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The feasibility of a 220 - 330 GHz zero order axicon generated Bessel beam for corneal water content was explored. Simulation and experimental data from the 25-degree cone angle hyperbolic-axicon lens illuminating metallic spherical targets demonstrate a monotonically decreasing, band integrated, backscatter intensity for increasing radius of curvature from 7 - 11 mm, when lens reflector and optical axis are aligned. Further, for radii >= 9.5 mm, maximum signal was obtained with a 1 mm transverse displacement between lens and reflector optical axes arising from spatial correlation between main lobe and out of phase side lobes. Thickness and permittivity parameter estimation experiments were performed on an 8 mm radius of curvature, 1 mm thick fused quartz dome over a 10 mm axial span. Extracted thickness and permittivity varied by less than ~ 25 $μ$m and 0.2 respectively after correction for superluminal velocity. Estimated water permittivity and thickness of water backed gelatin phantoms showed significantly more variation due to a time varying radius of curvature., 11 pages; submitted to IEEE Transactions on Terahertz Science and Technology

To highlight the progress and future direction of stem-cell based regenerative therapies for the treatment of corneal disease.Corneal stem cell-based therapies, such as limbal stem cell transplantation, corneal stromal stem cell transplantation, endothelial stem cell transplantation, and stem cell-derived extracellular vesicles have demonstrated promising results in the laboratory. Although most are still in preclinical development or early phase clinical trials, these stem cell-based therapies hold potential to facilitate tissue regeneration, restore native function, and inhibit pathologic disease processes such as fibrosis, inflammation, and neovascularization.Stem cell-based therapy offers a promising therapeutic option that can circumvent several of the challenges and limitations of traditional surgical treatment. This concise review summarizes the progress in stem-cell based therapies for corneal diseases along with their history, underlying mechanisms, limitations, and future areas for development.

The aim of this study was to investigate basal epithelial cell morphology (CM) in the central cornea and limbal areas of eyes with limbal stem cell deficiency (LSCD).This was a prospective, cross-sectional comparative study. We developed a CM scoring system based on basal epithelial cell phenotypes graded from 0 (normal) to 3 (severe morphologic alterations); this system was evaluated by 2 independent masked observers. The CM score was compared with the LSCD clinical score, mean best-corrected visual acuity, and in vivo laser scanning confocal microscopy parameters used to stage LSCD (ie, basal epithelial cell density, basal epithelial thickness, and subbasal corneal nerve fiber length density).One hundred sixty-eight eyes with LSCD and 63 normal eyes were included. Compared with the control group, the LSCD group had significantly higher mean (±SD) CM scores in the central cornea (1.8 ± 0.7 vs. 0.5 ± 0.4, respectively; P = 0.01) and limbal areas (1.6 ± 0.2 vs. 1.3 ± 0.0, respectively; P0.05). The mean CM score in the central cornea was positively correlated with the clinical score ( P0.01, r = 0.66) and negatively correlated with the best-corrected visual acuity ( P0.01, r = 0.42). The CM scores were positively correlated with all other in vivo laser scanning confocal microscopy parameters in the central cornea and limbal areas (all P0.001).Basal epithelial CM is altered in the central cornea and limbus of eyes with LSCD and thus can be used to stage the clinical severity of the disease.

With significant advancement and development of extracellular vesicle (EV)-based therapies, there is a growing need to understand how their storage affects their physical and functional characteristics. EVs were isolated from the conditioned medium of a corneal stromal stem cell line (imCSSC) using Total Exosome isolation kit (TEI) and ultracentrifugation (UC) combined protocol. Purified EVs were stored at 4°C, − 80°C, room temperature (RT) after lyophilization with or without trehalose for 4 weeks. EVs stored at − 80°C and RT (lyophilization with trehalose) demonstrated a comparable morphology, while the freeze-dried samples without trehalose showed aggregation and degradation under a transmission electron microscope (TEM). Lyophilized samples without trehalose demonstrated a decreased particle concentration, recovery rate and protein concentration, which was remediated by the addition of trehalose. EVs stored at − 80℃ showed no change in the protein expression of CD9, CD63, and CD81. Regardless of the storage condition, all EV samples investigated reduced inflammation, as well as inhibited expression of fibrotic markers in vitro. Lyophilization of EVs with trehalose was a feasible storage method that retained the physical property and in vitro biological activities of EVs after 4 weeks of storage, while − 80°C offered the best retention of imCSSC-derived EV physical properties. For the first time, this data demonstrated a practical and translatable method for the storage of CSSC-derived EVs for clinical use. Graphical Abstract

This article presents measurements of the permittivity of gelatin hydrogels between 220 and 330 GHz. Hydrated gelatin was treated as a binary mixture of free water and a compound consisting of water bound to collagen. Submillimeter-wave reflectometry was used to estimate the hydrated gelatin permittivity, hydrated gelatin density, and free-water volume fraction in phantoms composed of 62, 67, 72, and 77% water by weight. A hydrated dry/wet density ratio of 0.335 was validated with optical-coherence tomography. A constant nonfreezing bound-water mass of 0.6 g/g was observed and confirmed with differential-scanning calorimetry. Good agreement between results from different modalities supports the dielectric spectroscopy methods and data analysis. Depending on the hydrodynamics at the sample/air interface, measurements indicate a bound-water compound permittivity of 3.77−j2.52 to 3.95−j2.49—contrasting the pure-water average permittivity of 5.16–j5.63. The loss related to bound water was much higher than anticipated and characterization will help reduce uncertainty in measurements of gelatin hydrogel-based tissue phantoms; particularly corneal phantoms where adjacent free water creates complex hydration gradients. This is the first known, submillimeter-wave, frequency domain measurement of complex permittivity of the bound-water component in solid, proteinaceous matter.

