Pecina 2014 Abstract MiP2014
|Manifestation of mitochondrial disorders of nuclear origin in lymphocytes.|
Mitochondrial diseases belong to most severe inherited metabolic diseases affecting the pediatric population. Diagnostics of a substantial part of mitochondrial diseases with unknown genetic cause relies on clinical symptoms and biochemical analysis of energetic function and content of individual mitochondrial proteins in patient tissues – mainly in bioptic samples of skeletal muscle and cell cultures of skin fibroblasts. However, due to their invasive nature, the biopsies are often refused by the patient’s parents. Therefore, we tested the diagnostic suitability of easily obtainable patient material – lymphocytes isolated from peripheral blood.
High-resolution respirometry enables sensitive analysis of the mitochondrial oxidative phosphorylation system in isolated lymphocytes. Substrate-inhibitor measurements in digitonin-permeabilized lymphocytes provide a complex evaluation of individual respiratory complexes, coupled ATP synthesis and kinetic parameters of mitochondrial respiratory enzymes. We employed this approach in a large cohort of 48 children including mostly subjects with suspected mitochondrial disease, previously diagnosed patients with OXPHOS disorders, and controls. In combination with cytofluorometric detection of mitochondrial membrane potential and protein analysis by SDS and native electrophoreses, it was possible to diagnose specific defects of Complexes I, IV and V using small amounts of peripheral blood within 1-2 days. Importantly, functional manifestations of mitochondrial disorders caused by SURF1  and TMEM70  mutations in lymphocytes recapitulate previous findings in fibroblasts. Moreover, in contrast to cultured fibroblasts Complex IV deficiency was also manifested in lymphocytes harbouring SCO2 mutations.
The noninvasiveness, reliability and speed of such an approach demonstrate the high potential of isolated lymphocytes for diagnostics of oxidative phosphorylation disorders of nuclear origin in patients with suspected mitochondrial disease.
Labels: MiParea: Respiration, mt-Medicine, Patients
Organism: Human Tissue;cell: Blood cells, Lymphocyte Preparation: Permeabilized cells
Regulation: mt-Membrane potential Coupling state: OXPHOS Pathway: N, S, Gp, CIV HRR: Oxygraph-2k Event: C1, Oral MiP2014
1-Dep Bioenergetics, Inst Physiol, Acad Sc Czech Republic; 2-Dep Pediatrics Adolescent Medicine; 3-Clinic Pediatrics Adolescent Medicine, General Univ Hospital Prague; 4-Dep Neonatology, Thomayer Hospital; Prague, Czech Republic. - [email protected]
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