. Five patients required a regraft; four of those because of a graft detachment within one week.Evaluation of BL-preparation methods: Fs-BL preparation was successful until 14µm cuts (success rate: 12 out of 14, 86%). Fs/ex-BL graft preparation was most successful after an 80µm cut by femtosecond laser with subsequent 60µm ablation by excimer laser (success rate: 15 out of 21, 71%). After the femtosecond laser cut, traces of the femtosecond laser treatment were visible on the flap. While m-BL showed long protruding stromal fibers, they were shorter in fs-BL and absent in fs/ex-BL. CONCLUSION: BL-onlay grafting may be a feasible surgical technique, providing on average -3D of corneal flattening in eyes with advanced progressive keratoconus, while improving patient's visual acuity.Fs-BL and fs/ex-BL preparation may be faster alternatives to manual BL graft preparation.

P07-A139†Guttae in corneal donor tissue.

PURPOSE: To report on the occurrence of guttae in corneal donor tissue. MATERIAL & METHODS: Retrospective database study of discard reasons for corneal donor tissue at Amnitrans EyeBank Rotterdam (AER) for the period from January 2019 to December 2021 and the outcome of an eight-question survey sent to European Eye Bank Association corresponding members addressing the occurrence of corneal guttae and the practice pattern regarding donor tissue with guttae. RESULTS: Between 2019 and 2021 6039 donor corneas were processed at AER. Average discard rate because of guttae in this period was 9 (±4)% (n=552). Most corneas were discarded because of guttae at first evaluation (8%, n=481). Monthly discard rate because of guttae ranged from 3% to 19%. Yearly discard rates related to corneal guttae were 10 (±3)%, 8 (±3)% and 11 (±5)% in 2019, 2020 and 2021, respectively. Average endothelial cell density (ECD) at the first evaluation from 2019-2021 was 2486 (±93) cells/mm2, with average monthly ECD ranging from 2343 to 2642 cells/mm2.Twenty-nine eye banks completed the survey, including 4 located outside Europe. 70% reported a guttae-related discard rate of ≤4. The types of microscope used for the evaluation, the geographical location and the number of guttae permitted do not seem to influence the discard rates. 13 eye banks permitted 0 guttae while 10 banks accepted between 1-10 guttae.The 16 eye banks that responded 'no' to the question whether the contralateral cornea of a guttae-cornea was automatically discarded did report a lower guttae-related discard rate than the other eye banks. CONCLUSION: The high variability of the discard rate due to guttae in donor corneas (ranging from <1% and >12%) is an indication that it is not always easy to detect guttae in donor corneas. Although transplanting corneal grafts with guttae does not necessarily mean that a re-transplantation will be needed on the short term, a vital method to unequivocally determine the presence of guttae in the eye bank seems essential to prevent unnecessary waste of suspect tissue and unnecessary re-surgeries.

Effects of Rho-Associated Kinase (Rock) Inhibitors (Alternative to Y-27632) on Primary Human Corneal Endothelial Cells.