To evaluate safety and efficacy of autologous serum eye drops (AS) in the treatment of limbal stem cell deficiency (LSCD) associated with glaucoma surgery.Retrospective case series of eyes with glaucoma surgery-induced LSCD treated with AS. Diagnosis of LSCD was confirmed by anterior segment optical coherence tomography, in vivo confocal microscopy, and/or impression cytology. Limbal stem cell deficiency severity was staged using a clinical scoring system (2-10 points). Outcome measures were changes (≥2 points) of the LSCD score and best-corrected visual acuity (BCVA) from the baseline to the last follow-up.Thirteen eyes of 12 consecutive patients treated with 50% AS for at least 3 months were included. The mean age was 78.9±7.5 years and the mean duration of AS use was 20.9±16.8 months. Indications of AS included LSCD progression in eight eyes (61.5%) and visual axis threatening in five eyes (38.5%). The mean LSCD score at baseline (6.7±1.6) was similar to that at last follow-up (6.5±2.2, P =0.625). Two eyes (15.4%) showed improvement, nine eyes (69.2%) were stable, and two eyes (15.4%) worsened. The mean baseline BCVA (0.89±0.64 logMAR) was similar to the mean final BCVA (1.05±0.63 logMAR, P =0.173). There were no serious adverse complications related to AS.AS appears to be well tolerated and may stabilize the progression of LSCD with limited effects. A larger study is necessary to confirm the findings.

Purpose To review the published literature on the visual acuity results and complications of different surgical techniques for intraocular lens (IOL) implantation in the absence of zonular support. Methods Peer-reviewed literature searches were conducted last in PubMed and the Cochrane Library in July 2019. The searches yielded 734 citations of articles published in English. The panel reviewed the abstracts of these mostly retrospective case series studies, and 45 were determined to be relevant to the assessment objectives. Three articles were rated as level II evidence, and 42 articles were rated as level III evidence. Results Eight different types of IOL fixation techniques with at least 6-month follow-up were evaluated: anterior chamber IOL (ACIOL), iris-claw IOL, retropupillary iris-claw IOL, 10-0 polypropylene iris-sutured posterior chamber IOL (PCIOL), 10-0 polypropylene scleral-sutured PCIOL, 8-0 polypropylene scleral-sutured PCIOL, CV-8 polytetrafluoroethylene, and intrascleral haptic fixation (ISHF). Eight articles reported data comparing 2 techniques. The 45 studies had insufficient statistical power to compare the techniques conclusively. A qualitative analysis of similar types showed that trends in visual acuity outcomes were not inferior to those of ACIOL implantation, but the severity of preoperative pathologic features was not controlled for. Compared with ACIOL, complications of cystoid macular edema were higher in 10-0 polypropylene iris-sutured PCIOL and 8-0 polypropylene scleral-sutured PCIOL. Non–anterior chamber IOL techniques were less likely to report chronic uveitis. Chronic glaucoma was highest in the 8-0 polypropylene scleral-sutured PCIOL group. Although retinal detachment was infrequent overall, it was twice as common in both iris- and scleral-sutured PCIOLs (except CV-8 polytetrafluoroethylene suture) compared with nonsutured methods: ACIOL, iris-clipped IOL, and ISHF PCIOL. Conclusions The evidence reviewed shows no superiority of any single IOL implantation technique in the absence of zonular support. The various techniques seem to have equivalent visual acuity outcomes and safety profiles. Each technique has its own profile of inherent risk of postoperative complications. Surgeons must educate patients on the importance of close, long-term follow-up as a result of the uncertain nature of these techniques. Large prospective studies are needed to confirm the long-term complication profiles of these various IOL implantation techniques.

ObjectiveUsing anterior segment optical coherence tomography (AS-OCT), we investigated the epithelial thickness (ET) of the central cornea and limbal regions in patients with limbal stem cell deficiency (LSCD) as a diagnostic and staging parameter.DesignProspective, cross-sectional study.MethodsThe central corneal epithelium thickness (CET) and maximum limbal epithelium thickness (mLET) were measured in the superior, inferior, nasal, and temporal limbus on AS-OCT images of the normal and eyes with LSCD. CET was obtained by 1-point (OCT-CET1) and 3-point measurement (OCT-CET3). The values of OCT-CET1 and OCT-CET3 were compared to the CET obtained with invivo confocal microscopy (IVCM-CET).ResultsSixty-eight eyes of 50 patients with LSCD and 52 eyes of 34 normal subjects were included. The mean (±standard deviation) OCT-CET3 was 55.0 ± 3.0μm (range, 50.6-62.0μm) in the control group and 41.6 ± 10.8μm (range, 0-56.3μm) in the LSCD group (P < .001). OCT-CET3 had a better correlation with IVCM-CET (r= 0.91) than did OCT-CET1 (r= 0.87, P= .001). The degree of reduction in OCT-CET3 increased in more advanced clinical stages of LSCD (all P < .001). The OCT-CET3 cutoff value that suggests LSCD was 46.6μm. Compared with the control group, the LSCD group had decreases in mLET in all 4 limbal regions (all P < .001). The sensitivity and specificity of OCT-CET3 is the highest among all mLET in detecting LSCD.ConclusionsBoth CET and mLET were thinner in patients with LSCD than in normal subjects. OCT-CET3 appears to be a reliable parameter to confirm LSCD when there is clinical suspicion.

Purpose In recent decades, the medical and surgical treatment of limbal stem cell deficiency (LSCD) has evolved significantly through the incorporation of innovative pharmacological strategies, surgical techniques, bioengineering, and cell therapy. With such a wide variety of options, there is a need to establish a global consensus on the preferred approaches for the medical and surgical treatment of LSCD. Methods An international LSCD Working Group was established by the Cornea Society in 2012 and divided into subcommittees. Four face-to-face meetings, frequent email discussions, and teleconferences were conducted since then to reach agreement on a strategic plan and methods after a comprehensive literature search. A writing group drafted the current study. Results A consensus in the medical and surgical management of LSCD was reached by the Working Group. Optimization of the ocular surface by eyelid and conjunctival reconstruction, antiinflammatory therapy, dry eye and meibomian gland dysfunction treatment, minimization of ocular surface toxicity from medications, topical medications that promote epithelialization, and use of a scleral lens is considered essential before surgical treatment of LSCD. Depending on the laterality, cause, and stage of LSCD, surgical strategies including conjunctival epitheliectomy, amniotic membrane transplantation, transplantation of limbal stem cells using different techniques and sources (allogeneic vs. autologous vs. ex vivo-cultivated), transplantation of oral mucosal epithelium, and keratoprosthesis can be performed as treatment. A stepwise flowchart for use in treatment decision-making was established. Conclusions This global consensus provides an up-to-date and comprehensive framework for the management of LSCD.

