Samantha Bobba

Tracing the Fate of Limbal Epithelial Progenitor Cells in the Murine Cornea

Stem cell (SC) division, deployment, and differentiation are processes that contribute to corneal epithelial renewal. Until now studying the destiny of these cells in a living mammal has not been possible. However, the advent of inducible multicolor genetic tagging and powerful imaging technologies has rendered this achievable in the translucent and readily accessible murine cornea. K14CreERT2‐Confetti mice that harbor two copies of the Brainbow 2.1 cassette, yielding up to 10 colors from the stochastic recombination of fluorescent proteins, were used to monitor K‐14+ progenitor cell dynamics within the corneal epithelium in live animals. Multicolored columns of cells emerged from the basal limbal epithelium as they expanded and migrated linearly at a rate of 10.8 µm/day toward the central cornea. Moreover, the permanent expression of fluorophores, passed on from progenitor to progeny, assisted in discriminating individual clones as spectrally distinct streaks containing more than 1,000 cells within the illuminated area. The centripetal clonal expansion is suggestive that a single progenitor cell is responsible for maintaining a narrow corridor of corneal epithelial cells. Our data are in agreement with the limbus as the repository for SC as opposed to SC being distributed throughout the central cornea. This is the first report describing stem/progenitor cell fate determination in the murine cornea using multicolor genetic tracing. This model represents a powerful new resource to monitor SC kinetics and fate choice under homeostatic conditions, and may assist in assessing clonal evolution during corneal development, aging, wound‐healing, disease, and following transplantation. Stem Cells 2015;33:157–169

Clinical outcomes of xeno-free expansion and transplantation of autologous ocular surface epithelial stem cells via contact lens delivery: a prospective case series

IntroductionDepletion of limbal stem cells leads to a debilitating condition known as limbal stem cell deficiency, characterised by impaired corneal wound healing and poor vision. The aim of this study was to determine whether delivering progenitor cells on a contact lens is a viable and effective alternative to current transplantation techniques, which are complicated by biological and xenogeneic materials.MethodsSixteen eyes of 16 patients who had total (n = 14) and partial (n = 2) limbal stem cell deficiency (chemical burns, five eyes; iatrogenic causes, four eyes; aniridia, three eyes; trachoma-induced, two eyes; contact lens over-wear, one eye; and cicatrising conjunctivitis, one eye) and who had failed prior therapy were recruited prospectively into the study. Autologous limbal (n = 7) or conjunctival epithelial (n = 9) biopsies were harvested from patients and placed on the concave surface of silicone hydrogel contact lenses. Cells were expanded in culture with autologous serum and transplanted onto the ocular surface.ResultsRestoration of a transparent avascular and clinically stable corneal epithelium was attained in 10 of 16 eyes (63%) at a median follow-up time of 2.5 years (range of 0.8 to 5.8 years). Although minor complications occurred in two eyes of two patients because of contact lens insertion or removal, these were not associated with long-term sequelae.ConclusionsThis is the first and largest study to evaluate the mid-term outcomes of autologous limbal/conjunctival stem cell transplantation via a US Food and Drug Administration-approved contact lens, demonstrating that delivery of ocular progenitor cells via this procedure offers a viable, effective, and xeno-free alternative to current transplantation methodologies.Trial registrationAustralian New Zealand Clinical Trials Registry ACTRN012607000211460. Registered 17 April 2007.

Nature and incidence of severe limbal stem cell deficiency in Australia and New Zealand

This study aimed to determine the nature and incidence of severe limbal stem cell deficiency (LSCD) in Australia and New Zealand.

