The long term goal of this proposal is to develop a successful therapeutic strategy for treating corneal cystinosis. Cystinosis is a rare lysosomal storage disease caused by a defect in the cystine lysosomal transporter cystinosin (CTNS) that leads to cystine crystal formation in various tissues and organs resulting in multiple organ failure at an early age. In the cornea, progressive visual deterioration due to photophobia, blepharospasm and recurrent corneal erosions are associated with increasing concentrations of corneal crystals and are a major long-term burden for patients with cystinosis despite successful treatment of other organ systems. Recently, a cystinosin knockout mouse (Ctns-/-) has been generated that develops cystine crystals in tissues and shows progression of disease similar to that of cystinosis patients, including decreased kidney function and formation of corneal cystine crystals. In collaboration with Drs. Stephanie Cherqui at Scripps and Winston Kao at University of Cincinnati, we have conducted preliminary experiments to evaluate potential novel therapies for treating corneal cystinosis. These preliminary studies suggest that intra- corneal injection of human umbilical cord mesenchymal stem cells (hUCMSC) inhibits progression and scarring of Ctns-/- mouse corneas, and reduces the presence of intrastromal corneal crystals, suggesting that hUCMSC can replace dead or dying corneal keratocytes and restore corneal function. Based on these findings we propose the HYPOTHESIS: That hUCMSC intrastromal transplantation can rescue or block the development of corneal cystinosis in the Ctns-/- mouse. This hypothesis prompts the following important QUESTIONS that need to be addressed. What percent of the host keratocyte population needs to be replaced by hUCMSC to inhibit the development of corneal cystinosis (QUESTION 1)? Does stage of disease affect engraftment and differentiation of hUCMSC (QUESTION 2)? And, does engraftment of hUCMSC affect normal corneal responses to injury (QUESTION 3)? To address these questions have proposed the following specific aims: 1. Determine the effect of intrastromal injection of hUCMSC on the development of corneal cystinosis by injecting different numbers of hUCMSC into young (3 month old) mice and measuring in vivo cystine crystal volume, hUCMSC keratocyte differentiation and cell density over time. (QUESTION 1) 2. Assess the effect of stage of corneal cystinosis on hUCMSC engraftment by injecting the optimal number of hUCMSC in different aged mice and measuring in vivo cystine crystal volume, hUCMSC keratocyte differentiation and cell density over time, (QUESTION 2) 3. Characterize the wound healing response of hUCMSC engrafted corneas by performing epithelial scrape injuries in optimally engrafted mouse corneas and measure epithelial and keratocyte regeneration and return to keratocyte phenotype. (QUESTION 3)

Public Health Relevance

This study will evaluate the potential therapeutic effects of human umbilical cord mesenchymal stem cells (hUCMSC) transplantation into to the corneal stroma to treat a mouse model of corneal cystinosis. Progression of corneal cystinosis and the formation of intrastromal cystine crystals will be evaluated by quantitative in vivo confocal microscopy. Laser scanning confocal microscopy, immunocytochemistry and molecular biology will be used to evaluate differentiation of hUCMSC to mouse corneal keratocytes and their response to corneal injury.

National Institute of Health (NIH)
National Eye Institute (NEI)
Research Project (R01)
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Anterior Eye Disease Study Section (AED)
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Mckie, George Ann
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University of California Irvine
Schools of Medicine
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