Purpose To characterize the clinical presentation of limbal stem cell deficiency (LSCD) associated with glaucoma surgeries. Methods This is a retrospective cross-sectional study of patients with LSCD and glaucoma who presented to the Stein Eye Institute at the University of California, Los Angeles, between 2009 and 2018. Patients who underwent trabeculectomy and/or aqueous shunt surgery were included. The severity of LSCD was staged using global consensus guidelines and a clinical scoring system, and basal epithelial cell density was measured by in vivo confocal microscopy. Anatomic locations of glaucoma and non-glaucoma surgeries, locations of LSCD, and severity of LSCD were compared. Results Fifty-one eyes of 41 patients with LSCD associated with glaucoma surgery were included in this study. LSCD in these patients uniquely featured sectoral replacement of corneal epithelium by conjunctival epithelium, without corneal neovascularization or pannus. The sites of glaucoma surgery strongly correlated with the locations of LSCD (P = 0.002). There was a trend toward increased severity of LSCD in eyes with 2 or more glaucoma surgeries as compared to eyes with 1 glaucoma surgery, although the difference did not reach statistical significance (P = 0.3). Use of topical glaucoma medications correlated with LSCD severity, while the impact of antimetabolites did not reach statistical significance. The location of glaucoma drainage surgery is correlated with the location of LSCD. Conclusions LSCD associated with glaucoma surgery has clinical features distinct from LSCD resulting from other etiologies. Further study is required to delineate the full impact of glaucoma surgery on limbal stem cell function and survival.

Introduction: Human corneal stromal stem cells (CSSCs) have gained increasing attention in the treatment of corneal stromal scars. In view of this, the preparation and storage of CSSCs are critical...

Purpose To evaluate the efficacy and safety of topical cenegermin (recombinant human nerve growth factor) in patients with neurotrophic keratopathy. Design Multicenter, randomized, double-masked, vehicle-controlled trial. Participants Patients with neurotrophic persistent epithelial defect with or without stromal thinning. Methods The NGF0214 trial, conducted among 11 sites in the United States, randomized 48 patients 1:1 to cenegermin 20 μg/ml or vehicle eye drops, 6 drops daily for 8 weeks of masked treatment. Follow-up was 24 weeks. Safety was assessed in all patients who received study drug. Efficacy was assessed by intention to treat. Main Outcome Measures The primary end point was healing of the neurotrophic lesion (persistent epithelial defect or corneal ulcer) after 8 weeks of masked treatment. Masked central readers measured neurotrophic lesions in randomized clinical pictures, then assessed healing status conventionally ( Results Conventional assessment of corneal healing showed statistically significant differences at week 8: compared to 7 of 24 vehicle-treated patients (29.2%), 16 of 23 cenegermin-treated patients (69.6%) achieved less than 0.5 mm of lesion staining (+40.4%; 95% confidence interval [CI], 14.2%–66.6%; P = 0.006). Conservative assessment of corneal healing also reached statistical significance at week 8: compared to 4 of 24 vehicle-treated patients (16.7%), 15 of 23 cenegermin-treated patients (65.2%) achieved 0 mm of lesion staining and no other residual staining (+48.6%; 95% CI, 24.0%–73.1%; P Conclusions Cenegermin treatment showed higher rates of corneal healing than vehicle in neurotrophic keratopathy associated with nonhealing corneal defects.

Publisher Copyright: IEEE A system concept for online alignment verification of millimeter-wave, corneal reflectometry is presented. The system utilizes beam scanning to generate magnitude-only reflectivity maps of the cornea at 650 GHz and compares these images to a precomputed/measured template map to confirm/reject sufficient alignment. A system utilizing five off-axis parabolic mirrors, a thin film beam splitter, and two-axis galvanometric mirror was designed, simulated, and evaluated with geometric and physical optics. Simulation results informed the construction of a demonstrator system which was tested with a reference reflector. Similarity metrics computed with the aligned template and 26 misaligned positions, distributed on a 0.5 mm x 0.5 mm x 0.5 mm mesh, demonstrated sufficient misalignment detection sensitivity in 23 out of 26 positions. The results show that positional accuracy on the order of 0.5 mm is possible using 0.462 mm wavelength radiation due to the perturbation of coupling efficiency via beam distortion and beamwalk-off.

Etodolac is an FDA-approved nonsteroidal anti-inflammatory drug (NSAID) used to treat a variety of inflammatory diseases. The drug is administered as a racemate (50/50 mixture of R- and S- enantiomers), however, studies have shown that the two enantiomers have distinct biologic and pharmacokinetic differences. Wnt signaling, which plays key roles in cell proliferation, polarity, and differentiation, has been shown to be inhibited by R-etodolac; however, comparative analyses of R- and S-etodolac in this function have not been conducted. We used in silico molecular docking and TOPflash functional biologic assays to compare R- and S-enantiomers effect on Wnt signaling inhibition. Further, we used a cultivated limbal stem epithelial cell (cLSCs) model to investigate enantiospecific changes in the colony-forming efficiency (CFE) of cLSCs. The data shows that R-etodolac is a more potent inhibitor of Wnt signaling. In addition, consistently, while both enantiomers demonstrate a dose-dependent decrease in CFE of cLSCs, R-etodolac is a more potent inhibitor.

