Abstract 19502: A Human Induced Pluripotent Stem Cell-derived Cardiomyocyte Model of Pompe Disease.
Background: Infantile Pompe disease is an autosomal recessive genetic disorder characterized by a lack of acid alpha glucosidase (GAA) activity, the sole enzyme responsible for degrading glycogen within lysosomes. Pathological buildup of lysosomal glycogen in skeletal and cardiac muscle leads to respiratory insufficiency and hypertrophic cardiomyopathy causing death in infancy. A human cardiomyocyte model of Pompe disease does not exist.
Methods: Skin fibroblasts from a patient with infantile Pompe disease (ID: GM04912) caused by compound heterozygous mutations of GAA (1441delT/2237G>A) giving two null alleles and a heterozygote for GAA (ID: GM20123) with one null allele (deletion exon 18) giving a normal phenotype were reprogrammed into induced pluripotent stem (iPS) cells using six genes: OCT4, NANOG, SOX2, KLF4, LIN28, and CMYC. Several iPS and embryonic stem (ES) cells with wild type GAA alleles were used for comparison. Cardiomyocytes produced using a directed differentiation protocol were characterized biochemically and the cellular ultrastructure described with electron microscopy.
Results: The fibroblast cell lines were successfully reprogrammed into iPS cells based on colony properties and pluripotency marker expression. Differentiation was successful in generating contracting areas containing electrically-coupled cardiomyocytes. Assays confirmed the absence of GAA activity in GM04912 fibroblasts, iPS cells, and contracting areas compared to GM20123 and wild-type controls which exhibited GAA activity in the normal range (260±82 nmol glucose released/mg protein/hr). Western blots of GM04912 iPS cells lack GAA immunoreactivity compared to GAA bands at 110 (preproprotein), 76 (intermediate), and 70 (active protein) kDa in GM20123 and control iPS/ES cells. The ultrastructure of 1-month old cardiomyocytes from GM04912 exhibited distended glycogen-filled lysosomes, the hallmark feature of infantile Pompe disease. Cardiomyocytes from GM20123 and control lines only contained free cytoplasmic glycogen.
Conclusions: Human cardiomyocyte cell culture models of disease are possible via iPS technology. The generation of Pompe cardiomyocytes offers new strategies for understanding and developing therapies for the disease.
- Congenital heart disease
- Pediatric cardiology
- Stem cells
- Mitochondrial energetics, heart failure, arrhythmias
- Hypertrophic cardiomyopathy
- © 2010 by American Heart Association, Inc.