Native and synthetic scaffolds for limbal epithelial stem cell transplantation

Limbal stem cell deficiency (LSCD) is a complex blinding disease of the cornea, which cannot be treated with conventional corneal transplants. Instead, a stem cell (SC) graft is required to replenish the limbal epithelial stem cell (LESC) reservoir, which is ultimately responsible for regenerating the corneal epithelium. Current therapies utilize limbal tissue biopsies that harbor LESCs as well as tissue culture expanded cells. Typically, this tissue is placed on a scaffold that supports the formation of corneal epithelial cell sheets, which are then transferred to diseased eyes. A wide range of biological and synthetic materials have been identified as carrier substrates for LESC, some of which have been used in the clinic, including amniotic membrane, fibrin, and silicon hydrogel contact lenses, each with their own advantages and limitations. This review will provide a brief background of LSCD, focusing on bio-scaffolds that have been utilized in limbal stem cell transplantation (LSCT) and materials that are being developed as potentially novel therapeutics for patients with this disease. STATEMENT OF SIGNIFICANCE The outcome of patients with corneal blindness that receive stem cell grafts to restore eye health and correct vision varies considerably and may be due to the different biological and synthetic scaffolds used to deliver these cells to the ocular surface. This review will highlight the positive attributes and limitations of the myriad of carriers developed for clinical use as well as those that are being trialled in pre-clinical models. The overall focus is on developing a standardized therapy for patients, however due to the multiple causes of corneal blindness, a personal regenerative medicine approach may be the best option.

Incidence, clinical features and diagnosis of cicatrising conjunctivitis in Australia and New Zealand

AimsThis study aimed to determine the incidence, clinical features and management of cicatrising conjunctivitis in Australia and New Zealand, also enabling comparison with data from the United Kingdom.MethodsA prospective surveillance study was conducted over 17 months via the Australian and New Zealand Ophthalmic Surveillance Unit with a one-year follow-up period. Practicing ophthalmologists on the Surveillance Unit’s database were asked to report recently diagnosed cases of cicatrising conjunctivitis on a monthly basis. Initial and follow-up questionnaires were sent to ophthalmologists who had reported positive cases to obtain demographic and clinical data. The minimum incidence of cicatrising conjunctivitis was calculated based on cases reported during the study period and from population data.ResultsDuring the 17-month study period (December 2011–April 2013), 56 cases of cicatrising conjunctivitis were reported. Data was obtained for 35 cases (62%) with a mean age of 74 years (range, 28–94 years). The most common aetiologies were ocular mucus membrane pemphigoid (n = 18 cases, 51.4%), Stevens–Johnson Syndrome (n = 3, 8.6%) and graft versus host disease (n = 3, 8.6%). The minimum incidence of cicatrising conjunctivitis in Australia and New Zealand was 1.5 per million, comparable to incidence data from the United Kingdom.ConclusionsThis study is the first to prospectively record the incidence of cicatrising conjunctivitis in Australia and New Zealand and the second worldwide. It provides novel data on demographics and management of cicatrising conjunctivitis, as reported by treating ophthalmologists.

The current state of stem cell therapy for ocular disease

Herein, we review the safety, efficacy, regulatory standards and ethical implications of the use of stem cells in ocular disease. A literature review was conducted, registered clinical trials reviewed, and expert opinions sought. Guidelines and codes of conduct from international societies and professional bodies were also reviewed. Collated data is presented on current progress in the field of ocular regenerative medicine, future challenges, the clinical trial process and ethical considerations in stem cell therapy. A greater understanding of the function and location of ocular stem cells has led to rapid advances in possible therapeutic applications. However, in the context of significant technical challenges and potential long-term complications, it is imperative that stem cell practices operate within formal clinical trial frameworks. While there remains broad scope for innovation, ongoing evidence-based review of potential interventions and the development of standardized protocols are necessary to ensure patient safety and best practice in ophthalmic care.

Isolated painful trigeminal neuropathy as an unusual presentation of chronic lymphocytic inflammation with pontine perivascular enhancement responsive to steroids: A case report:

Background Chronic lymphocytic inflammation with pontine perivascular enhancement responsive to steroids (CLIPPERS) is an inflammatory central nervous system disorder, typically presenting with subacute symptoms referable to brainstem and cerebellar pathology. This is the first report of CLIPPERS presenting with a painful trigeminal neuropathy. Case report We report an unusual case of CLIPPERS presenting with facial pain and sensory symptoms, in the absence of other brainstem or cerebellar signs. Perivascular enhancement of peri-pontine structures on neuroimaging, lymphocytic infiltrate on histopathology and rapid clinical and radiological responsiveness to glucocorticosteroids were key to diagnosis. Extensive investigations excluded various differential aetiologies. Conclusion The pathogenesis of CLIPPERS is poorly understood, and the diagnostic criteria are yet to be validated. In this case, facial pain was not associated with other brainstem or cerebellar signs, broadening current understanding of how CLIPPERS may present. This has clinical implications in guiding future investigations for patients presenting with painful trigeminal neuropathy.