PURPOSE The purpose of this study was to evaluate the expression of cytokeratin (K) 13 on the corneal surface and to validate its application in the diagnosis of limbal stem cell deficiency (LSCD). METHODS This prospective comparative study included 26 corneal impression cytology (IC) specimens from patients diagnosed with LSCD. Twenty-three IC specimens from normal donors served as controls. K12 and K13 expression were detected on the IC specimens by immunohistochemistry study. The number of K12+ or K13+ cells in all areas of the IC was quantified using ImageJ software. RESULTS The epithelial cells harvested from IC specimens from control corneas were all K12+. In eyes with LSCD, K13+ and K12+/K13+ cells accounted for 93.8% and 2.6%, respectively, in the cornea. In eyes with sectoral LSCD, the median number of K13+ cells in the clinically affected area was higher than that in the unaffected area (810.0 vs. 115.0 cells/mm2; P < 0.001). No significant correlation was found between the LSCD severity and the number of K12+ cells (r = -0.284, P = 0.16) or K13+ cells (r = -0.011, P = 0.95). The presence of at least 16 K13+ cells/mm2 was suggestive of LSCD. CONCLUSIONS Identification of K13+ cells on IC specimens provides a simple and reliable method to detect conjunctival epithelial cells on the cornea. K13 is a marker for diagnosing LSCD and localizing the involved area in sectoral LSCD.

Limbal epithelial stem/progenitor cells (LSCs) reside in a niche that contains finely tuned balances of various signaling pathways including Wnt, Notch, BMP, Shh, YAP, and TGFβ. The activation or inhibition of these pathways is frequently dependent on the interactions of LSCs with various niche cell types and extracellular substrates. In addition to receiving molecular signals from growth factors, cytokines, and other soluble molecules, LSCs also respond to their surrounding physical structure via mechanotransduction, interaction with the ECM, and interactions with other cell types. Damage to LSCs or their niche leads to limbal stem cell deficiency (LSCD). The field of LSCD treatment would greatly benefit from an understanding of the molecular regulation of LSCs in vitro and in vivo. This review synthesizes current literature around the niche factors and signaling pathways that influence LSC function. Future development of LSCD therapies should consider all these niche factors to achieve improved long-term restoration of the LSC population.

Publisher Copyright: IEEE Absolute thickness and free-water-content gradients in gelatin-based corneal phantoms with physiologically accurate radii of curvature and aqueous backing were extracted via coherent submillimeter-wave reflectometry at 220-330 GHz. Fourier-domain-based calibration methods, utilizing temporal and spatial gating, were developed and yielded peak-to-peak amplitude and phase clutter of 10-3 and 0.1°, respectively, for signal-to-noise ratios (SNRs) between 40 and 50 dB. Total 12 phantoms were fabricated. Calibration methods enabled quantification of target sphericity that strongly correlated with optical-coherence tomography-based sphericity metrics via image segmentation. The extracted free-water volume fraction varied less than 5% in the five phantoms whose fabrication yielded the most spherical geometry. Submillimeter-wave-based thickness accuracy was better than 111 μm (∼λ/9) with an average of 65 μm (∼λ/17) and standard deviation of 44 μm (∼λ/25) for phantoms with physiologically relevant geometry. Monte-Carlosimulations of measurement noise and uncertainty limits agree with the experimental data analysis and indicate a lower thickness accuracy limit of 33 μm, and water-content sensitivities of 0.5% and 11.8% for the anterior and posterior segments, respectively. Numerical analysis suggests that the measurement fidelity was SNR limited and identified optical path length ambiguities within the cornea where a continuum of thickness/water gradient pairs produces statistically insignificant differences in complex reflection coefficient for finite SNR. This is the first known submillimeter-wave measurement technique, which is able to extract both the thickness and water-content gradients from a soft-tissue phantom, with a water backing, without the need for ancillary measurements.

An efficient method for rapid, non-contact scanning of human cornea is presented. The optics utilize two, 101.6-mm diameter off-axis parabolic mirrors fed by a goniometrically scanned, planar mirror. An evaluation system at 650 GHz was built and demonstrated ~ 1.5 mm beam radius on target and ~ 30° x 30° field of view. The system was inspired by confocal laser scanning principles and represents a system design where image acquisition time is limited by SNR, not mechanical translation.

Aims To evaluate the clinical and visual outcomes of fluid-filled scleral lens devices (SL) wear in patients with limbal stem cell deficiency (LSCD). Design Retrospective consecutive case series. Methods 27 eyes with LSCD confirmed by in vivo confocal microscopy at the Stein Eye Institute and fitted with SL were included. Correlations between corrected distance visual acuity (CDVA) and LSCD stage determined by clinical grading were performed between baseline (after the SL fit) and the last follow-up (the time of discontinuation of SL wear or the last visit in eyes in which SL were continued). In a subset of patients that had worsened LSCD while using SL, anterior segment optical coherence tomography (AS-OCT) and anterior segment fluorescein angiogram (AS-FA) were performed. Results Baseline LSCD grading was stage I in 12 eyes (44.4%), stage 2 in 12 eyes (44.4%), and stage III in 3 eyes (11.1%). At the last follow-up, CDVA was improved in 7 eyes (25.9%), remained stable in 13 eyes (48.1%) and decreased in 7 eyes (25.9%, P = 0.16). The LSCD stage was improved in 7 eyes (25.9%), remained stable in 8 eyes (29.6%) and worsened in 12 eyes (44.4%, P = 0.10). AS-OCT and AS-FA, performed in 5 eyes, showed limbal compression and delayed fluorescein filling. Conclusion SL can improve visual acuity and maintain the ocular surface in the majority of eyes. Worsening of the ocular surface might be a result of limbal hypoxia. Close monitoring of SL fit is necessary in these compromised eyes.