(1) Rho-associated coiled-coil protein kinase (ROCK) signaling cascade impacts a wide array of cellular events. For cellular therapeutics, scalable expansion of primary human corneal endothelial cells (CECs) is crucial, and the inhibition of ROCK signaling using a well characterized ROCK inhibitor (ROCKi) Y-27632 had been shown to enhance overall endothelial cell yield. (2) In this study, we compared several classes of ROCK inhibitors to both ROCK-I and ROCK-II, using in silico binding simulation. We then evaluated nine ROCK inhibitors for their effects on primary CECs, before narrowing it down to the two most efficacious compounds-AR-13324 (Netarsudil) and its active metabolite, AR-13503-and assessed their impact on cellular proliferation in vitro. Finally, we evaluated the use of AR-13324 on the regenerative capacity of donor cornea with an ex vivo corneal wound closure model. Donor-matched control groups supplemented with Y-27632 were used for comparative analyses. (3) Our in silico simulation revealed that most of the compounds had stronger binding strength than Y-27632. Most of the nine ROCK inhibitors assessed worked within the concentrations of between 100 nM to 30 µM, with comparable adherence to that of Y-27632. Of note, both AR-13324 and AR-13503 showed better cellular adherence when compared to Y-27632. Similarly, the proliferation rates of CECs exposed to AR-13324 were comparable to those of Y-27632. Interestingly, CECs expanded in a medium supplemented with AR-13503 were significantly more proliferative in (i) untreated vs. AR-13503 (1 µM; * p < 0.05); (ii) untreated vs. AR-13503 (10 µM; *** p < 0.001); (iii) Y-27632 vs. AR-13503 (10 µM; ** p < 0.005); (iv) AR-13324 (1 µM) vs. AR-13503 (10 µM; ** p < 0.005); and (v) AR-13324 (0.1 µM) vs. AR-13503 (10 µM; * p < 0.05). Lastly, an ex vivo corneal wound healing study showed a comparable wound healing rate for the final healed area in corneas exposed to Y-27632 or AR-13324. (4) In conclusion, we were able to demonstrate that various classes of ROCKi compounds other than Y-27632 were able to exert positive effects on primary CECs, and systematic donor-match controlled comparisons revealed that the FDA-approved ROCK inhibitor, AR-13324, is a potential candidate for cellular therapeutics or as an adjunct drug in regenerative treatment for corneal endothelial diseases in humans.

Early and late-onset cell migration from peripheral corneal endothelium.

In this study we describe peripheral corneal endothelial cell migration in vitro in the absence and presence of a ROCK-inhibitor. For this study, 21 corneal endothelial graft rims, with attached trabecular meshwork (TM), were prepared from Descemet membrane-endothelial cell sheets by 6.5 mm trepanation. For the initial proof-of-concept, 7 outer graft rims were cultured in a thermo-reversible hydrogel matrix for up to 47 days. To assess the effect of a ROCK-inhibitor, 14 paired outer rims were cultured either with or without ROCK-inhibitor for up to 46 days. At the end of culture, tissue was retrieved from the hydrogel matrix and examined for cell viability and expression of different endothelial cell markers (ZO-1, Na+/K+-ATPase, NCAM, glypican, and vimentin). All cultured rims remained viable and displayed either single regions (n = 5/21) or collective areas (n = 16/21) of cell migration, regardless of the presence or absence of ROCK-inhibition. Migration started after 4±2 days and continued for at least 29 days. The presence of ROCK-inhibitor seemed to contribute to a more regular cell morphology of migrating cells. In addition, 7 outer rims demonstrated a phenotypically distinct late-onset but fast-growing cell population emerging from the area close to the limbus. These cells emerged after 3 weeks of culture and appeared less differentiated compared to other areas of migration. Immunostaining showed that migrated cells maintained the expression patterns of endothelial cell markers. In conclusion, we observed 2 morphologically distinct migrating cell populations with the first type being triggered by a broken physical barrier, which disrupted contact inhibition and the second, late-onset type showing a higher proliferative capacity though appearing less differentiated. This cell subpopulation appeared to be mediated by stimuli other than loss of contact inhibition and ROCK-inhibitor presence. Further exploration of the differences between these cell types may assist in optimizing regenerative treatment options for endothelial diseases.

Long-Term Outcomes of Descemet Membrane Endothelial Keratoplasty: Effect of Surgical Indication and Disease Severity.

