Glare prediction and mechanism of adaptation following implantation of hydrophilic and hydrophobic intraocular lenses.

Purpose: Glare is a known side effect of intraocular lens (IOL) implantation, affected principally by IOL material and optics, although it is reported subjectively to decrease in impact with time. However, little objective data have been published on changes over time, how these relate to subjective reports, and whether those who will report greater glare symptoms can be predicted prior to IOL implantation. Methods: A total of 32 patients (aged 72.4 ± 8.0 years) with healthy eyes were implanted bilaterally with hydrophilic 600s (Rayner, Worthing, UK) or hydrophobic Acrysof (Alcon, Texas, USA) acrylic IOLs (n = 16 each, randomly assigned). Each patient reported their dysphotopsia symptoms subjectively using the validated forced choice photographic questionnaire for photic phenomena, and halo size resulting from a bright light in a dark environment was quantified objectively in eight orientations using the Aston Halometer. Assessment was performed binocularly pre-operatively and at 1, 2, 3, and 4 weeks after IOL implantation. Setting: The study was carried out at the National Health Service Ophthalmology Department, Queen Elizabeth Hospital, Birmingham, UK. Results: Visual acuity (average 0.37 ± 0.26 logMAR) did not correlate with subjective glare (r = 0.184, p = 0.494) or objective glare (r = 0.294, p = 0.270) pre-surgery. Objective halo size (F = 112.781, p < 0.001) decreased with cataract removal and IOL implantation and continued to decreased over the month after surgery. Subjective dysphotopsia complaints (p < 0.001) were also greater pre-surgery, but did not change thereafter (p = 0.228). In neither case was there a difference with IOL material (p > 0.05). It was not possible to predict post-surgery dysphotopsia from symptoms or a ratio of symptoms to halo size pre-surgery (p > 0.05). Conclusions: Subjective dysphotopsia and objective halos caused by cataracts are greatly reduced by implantation of IOL after cataract removal causing few perceivable symptoms. However, objective measures are able to quantify a further reduction in light scatter over the first month post-IOL implantation, suggesting that any subjective effects over this period are due to the healing process and not due to neuroadaptation.

Patient priorities in herpes simplex keratitis.

OBJECTIVE: Herpes simplex keratitis (HSK) is a sight-threatening disease and a leading cause of infectious corneal blindness. Involving patients in setting the research agenda maximises patient benefit and minimises research waste. With no published patient involvement exercises, patients' priorities in HSK are unclear. The objective of this study is to explore patients' priorities for research in HSK. METHODS: A literature review of publications in the year preceding recruitment of patients identified nine domains of research interest. A questionnaire was sent to participants asking them to rank these in order of priority. The ranking results were given a weighted-average score, and a thematic analysis was undertaken for the narrative data. RESULTS: Thirty-seven patients participated in the survey. Top priorities for patients were risk factors for recurrence of infection, diagnostic tests and treatment failure. The narrative data revealed three key clinical needs: difficulties in long-term symptom control, the need for rapid access care in acute infection and the desire for more accessible information. CONCLUSION: This study highlighted three major issues in our current approach to HSK. First, there may be a misalignment between research efforts and patient priorities. Second, high-quality patient information is not widely available. This may hamper patients' abilities to make informed decisions and contribute towards research. Third, clinical service priorities are of equal importance to patients as research. Researchers and clinicians are encouraged to address both needs in parallel.

Rotational Stability and Centration of a New Toric Lens Design Platform Using Objective Image Analysis Over 6 Months.

PURPOSE: To determine the rotational stability of the Rayner 600S intraocular lens (IOL) (Rayner, Worthing, United Kingdom) using objective image analysis techniques. METHODS: A total of 66 patients (mean age: 69.92 ± 8.45 years) with healthy eyes presenting for routine cataract surgery were implanted monocularly with the 600S IOL and followed up over 90 to 180 days postoperatively. Images of the IOL were captured through a maximally dilated pupil (tropicamide 1.0%, phenylephrine 2.5%) immediately after surgery and after 1 to 3 days, 30 days, and 90 to 180 days using a digital slit-lamp biomicro-scope. The orientation of the IOL markings were image analyzed compared to conjunctival blood vessel landmarks visible across all time points to correct for head and rotation. Centration was assessed by drawing ovals to circumscribe the IOL, pupil, and limbus to compare the centers (0.01 mm per-pixel resolution). RESULTS: Absolute values for rotation postoperatively were 1.60° ± 1.13° at 1 to 3 days, 1.58° ± 1.36° at 30 days, and 1.83° ± 1.44° at 90 to 180 days. No lens rotated more than 5°. The average centration relative to the limbus was -0.04 ± 0.25 mm horizontally and 0.04 ± 0.26 mm vertically at 1 to 3 days, 0.00 ± 0.38 mm vertically and 0.01 ± 0.28 mm horizontally at 30 days, and 0.08 ± 0.37 mm vertically and 0.07 ± 0.47 mm horizontally at 90 to 180 days. CONCLUSIONS: The Rayner 600S IOL showed exceptional rotational stability and centration after implantation, meeting the U.S. Food and Drug Administration prescribed American National Standards Institute standards for toric IOLs. [J Refract Surg. 2019;35(1):48-53.].