Wnt proteins are secreted morphogens that play critical roles in embryonic development, stem cell proliferation, self-renewal, tissue regeneration and remodeling in adults. While aberrant Wnt signaling contributes to diseases such as cancer, activation of Wnt/β-catenin signaling is a target of interest in stem cell therapy and regenerative medicine. Recent high throughput screenings from chemical and biological libraries, combined with improved gene expression reporter assays of Wnt/β-catenin activation together with rational drug design, led to the development of a myriad of Wnt activators, with different mechanisms of actions. Among them, Wnt mimics, antibodies targeting Wnt inhibitors, glycogen-synthase-3β inhibitors, and indirubins and other natural product derivatives are emerging modalities to treat bone, neurodegenerative, eye, and metabolic disorders, as well as prevent ageing. Nevertheless, the creation of Wnt-based therapies has been hampered by challenges in developing potent and selective Wnt activators without off-target effects, such as oncogenesis. On the other hand, to avoid these risks, their use to promote ex vivo expansion during tissue engineering is a promising application., Overview of the Wnt signaling pathway and targets of Wnt activators.

The corneal epithelium is consistently regenerated by limbal stem/progenitor cells (LSCs), a very small population of adult stem cells residing in the limbus. Several Wnt ligands, including Wnt6, are preferentially expressed in the limbus. To investigate the role of Wnt6 in regulating proliferation and maintenance of human LSCs in an in vitro LSC expansion setting, we generated NIH-3T3 feeder cells to overexpress different levels of Wnt6. Characterization of LSCs cultured on Wnt6 expressing 3T3 cells showed that high level of Wnt6 increased proliferation of LSCs. Medium and high levels of Wnt6 also increased the percentage of small cells (diameter ≤ 12 µm), a feature of the stem cell population. Additionally, the percentage of cells expressing the differentiation marker K12 was significantly reduced in the presence of medium and high Wnt6 levels. Although Wnt6 is mostly known as a canonical Wnt ligand, our data showed that canonical and non-canonical Wnt signaling pathways were activated in the Wnt6-supplemented LSC cultures, a finding suggesting that interrelationships between both pathways are required for LSC regulation.

Purpose To compare the outcomes of Descemet membrane endothelial keratoplasty (DMEK) with those of Descemet's stripping endothelial keratoplasty (DSEK) in eyes with prior glaucoma surgery. Design Case-matched retrospective comparative case series. Methods Setting/study population: 46 DMEK procedures were matched with 46 DSEK procedures at a single institution. Observation procedures: cases were matched based on preoperative visual acuity, lens status, and surgical indication. Main outcome measurements: the outcome measurements included visual acuity improvement, primary and secondary graft failure, endothelial rejection, intraocular pressure (IOP) elevation, and the need for additional glaucoma intervention. Results Best-corrected visual acuity (BCVA) improved by −0.89 logMAR in the DMEK group and −0.62 logMAR in the DSEK group (P = 0.005) at 1 year follow-up. Visual acuity was significantly better in the DMEK group at postoperative months 1, 3, and 12 and at last follow-up. The percentage of patients achieving 20/40 or better best-corrected visual acuity was higher in the DMEK group at all time points, notably 47% in the DMEK group versus 15% in the DSEK group at 1 year (P = 0.002). Secondary graft failure was lower in the DMEK group (DMEK 0% vs. DSEK 17%; P = 0.006). Primary graft failure rates and rebubling rates were similar. There were no differences in the rates of postoperative IOP elevation or in the need for additional glaucoma intervention. Conclusions In complex eyes with prior glaucoma surgery, DMEK offers faster visual recovery, better final visual acuity, and a lower rate of secondary graft failure compared to DSEK during the first postoperative year and beyond.

To evaluate in vivo parameters as biomarkers of limbal stem cell function and to establish an objective system that detects and stage limbal stem cell deficiency (LSCD).A total of 126 patients (172 eyes) with LSCD and 67 normal subjects (99 eyes) were included in this observational cross-sectional comparative study. Slit-lamp biomicroscopy, in vivo laser scanning confocal microscopy (IVCM), and anterior segment optical coherence tomography (AS-OCT) were performed to obtain the following: clinical score, cell morphology score, basal cell density (BCD), central corneal epithelial thickness (CET), limbal epithelial thickness (LET), total corneal nerve fiber length (CNFL), corneal nerve fiber density (CNFD), corneal nerve branch density (CNBD), and tortuosity coefficient. Their potential correlations with the severity of LSCD were investigated, and cutoff values were determined.An increase clinical score correlated with a decrease in central cornea BCD, limbal BCD, CET, mean LET, maximum LET, CNFL, CNFD, CNBD, and tortuosity coefficient. Regression analyses showed that central cornea BCD, CET and CNFL were the best parameters to differentiate LSCD from normal eyes (Coef = 3.123, 3.379, and 2.223; all p 0.05). The rank correlation analysis showed a similar outcome between the clinical scores and the central cornea BCD (ρ = 0.79), CET (ρ = 0.82), and CNFL (ρ = 0.71). A comprehensive LSCD grading formula based on a combination of these parameters was established.A comprehensive staging system combining clinical presentation, central cornea BCD, CET, and CNFL is established to accurately and objectively diagnose LSCD and stage its severity.

The purpose of this study was to determine longitudinal trends in prevalence and resistance profiles for infectious keratitis at referral centers in Southern California.Cultured infectious keratitis cases from January 1, 2006, through December 31, 2009, and January 1, 2016, through December 31, 2019, at the University of California, Los Angeles, were evaluated. Outcome measures included microbial isolate prevalence and antibiotic/antifungal susceptibility and resistance patterns.One hundred thirty-nine and 315 culture-positive isolates were obtained between 2006-2009 and 2016-2019, respectively. Gram-positive organisms accounted for 65% (2006-2009) and 74% (2016-2019) of bacterial isolates ( P = 0.076). Staphylococcus infections, the most common gram-positive and bacterial isolate in both study epochs, demonstrated increased prevalence from 2006-2009 to 2016-2019 (41% vs. 53%, P = 0.019). Although coagulase-negative Staphylococcus (CoNS) increased from 40% to 58% ( P = 0.0012), the prevalence of methicillin-resistant Staphylococcus aureus was unchanged (28% vs. 28%, P = 0.99). Pseudomonas aeruginosa , the most common gram-negative organism, demonstrated decreased prevalence from 18% to 10% ( P = 0.027). Candida species comprised 3.5% of culture-positive isolates in both epochs. All gram-positive isolates were susceptible to vancomycin, and all Staphylococcus isolates were susceptible to linezolid. Pseudomonas aeruginosa remained susceptible to tested fluoroquinolones (93%) and aminoglycosides (100%) over time.In southern California between 2006 and 2019, there was a shift toward Staphylococcus species, with increased CoNS, decreased methicillin-sensitive Staphylococcus aureus , and decreased prevalence of P. aeruginosa . Empiric therapy of vancomycin and a fluoroquinolone or aminoglycoside provides effective antibacterial coverage for predominant bacterial species when culture sensitivities are pending.

