Although the existence of the limbal stem cell (LSC) niche is accepted, precise knowledge of its three-dimensional (3D) architecture remains incomplete. The LSC niche was explored on freshly excised and organ-cultured corneoscleral rims from human donors (n = 47), pigs (n = 15) and mice (n = 27) with full-field optical coherence microscopy (FFOCM). Limbal crypt features were detected in 90% of organ-cultured human corneoscleral rims, extending between the palisades of Vogt as radially oriented rectangular (74% of eyes) and/or rounded (23% of eyes) forms, often branching off to, or becoming interconnected by, sub-scleral radially or circumferentially oriented crypts (in 56% of eyes). Mean crypt volume represented 16% of sampled limbal volume on the vertical axis and 8% on the horizontal axis. In pigs, palisades were finer and crypts wider with relatively uniform distribution around the eye, and radial orientation, connecting to numerous narrow criss-crossing invaginations beneath the scleral surface. In mice, only a circumferential limbal trough was detected. Mean crypt volume represented 13% of sampled limbal volume in humans and 9% in pigs. FFOCM combined with fluorescence, and confocal fluorescence microscopy, showed presence of p63-α+ cells and cytokeratin-3+ cells in the limbal crypts. To assess colony forming efficiency (CFE), limbal epithelial cells were cultured at low density with mitomycin-arrested 3T3 feeders. CFE increased with limbal crypt volume and was not significantly decreased in organ-cultured cornea, despite degradation of the epithelial roof, suggesting that stem cells remain protected at the base of crypts during organ culture. CFE in human samples was significantly greater than in pig, and CFE in mouse was zero. Crypt architecture in the three species appears associated with eye exposure to light. LSC density increased with percentage limbal volume occupied by crypts.

We uncover the significance of a previously unappreciated structural feature in corneal stroma, important to its biomechanics. Vogt striae are a known clinical indicator of keratoconus, and consist of dark, vertical lines crossing the corneal depth. However we detected stromal striae in most corneas, not only keratoconus. We observed striae with multiple imaging modalities in 82% of 118 human corneas, with pathology-specific differences. Striae generally depart from anchor points at Descemet’s membrane in the posterior stroma obliquely in a V-shape, whereas in keratoconus, striae depart vertically from posterior toward anterior stroma. Optical coherence tomography shear wave elastography showed discontinuity of rigidity, and second harmonic generation and scanning electron microscopies showed undulation of lamellae at striae locations. Striae visibility decreased beyond physiological pressure and increased beyond physiological hydration. Immunohistology revealed striae to predominantly contain collagen VI, lumican and keratocan. The role of these regions of collagen VI linking sets of lamellae may be to absorb increases in intraocular pressure and external shocks.

AimTo assess the influence of graft thickness and regularity on visual recovery and postoperative wavefront aberrations after endothelial keratoplasty (EK).Methods150 EKs performed in eyes with corneal endothelial disorders and no other ocular comorbidities, preoperative and postoperative assessment with spectral domain optical coherence tomography and postoperative assessment with whole eye wavefront aberrometry were retrospectively analysed. Eyes were classified into five groups: Descemet Membrane Endothelial Keratoplasty (DMEK), nanothin Descemet Stripping Automated Endothelial Keratoplasty (DSAEK) (15–49 µm), ultrathin DSAEK (50–99 µm), thin DSAEK (100–149 µm) and conventional DSAEK (150–250 µm).ResultsThe preoperative diagnosis was Fuchs dystrophy in 139 eyes (92.7%). The graft thickness measured after graft deswelling was in average 74 µm with a mean coefficient of variation of 17%. The average follow-up time was 32 months. The mean spectacle-corrected logarithm of minimum angle of resolution visual acuity improved from 0.76 (20/116) before surgery to 0.14 (20/27) at last follow-up visit. No significant differences in final visual acuity were found between the five groups. The time to reach 20/40 vision was significantly shorter in the DMEK and nanothin DSAEK groups compared with the remaining three DSAEK groups. No significant differences in postoperative aberrometry measurements were found between the five groups. Shorter time to reach 20/40 visual acuity was associated with better preoperative visual acuity and thinner graft. Higher final vision improvement was associated with poorer preoperative visual acuity. Higher postoperative high-order aberrations were associated with poorer preoperative visual acuity.ConclusionThe main advantage of DMEK and nanothin DSAEK over thicker DSAEKs was the rapidity of visual recovery. Final quality of vision was not influenced by graft thickness and regularity.