PURPOSE: The aim of this study was to evaluate clinical outcomes and graft survival in a large patient cohort up to 10 years after Descemet membrane endothelial keratoplasty (DMEK) based on surgical indication and Fuchs endothelial corneal dystrophy (FECD) severity. METHODS: The cohort in this retrospective study included 750 eyes that underwent DMEK for FECD (86%), bullous keratopathy (BK, 9%), and other indications (5%). Based on the modified Krachmer grading, 186 eyes (29%) had moderate FECD (Krachmer grade 3-4) and 440 eyes (68%) had advanced FECD (Krachmer grade 5-6). Main outcome measures were best-corrected visual acuity (BCVA), central corneal thickness, endothelial cell density (ECD), endothelial cell loss (ECL), postoperative complication rate, and graft survival. RESULTS: The mean 10-year BCVA was 0.08 ± 0.28 logMAR (n = 96), and the BCVA remained stable between 5 and 10 years postoperatively (all P > 0.05). The mean ECL at 5 and 10 years was 56% ± 17% (n = 460) and 66% ± 14% (n = 96), respectively ( P < 0.05). The 10-year graft survival rate for the total cohort was 0.85 (95% confidence interval, 0.82-0.89). BCVA and ECD differed at 1 to 5 years between eyes with different surgical indications (all P < 0.05), and FECD eyes had higher graft survival compared with BK eyes (0.90 vs. 0.60, P = 0.001). Moderate FECD eyes had better BCVA, ECD, and graft survival outcomes than advanced FECD eyes (all P < 0.05). Minor graft detachment ( P = 0.04) and lower donor ECD ( P = 0.01) were related to lower 10-year ECD. CONCLUSIONS: Long-term outcomes and graft survival after DMEK were better in FECD eyes than BK eyes and were also related to FECD disease severity. Eyes operated for moderate FECD showed the highest graft survival probability and excellent long-term outcomes.

Impact of Fuchs Endothelial Corneal Dystrophy Severity on Scheimpflug-Derived Parameters After Descemet Membrane Endothelial Keratoplasty.

PURPOSE: The aim of this study was to evaluate how Scheimpflug-derived parameters of eyes with Fuchs endothelial corneal dystrophy (FECD) are influenced by Descemet membrane endothelial keratoplasty (DMEK) depending on FECD severity and the presence of subclinical edema. METHODS: A retrospective cohort study including 115 eyes (115 patients) that underwent DMEK for FECD and a control group of 27 eyes with nonpathological corneas was conducted. Preoperative and 6 months postoperative Scheimpflug imaging was used to analyze pachymetry, presence of tomographic features (loss of isopachs/displacement of the thinnest point/focal posterior depression), and corneal backscatter. FECD severity was based on the modified Krachmer scale and the absence/presence of subclinical edema. RESULTS: Scheimpflug-derived pachymetry, tomographic, and corneal backscatter parameters were correlated with FECD severity, and all changed from preoperatively to postoperatively (all P < 0.05). Postoperative central corneal thickness, anterior and posterior corneal backscatter, and presence of focal posterior depression remained different from the control group (all P < 0.05). Of eyes without preoperative clinical edema (n = 75), 18.7% showed 0 or 1 tomographic feature (no edema group) and 82.4% had 2 or 3 features (subclinical edema group). Compared with the control group, postoperative best-corrected visual acuity for the "no edema" group did not differ (0.03 ± 0.12 vs. -0.02 ± 0.08 logarithm of the minimum angle of resolution, P = 0.150) but was worse for the subclinical edema group (0.06 ± 0.08 vs. -0.02 ± 0.08 logarithm of the minimum angle of resolution, P = 0.001). CONCLUSIONS: For eyes without preoperative edema, more parameters reversed back to 'normal' levels than for eyes with (sub)clinical edema. Although most analyzed parameters correlated with FECD severity, corneal tomography might be best suited for objective grading of disease severity to aid in surgical decision-making.

[Evolution in corneal transplantation techniques]. / Evolutie in hoornvliestransplantatie.

Over the past twenty years, there have been many developments in the treatment of corneal diseases, especially in the field of corneal transplantation. By targeting treatments solely to the affected layers of the cornea, the procedures have become less invasive and the cornea recovers significantly faster. Despite the success of these new surgical techniques, new (cell) regenerative therapies are emerging.

Selective endothelial removal: A case series of a phase I/II surgical trial with long-term follow up.

Peters anomaly is a congenital condition which results in a central corneal opacity from birth. Selective Endothelial Removal (SER) is a novel surgical technique and a form of regenerative therapy, which encourages clearance of the central corneal opacity by the patient's own corneal endothelial cells, and it may potentially be beneficial for the treatment of Peters anomaly. We have performed a phase I/II surgical trial, evaluating the safety of SER in four eyes (three patients) with Peters Anomaly. These patients underwent SER at between 9 and 39 months of age, each demonstrating clearance of central corneal opacities and improvements in vision post-operatively. No complications occurred in any of these eyes, at a minimal post-operative follow-up duration of 48 months. We conclude that SER for Peters anomaly is a safe surgical procedure. While encouraging efficacy outcomes have been observed, these findings should be further evaluated in a larger scale Phase II/III surgical trial.