The Role of Nerve Growth Factor in Maintaining Proliferative Capacity, Colony-Forming Efficiency, and the Limbal Stem Cell Phenotype.

Nerve growth factor (NGF) has demonstrated great benefit in the treatment of neurotrophic corneal ulcers. There is evidence for multiple modes of action in promoting corneal healing, but only indirect evidence exists for NGF's effects on limbal stem cells (LSCs). Understanding the role of NGF in LSC biology will improve our understanding of paracrine regulation of the limbal niche and the design of stem cell-based therapies for conditions such as LSC deficiency. In this article, we studied the regulation of NGF signaling components during LSC differentiation and the role of NGF in LSC proliferation and maintenance of the stem cell phenotype. LSC differentiation was induced by prolonged (40 day) culture which resulted in a significant increase in cell size, decrease in colony-forming efficiency and expression of putative LSC markers. A protein microarray measuring expression of 248 signaling proteins indicated the low affinity NGF receptor p75NTR to be the most downregulated protein upon differentiation. Further confirmation by Western blotting and real-time quantitative polymerase chain reaction indicated that NGF and p75NTR are expressed in early LSC cultures and downregulated upon differentiation. LSC cultures grown in the presence of anti-NGF antibody showed decreased colony-forming efficiency, DNA replication and expression of putative LSC markers ABCG2 and C/EBPδ. Supplementation of LSC culture medium with NGF extended the life span of LSC cultures in vitro and increased the expression of putative LSC markers ΔNp63α and ABCG2. Taken together, our data indicate that NGF signaling is a key promoter of LSC proliferation, colony-forming efficiency, and a maintainer of the LSC phenotype. Stem Cells 2019;37:139-149.

Toxic epidermal necrolysis: the red eye and red herrings in casualty.

A 38-year-old woman presented to casualty with bilateral red eyes associated with a recent upper respiratory tract infection. This was initially diagnosed as conjunctivitis, however systemic review revealed an erythematous facial and skin rash, mildly swollen lips and mild swallowing difficulties. The patient was referred for an urgent medical assessment, by which time she was found to have erythema affecting 54% of her body surface area and diagnosed with suspected toxic epidermal necrolysis (TEN). She rapidly deteriorated over 24 hours with a spreading blistering skin rash and airway compromise requiring urgent intubation and admission to the intensive treatment unit (ITU). Subsequent skin biopsies confirmed the diagnosis of TEN, attributed to recent use of ibuprofen. Treatment included broad-spectrum antibiotics and high-dose corticosteroids. The patient had a prolonged hospital stay and developed severe scarring of the ocular surface. She was discharged home and remains under continuing outpatient follow-up with ophthalmology and dermatology teams.

FOXC1 plays a crucial role in the growth of pancreatic cancer.

IGF-1R signaling controls various vital cellular functions and this signaling is deregulated in many cancers, including pancreatic cancer. Several efforts have mainly focused on inhibiting the IGF-1R signaling cascade. The outcomes of these focused preclinical studies have been positive, whereas clinical trials of IGF-1R inhibitors in pancreatic cancer have failed, raising the questions about this therapeutic approach. This necessitates a better understanding of the role of IGF-1R signaling in pancreatic cancer. We investigated the impact of IGF-1R signaling on crucial transcription factors and identified the FOXC1 as one of the crucial regulator of IGF-1R signaling. We employed genetic approaches to overexpress and silence FOXC1 in pancreatic cancer cells. Our results demonstrate that IGF-1R and FOXC1 seem to positively regulate each other. Further, FOXC1 increased the metastatic abilities of pancreatic cancer cells by enhancing cell proliferation, migration, invasion, epithelial-to-mesenchymal transition, and angiogenesis. The data from xenograft experiments further established the importance of FOXC1 in pancreatic tumorigenesis. In conclusion, FOXC1 is a potent oncogenic transcription factor, which promotes pancreatic cancer growth and metastasis. Thus, targeting FOXC1 could be a potential therapeutic strategy against pancreatic cancer.