Pterygium is an ocular surface disorder with high prevalence that can lead to vision impairment. As a pathological outgrowth of conjunctiva, pterygium involves neovascularization and chronic inflammation. Here, we developed a 3D multicellular in vitro pterygium model using a digital light processing (DLP)-based 3D bioprinting platform with human conjunctival stem cells (hCjSCs). A novel feeder-free culture system was adopted and efficiently expanded the primary hCjSCs with homogeneity, stemness and differentiation potency. The DLP-based 3D bioprinting method was able to fabricate hydrogel scaffolds that support the viability and biological integrity of the encapsulated hCjSCs. The bioprinted 3D pterygium model consisted of hCjSCs, immune cells, and vascular cells to recapitulate the disease microenvironment. Transcriptomic analysis using RNA sequencing (RNA-seq) identified a distinct profile correlated to inflammation response, angiogenesis, and epithelial mesenchymal transition in the bioprinted 3D pterygium model. In addition, the pterygium signatures and disease relevance of the bioprinted model were validated with the public RNA-seq data from patient-derived pterygium tissues. By integrating the stem cell technology with 3D bioprinting, this is the first reported 3D in vitro disease model for pterygium that can be utilized for future studies towards personalized medicine and drug screening.

Author(s): Bonnet, Clemence; Vazirnia, Parsia; Deng, Sophie X; Aldave, Anthony J; Miller, Kevin M | Abstract: PurposeTo assess safety and efficacy outcomes of sutured custom silicone artificial iris and intraocular lens implantation combined with penetrating keratoplasty (triple procedure).MethodsProspective consecutive surgical case series of patients who underwent the triple procedure between 2010 and 2019 at Stein Eye Institute, UCLA, followed up for 1 year minimum. Safety outcomes were changes from preoperative to last follow-up in corrected distance visual acuity (CDVA), endothelial cell count, intraocular pressure (IOP), and postoperative complications. Efficacy outcomes included changes in subjective glare (none to severe), cosmetic appearance (worse to very much improved), and visual function as assessed by the Visual Function Questionnaire-25 at 1-year follow-up.ResultsAmong 82 eyes implanted with an artificial iris, 14 eyes (17.1%) underwent the triple procedure. The median follow-up was 18.1 months (range 12.0-54.9 months). The median CDVA improved from 2.0 log of minimum angle of resolution (logMAR) (range 0.9-2.3 logMAR) to 0.7 logMAR (range 0.2-2.6 logMAR) (P = 0.02). Average endothelial cell count decreased 57.6% (P l 0.01). Six eyes (42.9%) experienced IOP elevations, 13 eyes (92.3%) developed iritis, and 11 eyes (78.6%) underwent secondary surgery. Graft rejection or secondary graft failure occurred in 7 eyes each (50.0%). Cosmesis improved in 12 eyes (85.7%; P l 0.01). The Visual Function Questionnaire-25 score improved from 72 to 77 (P l 0.01). Glare symptoms did not change significantly.ConclusionsThe triple procedure was effective at improving CDVA, cosmesis, and quality of life; however, it was associated with frequent postoperative complications, of which iritis, IOP elevation, and secondary graft failure were the most common.

Transplantation of human cultured limbal epithelial stem/progenitor cells (LESCs) has demonstrated to restore the integrity and functionality of the corneal surface in about 76% of patients with limbal stem cell deficiency. However, there are different protocols for the expansion of LESCs, and many of them use xenogeneic products, being a risk for the patients&rsquo, health. We compared the culture of limbal explants on the denuded amniotic membrane in the culture medium&mdash, supplemental hormone epithelial medium (SHEM)&mdash, supplemented with FBS or two differently produced human sera. Cell morphology, cell size, cell growth rate, and the expression level of differentiation and putative stem cell markers were examined. Several bioactive molecules were quantified in the human sera. In a novel approach, we performed a multivariate statistical analysis of data to investigate the culture factors, such as differently expressed molecules of human sera that specifically influence the cell phenotype. Our results showed that limbal cells cultured with human sera grew faster and contained similar amounts of small-sized cells, higher expression of the protein p63&alpha, and lower of cytokeratin K12 than FBS cultures, thus, maintaining the stem/progenitor phenotype of LESCs. Furthermore, the multivariate analysis provided much data to better understand the obtaining of different cell phenotypes as a consequence of the use of different culture methodologies or different culture components.

Background Keratitis-ichthyosis-deafness (KID) syndrome is characterized by a congenital triad of keratitis, ichthyosis, and deafness, and is most commonly associated with mutations in the gap junction protein beta 2 gene (GJB2) on chromosome 13q11-q12. Methods Multimodal anterior segment imaging and genetic testing were used to supplement clinical examination findings in the diagnosis and management of a 12-year-old boy with suspected KID syndrome. Results The patient presented with hearing loss, ichthyosis of the face and extremities, and corneal scarring and keratinization. The corneal limbal stem cell population was found to be normal on in vivo confocal microscopy, whereas the basal epithelium of the cornea demonstrated scarring and areas of cellular loss. Screening of GJB2 revealed a presumed pathogenic heterozygous missense mutation, c.148G>A, confirming the diagnosis of KID syndrome. Conclusions Multimodal imaging including in vivo confocal microscopy suggests that dysfunctional corneal basal epithelium maturation might contribute to the pathophysiology of keratopathy in KID syndrome.