32†Corneal guttae after descemet membrane endothelial keratoplasty (DMEK).

PURPOSE: To report on the occurrence of corneal guttae after Descemet membrane endothelial keratoplasty (DMEK) in eyes operated on for Fuchs endothelial corneal dystrophy (FECD). MATERIAL AND METHODS: Case series of 10 eyes of 10 patients operated on for FECD at a tertiary referral center between 2008 and 2019. Average patient age was 61±12 years and 3 patients were female and 6 were male. Five patients were phakic and 4 pseudophakic. Average donor age was 67±9 years. RESULTS: During routine postoperative consultation, specular microscopy images showed suspected recurrence of guttae in 10 eyes after DMEK. Presence of guttae was subsequently confirmed in 9 cases by confocal microscopy and in one case by histology. Six out of 10 patients (60%) had undergone bilateral DMEK, but all only showed recurrence of guttae in one eye. In 9 eyes guttae recurred after primary DMEK, while in one eye recurrence was after a re-DMEK that has been performed 56 months after the first DMEK with no signs of guttae after primary DMEK. Suspected guttae were visible on specular microscopy images already at 1 month after DMEK in most cases.No guttae had been noted during donor cornea processing in the eye bank. Preoperative donor endothelial cell density (ECD) had been 2643±145 cells/mm2 and 1-year postoperative ECD was 1047±458 cells/mm2 (n=8). CONCLUSION: Recurrence of guttae after DMEK is most likely due to guttae on the donor graft that were not detectable by routine slit-lamp and light-microscopy evaluation in the eye bank. Better screening methods for guttae detection need to be developed for eye banks to avoid releasing tissue for transplantation that contains guttae or is prone for postoperative guttae formation.

24†Effect of graft preparation techniques on clinical outcomes after Descemet membrane endothelial keratoplasty (DMEK).

PURPOSE: To compare the clinical outcomes after Descemet membrane endothelial keratoplasty (DMEK) for grafts prepared by the manual no-touch peeling technique and grafts prepared by a modified liquid bubble technique. MATERIAL AND METHODS: For this study, 236 DMEK grafts were included that were prepared at Amnitrans EyeBank Rotterdam by experienced eye bank personnel. 132 grafts were prepared by using the 'no-touch' DMEK preparation technique and 104 grafts by using a modified liquid bubble technique. The liquid bubble technique was modified to render it a no-touch technique while maintaining the ability to save the anterior donor button as a potential Deep Anterior lamellar keratoplasty (DALK) or Bowman layer (BL) graft. DMEK surgeries were performed at Melles Cornea Clinic Rotterdam by experienced DMEK surgeons. All patients underwent DMEK for Fuchs endothelial dystrophy. Average patient age was 68 (±10) years and average donor age was 69 (±9) years with no difference between the two groups. Endothelial cell density (ECD) was evaluated after graft preparation by light microscopy in the eye bank and at 6-month postoperatively by specular microscopy. RESULTS: Endothelial cell density (ECD) decreased from 2705 (±146) cells/mm2 (n=132) before to 1570 (±490) cells/mm2 (n=130) at 6 months postoperatively for grafts prepared by the no-touch technique. For grafts prepared by the modifiedliquid bubble technique, ECD decreased from 2627 (±181) cells/mm2 (n=104) before to 1553 (±513) cells/mm2 (n=103) after surgery. Postoperative ECD did not differ for grafts prepared by the two techniques (P=0.79). Central corneal thickness (CCT) decreased from 660 (±124) µm to 513 (±36) µm postoperatively in the no-touch group and from 684 (±116) µm to 515 (±35) µm postoperatively in the modified liquid bubble group, with no postoperative CCT difference between groups (P=0.59). In total 3 eyes underwent re-surgery within the study period (n=2 (1.5%) in the no-touch group, n=1 (1.0%) in the liquid bubble group; P=0.71) and 26 eyes required a re-bubbling procedure for incomplete graft adherence (n=16 (12%) in the no-touch group, n=10 (10%) in the liquid bubble group; P=0.37). CONCLUSION: Clinical outcomes after DMEK are comparable for grafts prepared by either the manual no-touch peeling technique or the modified liquid bubble technique. While both techniques are safe and useful techniques to prepare DMEK grafts, the modified liquid bubble technique offers advantages for corneas with scars.