Vitamin A deficiency due to chronic malabsorption: an ophthalmic manifestation of a systemic condition.

A 47-year-old woman presented with a 4-week history of progressive loss of vision, first manifesting as night blindness. Additionally, the patient reported frequent severe episodes of diarrhoea over the past month. Her medical history included end-stage renal failure for which she was currently on haemodialysis after a failed renal transplant, chronic pancreatitis and autonomic diabetes mellitus. Ophthalmological examination revealed severe bilateral corneal xerosis, bilateral Bitot's spots and inferior ulceration of the right cornea. A diagnosis of xerophthalmia due to vitamin A deficiency was made, most likely due to the presence of small intestinal bacterial overgrowth and the patient's chronic malabsorptive state. Standard management using oral vitamin A tablets was ineffective, resulting in the patient requiring intravenous supplementation. The extent of visual deterioration on presentation and the difficulties encountered managing the patient resulted in the patient's vision failing to improve.

Successful application of ex vivo expanded human autologous oral mucosal epithelium for the treatment of total bilateral limbal stem cell deficiency.

Ocular surface reconstruction with ex vivo expanded limbal stem cells (LSCs) is a widely used clinical treatment for patients with limbal stem cell deficiency (LSCD). This is not applicable to patients with bilateral LSCD where there are no remaining LSCs. Cultivated oral mucosa epithelium (OME) has been used as an alternative source of autologous epithelial stem cells for ocular reconstruction in few clinical trials. However, successful generation of stratified OME epithelium has only been achieved in the presence of animal feeder cells and/or animal-derived products in the culture media, likely to contribute to increased risk of pathogen transmission and graft rejection. In this study, we report generation of multilayered OME epithelium that shares many of the characteristics of corneal epithelium using a fully compliant good manufacturing practice, feeder- and animal product-free method. Proof of concept was achieved by transplantation of autologous ex vivo expanded OME in two patients with histologically confirmed bilateral total LSCD that resulted in successful reversal of LSCD in the treated eye up to 24 months.

Urrets-Zavalia syndrome following penetrating keratoplasty for keratoconus.

BACKGROUND: Urrets-Zavalia syndrome (UZS) consists of a fixed dilated pupil associated with iris atrophy. It is a poorly understood complication following penetrating keratoplasty (PKP) for keratoconus (KC). In this work, we aim to establish the incidence, visual outcomes, and an understanding of UZS. METHODS: This was a retrospective single-center study in a tertiary eye service in the United Kingdom of consecutive patients with UZS following PKP for KC in a 10-year period. Post-operative complications, including raised intraocular pressure (IOP), were recorded. UZS patients and age-matched control patients who had undergone PKP for KC without developing UZS attended a comprehensive clinical assessment. Anterior segment indocyanine green (ICG) angiography assessed iris perfusion. RESULTS: The incidence of UZS was 16.2 %. There was no difference in LogMAR VA or Pelli-Robson contrast sensitivity between groups. There was higher first-day post-operative IOP in UZS (p = 0.02). UZS patients had increased pupil size (p = 0.09) with reduced response to pilocarpine 2 % (p < 0.001). ICG angiography revealed delayed/reduced iris vasculature filling in UZS compared with normal filling patterns of controls. CONCLUSIONS: Elevated post-operative IOP within 24 h was a significant factor in the development of UZS. Visual function in UZS patients was unaffected. UZS patients developed longstanding mydriasis with reduced reactivity to topical pilocarpine 2 %. ICG angiography confirmed iris vessel ischemia; supporting the theory that iris ischemia resulting from occlusion of iris root vessels due to elevated IOP causes UZS. We advocate rigorous intraoperative management of ocular viscoelastic devices and aggressive postoperative IOP control in patients undergoing PKP for KC.

Stability of therapeutic retreatment of corneal wavefront customized ablation with the SCHWIND CAM: 4-year data.