Summary Ex vivo cultured limbal stem/progenitor cells is an effective alternative to other surgical treatments for limbal stem cell deficiency, but a standard xenobiotic-free method for culturing the LSCs in vitro needs to be optimized. Because Wnt ligands are required for LSC expansion and preservation in vitro, to create a small-molecule Wnt mimic, we created a consolidated compound by linking a Wnt inhibitor that binds to the Wnt co-receptor Frizzled to a peptide derived from the N-terminal Dickkopf-1 that binds to Lrp (low-density lipoprotein receptor-related protein) 5/6, another Wnt co-receptor. This Wnt mimic not only enhances cellular Wnt signaling activation, but also improves the progenitor cell phenotype of in vitro cultured limbal epithelial cells. As the maintenance of stem cell characteristics in the process of culture expansion is essential for the success of ocular surface reconstruction, the small molecules generated in this study may be helpful in the development of pharmaceutical reagents for treating corneal wounds., Graphical Abstract, Highlights • MFH-ND is generated by linking a Frizzled inhibitor and an LRP5/6 inhibitor • MFH-ND activates the canonical Wnt pathway by oligomerizing Frizzled and LRP5/6 • MFH-ND improves the stem cell phenotype of cultivated limbal epithelial cells • MFH-ND has therapeutic potential to improve limbal stem cell deficiency treatment, Molecular Biology; Stem Cells Research

IMPORTANCE: Limbal stem cell transplant (LSCT) can be categorized as direct autologous limbal transplant (AULT), direct allogenic limbal transplant (ALLT), cultivated autologous limbal stem cells transplant (cAULT), and cultivated allogenic limbal stem cells transplant (cALLT). To our knowledge, there is no study directly comparing the outcomes and complications of these procedures. OBJECTIVE: To evaluate the outcomes of different LSCT procedures. DATA SOURCE: We searched PubMed, EMBASE, Web of Science, and Cochrane without language filter for peer-reviewed articles about LSCT. The latest search was performed on June 30, 2019. STUDY SELECTION: Clinical studies with the outcome of at least 20 eyes after LSCT were included. Animal studies and studies of other surgical interventions were excluded. DATA EXTRACTION AND SYNTHESIS: Two reviewers independently abstracted the data from each study. Heterogeneity was evaluated with the I(2) statistic, and a meta-analysis was performed using the random-effects model. MAIN OUTCOMES AND MEASURES: Outcome measures included the improvement of ocular surface, visual acuity (VA), and adverse events of recipient eyes and donor eyes. RESULTS: Forty studies (2202 eyes) with a mean (SD) follow-up of 31.3 (20.9) months met the inclusion criteria. The mean (SD) age of study participants was 38.4 (13.1) years, and men accounted for 74%. The number of eyes that underwent AULT, ALLT, cAULT, and cALLT were 505, 742, 771, and 184, respectively. Improvement of the ocular surface was achieved in 74.5% of all eyes, 85.7% of eyes after AULT (95% CI, 79.5%-90.3%), 84.7% after cAULT (95% CI, 77.2%-90.0%), 57.8% after ALLT (95% CI, 49.0%-66.1%), and 63.2% after cALLT (95% CI, 49.3%-75.2%). Autologous limbal transplantation resulted in a greater VA improvement rate (76%) than did the other 3 procedures (cAULT: 56.4%; ALLT: 52.3%; cALLT: 43.3%; all P

The purpose of this study was to evaluate the long-term outcomes of Descemet membrane endothelial keratoplasty (DMEK) in eyes that had previously undergone trabeculectomy and/or drainage device implantation.Retrospective, noncomparative case series.Medical records of 251 consecutive DMEK procedures performed by 1 surgeon (S.X.D.) from 2013 to 2017 were reviewed. Patients with ≥2 years of follow-up were divided into 3 groups: eyes with prior glaucoma surgery (ST), eyes with medically treated glaucoma (MT), and eyes without glaucoma (NG). Main outcomes measured were visual acuity, endothelial cell count (ECC), rates of secondary graft failure (SGF), and postoperative complications.Ninety procedures (87 eyes) met inclusion criteria. The mean follow-up period of all eyes was 38.4 ± 11.2 months (range, 24.2-64.4 months). At last follow-up, the proportion of eyes reaching a vision of ≥20/40 was higher than that before the DMEK procedure in each group (all P.05). The rate of ECC loss was the highest in the ST group compared to that in the MT and NG groups (63.8% vs 47.6% vs 44.0%, respectively; P.05) as well as the rate of SGF (41.6% vs 0% vs 2.4%, respectively; P.05). The rate of SGF of repeat DMEK was higher than that of primary DMEK (P.05). The rates of postoperative complications were similar among all groups (all P.05).In eyes with prior glaucoma surgery, DMEK achieved good long-term visual outcomes but experienced a higher rate of SGF than eyes without such comorbidity.

An accurate diagnosis of limbal stem cell deficiency (LSCD) is the premise of an appropriate treatment; however, there is no consensus about the diagnostic criteria for LSCD. We performed a systematic literature search of the peer-reviewed articles on PubMed, Medline, and Ovid to investigate how LSCD was diagnosed before surgical intervention. The methods used to diagnose LSCD included clinical presentation, impression cytology, and invivo confocal microscopy. Among 131 eligible studies (4054 eyes), 26 studies (459 eyes, 11.3%) did not mention the diagnostic criteria. In the remaining 105 studies, the diagnosis of LSCD was made on the basis of clinical examination alone in 2398 eyes (62.9%), and additional diagnostic tests were used in 1047 (25.8%) eyes. Impression cytology was used in 981 eyes (24.2%), invivo confocal microscopy was used in 29 eyes (0.7%), and both impression cytology and invivo confocal microscopy were used in 37 eyes (0.9%). Our findings suggest that only a small portion of patients underwent a diagnostic test to confirm the diagnosis of LSCD. Treating physicians should be aware of the limitations of clinical examination in diagnosing LSCD and perform a diagnostic test whenever possible before surgical intervention.