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We aimed to evaluate efficiency and safety of transplantation of limbal stem cells (LSC) cultured on human amniotic membrane with no feeders and to compare cultured LSC with limbal tissue transplantation. Thirty eyes with stage III LSC deficiency were treated with autologous (autoLSC) or allogeneic (alloLSC) cultured LSC transplantation (prospective phase II clinical trial; average follow-up time, 72 months) or autologous (autoLT) or allogeneic (alloLT) limbal tissue transplantation (retrospective control group; average follow-up time, 132 months) between 1993 and 2014. The 5-year graft survival defined by absence of recurrence of the clinical signs of limbal deficiency was 71% for autoLSC, 0% for alloLSC, 75% for autoLT, and 33% for alloLT. Visual acuity improved by 9.2 lines for autoLSC and 3.3 lines for autoLT. It decreased by 0.7 lines for alloLSC and 1.9 lines for alloLT. Adverse events were recorded in 1/7 autoLSC, 7/7 alloLSC, 6/8 autoLT, and 8/8 alloLT patients. Corneal epithelial defect was the only adverse event recorded after autoLSC, whereas severe sight-threatening adverse events were recorded in the remaining three groups. Compared with failed grafts, successful grafts featured greater decrease in fluorescein staining, greater superficial vascularization-free corneal area, lower variability of the corneal epithelial thickness, and higher corneal epithelial basal cell density. Autologous cultured LSC transplantation was associated with high long-term survival and dramatic improvement in vision and was very safe. Autologous limbal tissue transplantation resulted in similar efficiency but was less safe. Cadaver allogeneic grafts resulted in low long-term success rate and high prevalence of serious adverse events. STEM CELLS TRANSLATIONAL MEDICINE 2019;8:1230-1241 LESSONS LEARNED• Transplantation of autologous cultured limbal stem cells was associated with high long-term survival, dramatic improvement in vision, and high safety. • Transplantation of autologous limbal tissue resulted in similar efficiency but lower safety.• Cadaver allogeneic grafts resulted in low long-term success rate and high prevalence of serious adverse events. SIGNIFICANCE STATEMENTAutologous cultured limbal stem cell transplantation was associated with high long-term survival, vision improvement, and safety, whereas limbal grafts featured lower safety and allogeneic grafts featured low success rate and serious adverse events.

The purpose of this study was to analyze the role of corneal epithelial thickness (ET) mapping provided by spectral domain optical coherence tomography in the diagnosis of ocular surface disorders (OSDs) involving the corneal epithelium.Design: This was a retrospective comparative study.Methods: Institutional settings are as follows. Study population includes 303 eyes with an OSD and 55 normal eyes (controls). Observation procedures include spectral domain optical coherence tomography with epithelial mapping in the central 6 mm. Main outcome measures include ET map classification (normal, doughnut, spoke-wheel, localized/diffuse, and thinning/thickening patterns) and ET data and statistics (minimum, maximum, and SD). A quantitative threshold was determined with receiver operating curves to distinguish pathological from normal corneas. Sensitivity and specificity of classification and quantitative data were calculated using all eyes to assess the ability to distinguish corneas with a given corneal disorder from other conditions.Results: Classification of full agreement between 3 readers was obtained in 75.4% to 99.4% of cases. Main OSD features were keratoconus (135 eyes), doughnut pattern (sensitivity/specificity = 56/ 94%), and max-min ET $ 13 mm (84/43%); limbal deficiency (56 eyes), spoke-wheel pattern (66/98%), and max-min ET $ 14 mm (91/59%); epithelial basement membrane dystrophy (55 eyes), inferior thickening pattern (55/92%), and central ET . 56 mm (53/ 81%); dry eye (21 eyes), superior thinning pattern (67/88%), and minimal ET # 44 mm (86/48%); pterygium (10 eyes), nasal thickening pattern (100/86%), and nasal ET . 56 mm (80/71%); and in situ carcinoma (11 eyes), max ET . 60 mm (91/60%), and ET SD .5 mm (100/58%). Conclusions:The epithelial map pattern recognition combined with quantitative analysis of ET is relevant for the diagnosis of OSDs and for distinguishing various OSDs from each other. Deep learning analysis of big data could lead to the fully automated diagnosis of these disorders.

Objective Big bubble (BB)-deep anterior lamellar keratoplasty (DALK) has become the reference transplantation technique for corneal stromal disorders. Type 1 BB is the desired aspect but it is not constant. We aimed to determine the predictive factors of type 1 BB success. Methods Observational cohort study including 77 consecutive eyes of 77 patients undergoing DALK by one surgeon at a single reference center without any selection. Clinical and spectral domain optical coherence tomography data were collected pre- and postoperatively. Results Stromal scars were found in 91.8% of cases and were located in the anterior (90.9%), mid (67.5%), and posterior (36.4%) stroma. Type 1 BB (49.3% of cases) was significantly associated with the absence of scars in the posterior stroma, stage 1–3 keratoconus, and deep trephination. Among eyes with posterior scars, type 1 BB was associated with higher minimal corneal thickness, maximum-minimum corneal thickness < 220 μm, and diagnosis other than keratoconus. Eyes with type 1 BB featured significantly thinner residual stromal bed (22 ± 8 µm versus 61 ± 28 µm), thinner corneas at 12, 24, and 36 months, and better visual acuity at 12 months compared with eyes with no type 1 BB. Conversely, no significant differences between both groups were observed for graft survival, visual acuity at 24 and 36 months, and endothelial cell density at 12 and 36 months. Conclusion OCT assessment before DALK is useful for choosing trephination depth that should be as deep as possible and for looking for posterior scars. The BB technique may not be the most appropriate method in keratoconus with posterior scars. Follow-up data do not support the need for conversion to penetrating keratoplasty when type 1 BB cannot be obtained nor does it support the need for performing a penetrating keratoplasty as a first-choice procedure in eyes with posterior stromal scars.