PURPOSE: To evaluate the long-term outcomes of aspheric corneal wavefront ablation profiles for excimer laser retreatment. METHODS: Eighteen eyes that had previously undergone LASIK or photorefractive keratectomy (PRK) were retreated with LASIK using the corneal wavefront ablation profile. Custom Ablation Manager (SCHWIND eye-tech-solutions, Kleinostheim, Germany) software and the ESIRIS flying spot excimer laser system (SCHWIND) were used to perform the ablations. Refractive outcomes and wavefront data are reported up to 4 years after retreatment. Pre- and postoperative data were compared with Student t tests and (multivariate) correlation tests. P<.05 was considered statistically significant. A bilinear correlation of various postoperative wavefront aberrations versus planned correction and preoperative aberration was performed. RESULTS: Mean manifest refraction spherical equivalent (MRSE) before retreatment was -0.38±1.85 diopters (D) and -0.09±0.22 D at 6 months and -0.10±0.38 D at 4 years postoperatively. The reduction in MRSE was statistically significant at both postoperative time points (P<.005). Postoperative aberrations were statistically lower (spherical aberration P<.05; coma P<.005; root-mean-square higher order aberration P<.0001) at 4 years postoperatively. Distribution of the postoperative uncorrected distance visual acuity (P<.0001) and corrected distance visual acuity (P<.01) were statistically better than preoperative values. CONCLUSIONS: Aspheric corneal wavefront customization with the ESIRIS yields visual, optical, and refractive results comparable to those of other wavefront-guided customized techniques for the correction of myopia and myopic astigmatism. The corneal wavefront customized approach shows its strength in cases where abnormal optical systems are expected. Systematic wavefront customized corneal ablation appears safe and efficacious for retreatment cases.

Femtosecond LASIK retreatment using side cutting only.

PURPOSE: To present visual and refractive outcomes in 11 eyes that underwent femtosecond LASIK and subsequent retreatment with flap lifting using only side cutting. METHODS: Seven patients (11 eyes) with a mean age of 30.55±5.42 years (range: 24 to 39 years) underwent retreatment 18.18±5.41 months (range: 12 to 26 months) after primary LASIK treatment with side cutting only using a femtosecond laser for the correction of residual refractive error. RESULTS: Mean follow-up after retreatment was 7.72±1.48 months (range: 6 to 10 months). No intra- or postoperative complications were found after LASIK retreatment. Uncorrected distance visual acuity (UDVA) improved in all patients. Mean UDVA improved from 0.54±0.12 (decimal scale) (range: 0.4 to 0.7) preoperatively to 0.99±0.03 (range: 0.9 to 1.0) after retreatment. No patient lost lines of corrected distance visual acuity. CONCLUSIONS: Retreatment using a femtosecond laser to create only a side cut is an effective modality to treat residual refractive errors in postoperative LASIK patients.

The effect of static cyclotorsion compensation on refractive and visual outcomes using the Schwind Amaris laser platform for the correction of high astigmatism.

PURPOSE: To compare the refractive and visual outcomes using the Schwind Amaris excimer laser in patients with high astigmatism (>1D) with and without the static cyclotorsion compensation (SCC) algorithm available with this new laser platform. METHODS: 70 consecutive eyes with ≥1D astigmatism were randomized to treatment with compensation of static cyclotorsion (SCC group- 35 eyes) or not (control group- 35 eyes). A previously validated optimized aspheric ablation algorithm profile was used in every case. All patients underwent LASIK with a microkeratome cut flap. RESULTS: The SCC and control group did not differ preoperatively, in terms of refractive error, magnitude of astigmatism or in terms of cardinal or oblique astigmatism. Following treatment, average deviation from target was SEq +0.16D, SD±0.52 D, range -0.98 D to +1.71 D in the SCC group compared to +0.46 D, SD±0.61 D, range -0.25 D to +2.35 D in the control group, which was statistically significant (p<0.05). Following treatment, average astigmatism was 0.24 D (SD±0.28 D, range -1.01 D to 0.00 D) in the SCC group compared to 0.46 D (SD±0.42 D, range -1.80 D to 0.00 D) in the control group, which was highly statistically significant (p<0.005). There was no statistical difference in the postoperative uncorrected vision when the aspheric algorithm was used although there was a trend to increased number of lines gained in the SCC group. CONCLUSIONS: This study shows that static cyclotorsion is accurately compensated for by the Schwind Amaris laser platform. The compensation of static cyclotorsion in patients with moderate astigmatism produces a significant improvement in refractive and astigmatic outcomes than when not compensated.

Safety and efficacy of collagen crosslinking for the treatment of keratoconus.

