Stem Cell Therapy for Retinal Disorders: Generation of Human RPE from Skin Biopsies of AMD Patients


ABSTRACT

Purpose:

Retinal pigmented epithelium (RPE) derived from human induced pluripotent stem (iPS) cells is a promising source of cells for the replacement of degenerated RPE in patients with age-related macular degeneration (AMD), as use of patient-specific iPS will avoid the issue of graft rejection. Here we demonstrate that iPS can be derived from fibroblasts obtained from older patients with AMD, and that these cells can be used to generate autologous RPE for transplantation.

Methods:

Patients with advanced AMD exhibiting either geographic atrophy or exudative AMD were recruited. Fibroblasts obtained from skin biopsies were grown to confluence and iPS cells were generated by Sendai virus reprogramming (Life Technologies, Carlsbad, CA). We then determined the ability of these cells to first form iPS, and then for RPE to be generated from the iPS of these older patients.

Results:

Fibroblasts were obtained successfully from over 33 patients with advanced AMD and age-match controls. To date, patient-specific iPS cells have been generated from 18 of these samples, and 8 RPE cell lines have been generated. iPS cells were all positive for stem cell markers including SSEA4, TRA-1-60, OCT4 and TRA-1-81. Cells differentiated towards an RPE fate formed a hexagonal monolayer within 30 days. These monolayers were uniformly positive for RPE markers Bestrophin, MITF and RPE65 and the tight junction marker ZO-1. iPS-derived RPE secrete VEGF and PEDF in a polarized fashion.

Conclusions:

RPE derived from iPS cells exhibited similar physiological properties to human RPE cells. Patient-specific fibroblasts have been reprogrammed into iPS cells from AMD patients to at least age 80, with no decline is this ability with advancing patient age. iPS cells can be differentiated towards RPE, with cells expressing RPE markers and polarized secretion of VEGF and PEDF. The presence of disease or advanced patient age does not limit the ability to generate functional RPE from skin biopsies.


CONTACT DETAILS

 

Lucian V. Del Priore
New Haven, CT, United States
Email : ldelpriore@yahoo.com
Cell Phone: +19086250395
Work Phone: +12037371413