PurposeTo grade the severity of limbal stem cell deficiency (LSCD) based on the extent of clinical presentation and central corneal basal epithelial cell density (BCD).MethodsThis is a retrospective observational comparative study of 48 eyes of 35 patients with LSCD and 9 eyes of 7 normal subjects (controls). Confocal images of the central cornea were acquired. A clinical scoring system was created based on the extent of limbal and corneal surface involvement. LSCD was graded as mild, moderate, and severe stages based on the clinical scores. The degree of BCD reduction was given a score of 0 to 3.ResultsCompared with BCD in controls, BCD decreased by 23.0%, 40.4%, and 69.5% in the mild, moderate, and severe stages of LSCD classified by the clinical scoring system, respectively. The degree of BCD reduction was positively correlated with larger limbal and corneal surface involvement and when the central visual axis was affected (all P ≤ 0.0005). Mean corrected distance visual acuity logarithm of the minimum angle of resolution was 0.0 ± 0.0 in control eyes, 0.2 ± 0.5 in mild LSCD, 0.6 ± 0.4 in moderate LSCD, and 1.6 ± 1.1 in severe LSCD (P < 0.0001). There was a significant correlation between a higher clinical score and corrected distance visual acuity logarithm of the minimum angle of resolution (rho = 0.82; P < 0.0001) and a greater decrease in BCD (rho = -0.78; P < 0.0001).ConclusionsA clinical scoring system was developed to assess the extent of clinical presentation of LSCD. A classification system to grade the severity of LSCD can be established by combining the BCD score with the clinical score.

Objective To review the published literature on safety and outcomes of Descemet's stripping endothelial keratoplasty (DSEK) for the surgical treatment of endothelial diseases of the cornea. Design Peer-reviewed literature searches were conducted in PubMed and the Cochrane Library with the most recent search in February 2009. The searches yielded 2118 citations in English-language journals. The abstracts of these articles were reviewed and 131 articles were selected for possible clinical relevance, of which 34 were determined to be relevant to the assessment objectives. Results The most common complications from DSEK among reviewed reports included posterior graft dislocations (mean, 14%; range, 0%–82%), followed by endothelial graft rejection (mean, 10%; range, 0%–45%), primary graft failure (mean, 5%; range, 0%–29%), and iatrogenic glaucoma (mean, 3%; range, 0%–15%). Average endothelial cell loss as measured by specular microscopy ranged from 25% to 54%, with an average cell loss of 37% at 6 months, and from 24% to 61%, with an average cell loss of 42% at 12 months. The average best-corrected Snellen visual acuity (mean, 9 months; range, 3–21 months) ranged from 20/34 to 20/66. A review of postoperative refractive results found induced hyperopia ranging from 0.7 to 1.5 diopters (D; mean, 1.1 D), with minimal induced astigmatism ranging from −0.4 to 0.6 D and a mean refractive shift of 0.11 D. A review of graft survival found that clear grafts at 1 year ranged from 55% to 100% (mean, 94%). Conclusions The evidence reviewed is supportive of DSEK being a safe and effective treatment for endothelial diseases of the cornea. In terms of surgical risks, complication rates, graft survival (clarity), visual acuity, and endothelial cell loss, DSEK appears similar to penetrating keratoplasty (PK). It seems to be superior to PK in terms of earlier visual recovery, refractive stability, postoperative refractive outcomes, wound and suture-related complications, and intraoperative and late suprachoroidal hemorrhage risk. The most common complications of DSEK do not appear to be detrimental to the ultimate vision recovery in most cases. Long-term endothelial cell survival and the risk of late endothelial rejection are beyond the scope of this assessment. Financial Disclosure(s) Proprietary or commercial disclosure may be found after the references.

Terahertz (THz) imaging of corneal tissue water content (CTWC) is a proposed method for early, accurate detection and study of corneal diseases. Despite promising results from ex vivo and in vivo cornea studies, interpretation of the reflectivity data is confounded by the contact between corneal tissue and rigid dielectric window used to flatten the imaging field. This work develops a novel imaging system and image reconstruction methods specifically for nearly spherical targets such as human cornea. A prototype system was constructed using a 650-GHz multiplier source and Schottky diode detector. Resolution and imaging field strength measurement from characterization targets correlate well with those predicted by the quasioptical theory and physical optics analysis. Imaging experiments with corneal phantoms and ex vivo corneas demonstrate the hydration sensitivity of the imaging system and reliable measurement of CTWC. We present successful acquisition of noncontact THz images of in vivo human cornea, and discuss strategies for optimizing the imaging system design for clinical use.

Limbal stem cells (LSCs) maintain the normal homeostasis and wound healing of corneal epithelium. Limbal stem cell deficiency (LSCD) is a pathologic condition that results from the dysfunction and/or an insufficient quantity of LSCs. The diagnosis of LSCD has been made mainly based on medical history and clinical signs, which often are not specific to LSCD. Methods to stage the severity of LSCD have been lacking. With the application of newly developed ocular imaging modalities and molecular methods as diagnostic tools, standardized quantitative criteria for the staging of LSCD can be established. Because of these recent advancements, effective patient-specific therapy for different stages of LSCD may be feasible.

The corneal epithelium is continuously renewed by limbal stem/progenitor cells (LSCs), a cell population harbored in a highly regulated niche located at the limbus. Dysfunction and/or loss of LSCs and their niche cause limbal stem cell deficiency (LSCD), a disease that is marked by invasion of conjunctival epithelium into the cornea and results in failure of epithelial wound healing. Corneal opacity, pain, loss of vision, and blindness are the consequences of LSCD. Successful treatment of LSCD depends on accurate diagnosis and staging of the disease and requires restoration of functional LSCs and their niche. This review highlights the major advances in the identification of potential LSC biomarkers and components of the LSC niche, understanding of LSC regulation, methods and regulatory standards in bioengineering of LSCs, and diagnosis and staging of LSCD. Overall, this review presents key points for researchers and clinicians alike to consider in deepening the understanding of LSC biology and improving LSCD therapies.