AAV Gene Therapy for MPS1-associated Corneal Blindness.

Although cord blood transplantation has significantly extended the lifespan of mucopolysaccharidosis type 1 (MPS1) patients, over 95% manifest cornea clouding with about 50% progressing to blindness. As corneal transplants are met with high rejection rates in MPS1 children, there remains no treatment to prevent blindness or restore vision in MPS1 children. Since MPS1 is caused by mutations in idua, which encodes alpha-L-iduronidase, a gene addition strategy to prevent, and potentially reverse, MPS1-associated corneal blindness was investigated. Initially, a codon optimized idua cDNA expression cassette (opt-IDUA) was validated for IDUA production and function following adeno-associated virus (AAV) vector transduction of MPS1 patient fibroblasts. Then, an AAV serotype evaluation in human cornea explants identified an AAV8 and 9 chimeric capsid (8G9) as most efficient for transduction. AAV8G9-opt-IDUA administered to human corneas via intrastromal injection demonstrated widespread transduction, which included cells that naturally produce IDUA, and resulted in a >10-fold supraphysiological increase in IDUA activity. No significant apoptosis related to AAV vectors or IDUA was observed under any conditions in both human corneas and MPS1 patient fibroblasts. The collective preclinical data demonstrate safe and efficient IDUA delivery to human corneas, which may prevent and potentially reverse MPS1-associated cornea blindness.

Complications related to a cosmetic eye-whitening procedure.

PURPOSE: To report sight-threatening complications following extensive bulbar conjunctival resection and postoperative mitomycin C therapy for cosmetic eye-whitening in the United States. DESIGN: Retrospective noncomparative case series. METHODS: Multicenter report of 9 patients referred for evaluation and management of complications following bilateral cosmetic eye whitening. RESULTS: Seventeen eyes of 9 patients underwent cosmetic eye-whitening performed between 2 and 48 months prior to referral to one of the centers. Sixteen of the 17 eyes had persistent conjunctival epithelial defects, with 10 eyes requiring amniotic membrane grafting to facilitate re-epithelialization. Four eyes of 2 patients developed limbal stem cell compromise confirmed with in vivo confocal laser scanning microscopy. One patient developed infectious scleritis and diplopia resulting from Tenon capsule scarring. Another patient developed scleral necrosis, secondary infectious scleritis, and infectious endophthalmitis. This patient subsequently developed noninfectious scleritis that required 3-drug-regimen immunosuppression. CONCLUSION: Severe adverse effects can occur after extensive cosmetic conjunctival resection followed by topical mitomycin C application. Patients and physicians should be aware of the potential sight-threatening complications associated with this eye-whitening procedure.

Impact of parenteral nutrition-associated liver disease on intestinal transplant waitlist dynamics.

BACKGROUND: United Network for Organ Sharing (UNOS) reports indicate that waiting list mortality for intestinal transplant candidates greatly exceeds that for all other organ transplant candidates. But United Network for Organ Sharing outcomes reports have not routinely distinguished between the intestine candidate subgroups that are listed only for an intestine and those that are also listed for a liver. STUDY DESIGN: Data were obtained by request from the collaborative Organ Procurement and Transplantation Network (United Network for Organ Sharing)/Scientific Registry of Transplant Recipients (University Research and Education Association) database. Waiting list data for intestinal transplant recipients from 1995 to 2004 were divided into those candidates listed for only an intestine and those listed for both an intestine and a liver. Additional data concerning overall waiting list outcomes and posttransplant survival rates stratified into pediatric and adult subsets were also obtained and analyzed. RESULTS: The overall number of candidates on the intestinal transplant waiting list has increased steadily since 1995 and, consistently, the majority of candidates have also been listed for a liver. This subset was found to have both a higher relative risk of dying while awaiting transplantation and lower relative odds of receiving transplants. In addition, parenteral nutrition-associated liver disease is a major problem across all age groups, as evidenced by the combined liver and intestine listings that compose the majority of both adult and pediatric waiting list populations. Posttransplant survival data were found to be superior for isolated intestine recipients compared with liver-intestine recipients. CONCLUSIONS: The preponderance of dual listings and their associated inferior outcomes, before and after transplantation, has skewed overall intestinal transplant outcomes. Because progression of parenteral nutrition-associated liver disease can be insidious, and recognition of irreversibility is often difficult, intestine-only transplants should be considered early for high-risk patients before parenteral nutrition-associated liver disease progression mandates inclusion of a liver graft also.

Proteomic analysis of reporter genes for molecular imaging of transplanted embryonic stem cells.

Study of stem cells may reveal promising treatment for diseases. The fate and function of transplanted stem cells remain poorly defined. Recent studies demonstrate that reporter genes can monitor real-time survival of transplanted stem cells in living subjects. We examined the effects of a novel and versatile triple fusion (TF) reporter gene construction on embryonic stem (ES) cell function by proteomic analysis. Murine ES cells were stably transduced with a self-inactivating lentiviral vector containing fluorescence (firefly luciferase; Fluc), bioluminescence (monomeric red fluorescence protein; mRFP), and positron emission tomography (herpes simplex virus type 1 truncated thymidine kinase; tTK) reporter genes. Fluorescence-activated cell sorting (FACS) analysis isolated stably transduced populations. TF reporter gene effects on cellular function were evaluated by quantitative proteomic profiling of control ES cells versus ES cells stably expressing the TF construct (ES-TF). Overall, no significant changes in protein quantity were observed. TF reporter gene expression had no effect on ES cell viability, proliferation, and differentiation capability. Molecular imaging studies tracked ES-TF cell survival and proliferation in living animals. In summary, this is the first proteomic study, demonstrating the unique potential of reporter gene imaging for tracking ES cell transplantation non-invasively, repetitively, and quantitatively.