37†Descemet membrane endothelial keratoplasty (DMEK): 10-year clinical outcomes and graft survival.

PURPOSE: To evaluate graft survival and clinical outcomes up to 10 years after Descemet membrane endothelial keratoplasty (DMEK). SETTING/VENUE: Retrospective cohort study conducted at the Netherlands Institute for Innovative Ocular Surgery. METHODS: 750 consecutive DMEK eyes, not including the very first 25 DMEK eyes that constitute the technique learning curve, were included. Main outcome parameters (survival, best-corrected visual acuity (BCVA), central endothelial cell density (ECD)) was evaluated up to 10-years postoperatively and postoperative complications were documented. Outcomes were analyzed for the entire study group and separately for the subgroup of the first 100 DMEK eyes. RESULTS: For the subgroup of 100 DMEK eyes, 82% and 89% reached a BCVA of ≥20/25 (Decimal VA ≥0.8) at 5- and 10 years postoperatively, respectively, and preoperative donor ECD decreased by 59% at 5 years and 68% at 10 years postoperatively. Graft survival probability for the first 100 DMEK eyes was 0.83 [95% Confidence Interval (CI), 0.75-0.92] and 0.79 [95% CI, 0.70 -0.88] at 5- and 10-years postoperatively, respectively. For the total study group, clinical outcome in terms of BCVA and ECD were comparable, but graft survival probability was significantly higher at 5- and 10-year postoperatively. CONCLUSIONS: Most eyes operated in the pioneering phase of DMEK showed excellent and stable clinical outcomes with a promising graft longevity over the first decade after surgery. The increase in DMEK experience resulted in a lower graft failure rate and positively affected longer-term graft survival probability.

New developments in corneal endothelial cell replacement.

Corneal transplantation is currently the most effective treatment to restore corneal clarity in patients with endothelial disorders. Endothelial transplantation, either by Descemet membrane endothelial keratoplasty (DMEK) or by Descemet stripping (automated) endothelial keratoplasty (DS(A)EK), is a surgical approach that replaces diseased Descemet membrane and endothelium with tissue from a healthy donor eye. Its application, however, is limited by the availability of healthy donor tissue. To increase the pool of endothelial grafts, research has focused on developing new treatment options as alternatives to conventional corneal transplantation. These treatment options can be considered as either 'surgery-based', that is tissue-efficient modifications of the current techniques (e.g. Descemet stripping only (DSO)/Descemetorhexis without endothelial keratoplasty (DWEK) and Quarter-DMEK), or 'cell-based' approaches, which rely on in vitro expansion of human corneal endothelial cells (hCEC) (i.e. cultured corneal endothelial cell sheet transplantation and cell injection). In this review, we will focus on the most recent developments in the field of the 'cell-based' approaches. Starting with the description of aspects involved in the isolation of hCEC from donor tissue, we then describe the different natural and bioengineered carriers currently used in endothelial cell sheet transplantation, and finally, we discuss the current 'state of the art' in novel therapeutic approaches such as endothelial cell injection.

Fuchs endothelial corneal dystrophy: The vicious cycle of Fuchs pathogenesis.