IMPORTANCE OF THE FIELD: Keratoconus is a condition that causes corneal ectasia and reduced vision in young adults. A proportion of these patients have progressive disease requiring corneal transplantation. A revolutionary new treatment that is purported to halt progression of keratoconus, known as collagen crosslinking (CXL), has recently been introduced into clinical practice. AREAS COVERED IN THIS REVIEW: CXL involves the treatment of the cornea with riboflavin followed by photoactivation with UVA light leading to corneal strengthening. This article reviews the basic science, clinical protocols, safety aspects and clinical results of CXL. WHAT THE READER WILL GAIN: The reader will gain a comprehensive understanding of: i) the basic science of CXL; ii) the optimised protocols for clinical use of CXL; iii) the results of all the main clinical trials in the literature; iv) contraindications to treatment and v) full clinical safety profile of CXL. TAKE HOME MESSAGE: CXL represents a new treatment that uniquely allows the halt of progression of keratoconus, thus preventing visual loss and the need for surgical intervention. Available data suggest that this treatment has high efficacy and is very safe and may represent the future standard treatment for progressive keratoconus.

Stem cell therapies for ocular surface disease.

Transparency of the cornea on the front surface of the eye is essential for vision. A variety of blinding ocular surface diseases involve the cornea. This review focusses on vision loss caused by disruption of the integrity and function of the outermost corneal layer (the epithelium) and the stem-cell-based therapeutic strategies in use and under development to restore sight in affected patients.

Successful clinical implementation of corneal epithelial stem cell therapy for treatment of unilateral limbal stem cell deficiency.

The corneal epithelium is maintained by a population of stem cells known as limbal stem cells (LSCs) due to their location in the basal layer of the outer border of the cornea known as the limbus. Treatment of limbal stem cell deficiency (LSCD) has been achieved with transplantation of ex vivo expanded LSCs taken from a small biopsy of limbus. This is a relatively new technique, and as such, specific national or international guidance has yet to be established. Because of the lack of such specific guidance, our group has sought to minimize any risk to the patient by adopting certain modifications to the research methodologies in use at present. These include the replacement of all non-human animal products from the culture system and the production of all reagents and cultures under Good Manufacturing Practice conditions. In addition, for the first time, a strictly defined uniform group of patients with total unilateral LSCD and no other significant ocular conditions has been used to allow the success or failure of treating LSCD to be attributable directly to the proposed stem cell therapy. A prospectively designed study with strict inclusion and exclusion criteria was used to enroll patients from our database of patients with unilateral LSCD. Eight eyes of eight consecutive patients with unilateral total LSCD treated with ex vivo expanded autologous LSC transplant on human amniotic membrane (HAM) with a mean follow-up of 19 (RANGE) months were included in the study. Postoperatively, satisfactory ocular surface reconstruction with a stable corneal epithelium was obtained in all eyes (100%). At last examination, best corrected visual acuity improved in five eyes and remained unchanged in three eyes. Vision impairment and pain scores improved in all patients (p < .05). This study demonstrates that transplantation of autologous limbal epithelial stem cells cultured on HAM without the use of non-human animal cells or products is a safe and effective method of reconstructing the corneal surface and restoring useful vision in patients with unilateral total LSCD.

Loss of corneal epithelial stem cell properties in outgrowths from human limbal explants cultured on intact amniotic membrane.

BACKGROUND: The corneal epithelium is renewed by stem cells located at the limbus, the so-called limbal stem cells (LSCs). Absence, damage or loss of the LSC population leads to the painful and blinding condition of LSC deficiency (LSCD). Ex vivo expansion of LSCs is an increasingly well recognized treatment modality for LSCD. One method of ex vivo expansion of LSCs involves the culture of limbal explants on amniotic membrane (AM). The purpose of this study was to analyze the outgrowths from human cadaveric limbal explants cultured on AM for properties associated with LSCs. In particular, the expression of putative stem cell markers and the colony-forming efficiency of the different zones of the outgrowth were studied. METHODS: The limbal explants were expanded in the standard way used for clinical transplantation and the outgrowths were divided into three zones depending on proximity to the explant (inner, middle and outer zones). The colony-forming efficiencies (CFEs) of cells from each zone were calculated. In addition, the expression of DeltaNp63, ABCG2 (both putative positive LSC markers) and cytokeratin K3 (marker of corneal differentiation) were assessed using quantitative reverse transcription PCR (RT-PCR). Immunohistochemistry on paraffin-embedded sections was also performed to demonstrate protein localization and allow further confirmation of the quantitative RT-PCR results. RESULTS: Successful cultures for both the explant outgrowths and the CFE calculations were obtained in every case. CFE showed a successive decline in zones further away from the explant (p < 0.00005). Quantitative RT-PCR revealed that the expression of the positive putative LSC markers DeltaNp63 and ABCG2 also showed a steady decrease in the zones furthest from the explant (p < 0.05 and p < 0.005, respectively). The expression of cytokeratin K3 was increased in zones furthest from the explant (p < 0.005). Immunohistochemistry on paraffin-embedded sections of intact ex vivo-expanded limbal epithelium for the putative positive marker p63 and cytokeratin K3 confirmed the findings of the quantitative RT-PCR and CFE results. CONCLUSIONS: We demonstrate for the first time that outgrowths from human limbal explants, a widely used technique in ex vivo expansion of LSCs for clinical transplantation, show a steady decline in a wide range of stem cell properties with distance from the central explant. These findings support the importance of proximity of stem cells to their niche environment in maintaining their undifferentiated state. These findings suggest the need for modifications of existing techniques to ensure maximum numbers of LSCs following ex vivo expansion protocols, which will then ensure the success of subsequent engraftment.

