NIH Research Festival
Fabry disease is a glycosphingolipid storage disorder caused by a deficiency of the enzyme alpha-galactosidase A (AGA) which results in the storage of Gb3 (globotriaosylceramide). This buildup of Gb3 leads to a multisystem disease that affects the vascular, cardiac, renal, and nervous systems. One of the hallmarks of this disorder is neuropathic pain in hands and feet and nervous dysfunction. To better understand this complex phenotype, we sought to generate a human neuronal cell line lacking in AGA. We transfected the human embryonic stem cell line WA-14 with two different short-hairpin RNA plasmids against AGA message, and selected stable transformants with puromycin. We demonstrated that these cell lines have severely reduced AGA activity and decreased breakdown of Gb3 that is characteristic of the Fabry phenotype. In addition, they retained all markers of pluripotency tested. Using dual-SMAD inhibition, we showed our genetically engineered stem cells could be differentiated to peripheral neurons, confirmed by b-tubulin and peripherin immunostaining. In addition, the differentiated cells were positive by immunostaining for the pain receptors TRPV-1 and NAV-1.7. Our human stem cell model can now be used to study the mechanisms of pain in Fabry patients and could be used for screening of potential treatments.
Scientific Focus Area: Stem Cell Biology
This page was last updated on Friday, March 26, 2021