Fuchs endothelial corneal dystrophy (FECD) is the most common primary corneal endothelial dystrophy and the leading indication for corneal transplantation worldwide. FECD is characterized by the progressive decline of corneal endothelial cells (CECs) and the formation of extracellular matrix (ECM) excrescences in Descemet's membrane (DM), called guttae, that lead to corneal edema and loss of vision. FECD typically manifests in the fifth decades of life and has a greater incidence in women. FECD is a complex and heterogeneous genetic disease where interaction between genetic and environmental factors results in cellular apoptosis and aberrant ECM deposition. In this review, we will discuss a complex interplay of genetic, epigenetic, and exogenous factors in inciting oxidative stress, auto(mito)phagy, unfolded protein response, and mitochondrial dysfunction during CEC degeneration. Specifically, we explore the factors that influence cellular fate to undergo apoptosis, senescence, and endothelial-to-mesenchymal transition. These findings will highlight the importance of abnormal CEC-DM interactions in triggering the vicious cycle of FECD pathogenesis. We will also review clinical characteristics, diagnostic tools, and current medical and surgical management options for FECD patients. These new paradigms in FECD pathogenesis present an opportunity to develop novel therapeutics for the treatment of FECD.

Automated Clinical Assessment of Corneal Guttae in Fuchs Endothelial Corneal Dystrophy.

PURPOSE: To describe the validation and implementation of an automated system for the detection and quantification of guttae in Fuchs endothelial corneal dystrophy (FECD). DESIGN: Observational reliability study. METHODS: Patients with FECD underwent retroillumination corneal photography, followed by determination of the distributions and sizes of corneal guttae by an automated image analysis algorithm. Performance of the automated system was assessed via (1) validation against manual guttae segmentation, (2) reproducibility studies to ensure consistency, and (3) evaluation for agreement with the Krachmer scale. It was then deployed to perform large-scale guttae assessment with anatomic subregion analysis in a batch of 40 eyes. RESULTS: Compared to manual segmentation, the automated system was reasonably accurate in identifying the correct number of guttae (mean count of 78 guttae per 1 × 1 mm test frame, overestimation: +10 per frame), but had a tendency to significantly overestimate guttae size (mean guttae size 1073 µm2, overestimation: +255 µm2). Automated measurements of guttae counts and sizes were reproducible within a 1% discrepancy range across repeat intra-eye assessments. Automated guttae counts, interguttae distances, and density of interguttae gaps lesser than 40 µm (ie, D40 density) were highly correlated with the Krachmer scale (P < .001 for all). Large-scale guttae assessment demonstrated the automated system's potential to selectively identify a region of the corneal endothelium most affected by densely packed guttae. CONCLUSIONS: Automated guttae assessment facilitates the precise identification and quantification of guttae characteristics in FECD patients. This can be used clinically as a personalized descemetorrhexis zone for Descemet stripping only and/or Descemet membrane transplantation.

Regenerative capacity of the corneal transition zone for endothelial cell therapy.

The corneal endothelium located on the posterior corneal surface is responsible for regulating stromal hydration. This is contributed by a monolayer of corneal endothelial cells (CECs), which are metabolically active in a continuous fluid-coupled efflux of ions from the corneal stroma into the aqueous humor, preventing stromal over-hydration and preserving the orderly arrangement of stromal collagen fibrils, which is essential for corneal transparency. Mature CECs do not have regenerative capacity and cell loss due to aging and diseases results in irreversible stromal edema and a loss of corneal clarity. The current gold standard of treatment for this worldwide blindness caused by corneal endothelial failure is the corneal transplantation using cadaveric donor corneas. The top indication is Fuchs corneal endothelial dystrophy/degeneration, which represents 39% of all corneal transplants performed. However, the global shortage of transplantable donor corneas has restricted the treatment outcomes, hence instigating a need to research for alternative therapies. One such avenue is the CEC regeneration from endothelial progenitors, which have been identified in the peripheral endothelium and the adjacent transition zone. This review examines the evidence supporting the existence of endothelial progenitors in the posterior limbus and summarizes the existing knowledge on the microanatomy of the transitional zone. We give an overview of the isolation and ex vivo propagation of human endothelial progenitors in the transition zone, and their growth and differentiation capacity to the corneal endothelium. Transplanting these bioengineered constructs into in vivo models of corneal endothelial degeneration will prove the efficacy and viability, and the long-term maintenance of functional endothelium. This will develop a novel regenerative therapy for the management of corneal endothelial diseases.