A putative role for RHAMM/HMMR as a negative marker of stem cell-containing population of human limbal epithelial cells.

The corneal epithelium is maintained by stem cells located at the periphery of the cornea in a region known as the limbus. Depletion of limbal stem cells (LSCs) results in limbal stem cell deficiency. Treatments for this disease are based on limbal replacement or transplantation of ex vivo expanded LSCs. It is, therefore, crucial to identify cell surface markers for LSCs that can be used for their enrichment and characterization. Aldehyde dehydrogenases (ALDHs) are enzymes which protect cells from the toxic effects of peroxidic aldehydes. In this manuscript, we show for the first time that ALDH1 is absent from the basal cells of the limbal and corneal epithelium. We separated limbal epithelial cells on the basis of ALDH activity and showed that ALDH(dim) cells expressed significantly higher levels of DeltaNp63 and ABCG2 as well as having a greater colony forming efficiency (CFE) when compared to ALDH(bright) cells. Large scale transcriptional analysis of these two populations led to identification of a new cell surface marker, RHAMM/HMMR, which is located in all layers of corneal epithelium and in the suprabasal layers of the limbal epithelium but is completely absent from the basal layer of the limbus. Our studies indicate that absence of RHAMM/HMMR expression is correlated with properties associated with LSCs. RHAMM/HMMR- limbal epithelial cells are smaller in size, express negligible CK3, have higher levels of DeltaNp63 and have a higher CFE compared to RHAMM/HMMR+ cells. Taken together these results suggest a putative role for RHAMM/ HMMR as a negative marker of stem cell containing limbal epithelial cells. Cell selection based on Hoechst exclusion and lack of cell surface RHAMM/HMMR expression resulted in increased colony forming efficiency compared to negative selection using RHAMM/HMMR alone or positive selection using Hoechst on its own. Combination of these two cell selection methods presents a novel method for LSC enrichment and characterization. Disclosure of potential conflicts of interest is found at the end of this article.

Differentiation of human embryonic stem cells into corneal epithelial-like cells by in vitro replication of the corneal epithelial stem cell niche.

Human embryonic stem cells (hESCs) are pluripotent cells capable of differentiating into any cell type of the body. It has long been known that the adult stem cell niche is vital for the maintenance of adult stem cells. The cornea at the front of the eye is covered by a stratified epithelium that is renewed by stem cells located at its periphery in a region known as the limbus. These so-called limbal stem cells are maintained by factors within the limbal microenvironment, including collagen IV in basement membrane and limbal fibroblasts in the stroma. Because this niche is very specific to the stem cells (rather than to the more differentiated cells) of the corneal epithelium, it was hypothesized that replication of these factors in vitro would result in hESC differentiation into corneal epithelial-like cells. Indeed, here we show that culturing of hESC on collagen IV using medium conditioned by the limbal fibroblasts results in the loss of pluripotency and differentiation into epithelial-like cells. Further differentiation results in the formation of terminally differentiated epithelial-like cells not only of the cornea but also of skin. Scanning electron microscopy shows that some differences exist between hESC-derived and adult limbal epithelial-like cells, necessitating further investigation using in vivo animal models of limbal stem cell deficiency. Such a model of hESC differentiation is useful for understanding the early events of epithelial lineage specification and to the eventual potential application of epithelium differentiated from hESC for clinical conditions of epithelial stem cell loss. Disclosure of potential conflicts of interest is found at the end of this article.