Generation of an induced pluripotent stem cell (iPSC) line from a 40-year-old patient with the A8344G mutation of mitochondrial DNA and MERRF (myoclonic epilepsy with ragged red fibers) syndrome

Abstract

Mitochondrial defects are associated with clinical manifestations from common diseases to rare genetic disorders. Myoclonus epilepsy associated with ragged-red fibers (MERRF) syndrome results from an A to G transition at nucleotide position 8344 in the tRNA gene of mitochondrial DNA (mtDNA) and is characterized by myoclonus, myopathy and severe neurological symptoms. In this study, Sendai reprogramming method was used to generate an iPS cell line carrying the A8344G mutation of mtDNA from a MERRF patient. This patient-specific iPSC line expressed pluripotent stem cell markers, possessed normal karyotype, and displayed the capability to differentiate into mature cells in three germ layers.

Mitochondrial impairment and synaptic dysfunction are associated with neurological defects in iPSCs-derived cortical neurons of MERRF patients

“…The 6 lines of human iPSCs used in this study were generated from the skin fibroblasts established from 3 patients harboring the MERRF syndrome-specific m.8344A > G mutation. Among them, two patients are the members of a Taiwanese MERRF family, a 15-year-old girl (M1-iPSCs) and her 13-year-old sister (M2-iPSCs), and the third patient is a 25-year-old female (M3-iPSCs) as described previously [ 12 , 14 , 16 ]. M1 patient had poor learning ability in childhood and developed myoclonic epilepsy at 12 years of age, and exhibited severe clinical symptoms, including unsteady gait with tremor, intermittent myoclonus and polyneuropathy [ 17 ].…”

“…Among all iPSCs lines, M1-iPSC sublines (M1 Low and M1 High iPSCs) and M2-iPSCs (M2 High iPSCs) were generated at Taipei Veterans General Hospital and have been used for the studies reported previously [ 12 ]. M3-iPSCs sublines (M3 Low and M3 Med iPSCs) were generated from another MERRF patient in collaboration with Prof. Patrick C. H. Hsieh at the Institute of Biomedical Sciences, Academia Sinica supported by Taiwan Human Disease iPSC Service Consortium [ 14 ]. Prof. Y. C. Hsu at Mackay Medical College kindly provided us the normal iPSCs (IBMS-iPSC-02-07), which were generated from the peripheral blood of a healthy 22-year-old Asian male volunteer and are commercially available from the Food Industry Research and Development Institute, Hsinchu, Taiwan.…”

“…For this study, the IRB waived the informed consent requirement for the use of these iPSCs cell lines. The ability of the MERRF iPSCs to differentiate into three germ layers in vitro and form teratoma and three germ layers in vivo was confirmed during characterization of iPSCs [ 12 , 14 ]. The iPSCs were expanded in Gibco™ StemFlex™ medium on Geltrex-coated plates at 37 °C in a humidified CO 2 incubator.…”

“…Several studies have used iPSCs to investigate the mechanism of pathogenesis of mitochondrial diseases, such as LHON and the MELAS syndrome [ 8 – 11 ]. In the past few years, we have generated several iPSCs lines by using the primary culture of skin fibroblasts from patients with MERRF syndrome and other mitochondrial diseases, respectively [ 12 – 14 ]. We have successfully differentiated MERRF iPSCs into inner ear hair cell-like cells [ 15 ] and cardiomyocytes [ 12 ], respectively, and both of which displayed the phenotype caused by mitochondrial dysfunction.…”

Development of a bioactive microencapsulation platform incorporating decellularized extracellular matrix to entrap human induced pluripotent stem cells for versatile biomedical applications

“…The cells were found to exhibit characteristic human pluripotent stem cell morphology (Figure 1A). Moreover, they were positive for key markers that are typically screened for to validate pluripotency—Nanog, Sox2, SSEA4, Tra‐1‐60, and Oct‐4, 28–30 thus confirming their identity (Figure 1B–F, respectively).…”