Iyer 2017 MiP2017
Mutations in the mitochondrial DNA (mtDNA) lead to many human mitochondrial disorders with impaired bioenergetics. The clinical severity of the disease may also depend on the load of mutant mtDNA and bioenergetic dysfunction. The development of FDA –approved therapies for primary mitochondrial disorders have been hindered by genetic and clinical heterogeneity, limited knowledge of natural history of disease progression, and lack of clear clinical endpoints. Our long-term goal is the development of patient- and disease-specific cell based models that will provide us novel insights into disease origins and severity.
In this study, we have focused our efforts on Leigh’s syndrome (LS), as there is no current cure for LS nor an adequate model for understanding the rapid fatality associated with the disease. We characterized the mutation and mitochondrial function of two patient-derived fibroblasts from LS carrying T8993G point mutation encoding subunit of ATP synthase (Complex V); and T10185C point mutation encoding subunit of NADH dehydrogenase (Complex I). Using Sanger sequencing approaches, we confirmed the presence of the mutation in the patient fibroblasts. Immunocytochemical analysis based on use of MitoTracker Red CM-H2Xros and MitoTracker™ Green FM demonstrated that T8993G and T10185C cells have a low number of active mitochondria compared to healthy fibroblasts. Our preliminary observations indicate that the basal respiration and ATP production in the diseased cell lines were significantly lower than healthy fibroblast cells, while the proton leak and non-mitochondrial oxygen consumption of T8993G cells was higher compared to T10185C and healthy cells. Our findings demonstrate that both the patient fibroblasts have mitochondrial dysfunction that correlate with the pathogenic mtDNA burden. We are currently monitoring these parameters in the reprogrammed and differentiated cells carrying the mutation.
Labels: MiParea: Respiration, mtDNA;mt-genetics, Patients
Stress:Mitochondrial disease Organism: Human Tissue;cell: Fibroblast
Coupling state: LEAK, ROUTINE
Dept Biological Sciences, Fulbright College Arts Sciences, Univ Arkansas, Fayetteville, AR, USA. - firstname.lastname@example.org