Single-fiber PCR in MELAS(3243) patients: Correlations between intratissuedistribution and phenotypic expression of the mtDNAA(3243G) genotype

We performed morphological, biochemical, and genetic studies, including sin gle-fiber PCR (sf PCR), on muscle biopsies obtained from a mother and daugh ter with MELAS syndrome due to the A3243G transition of mitochondrial DNA ( mtDNA). The severity of muscle involvement appeared quite distinct, in spit e of the fact that both patients segregated similar mutant mtDNA levels on total muscle DNA. The daughter did not show any clinical muscle involvement : muscle biopsy revealed many ragged red fibers (RRFs) mostly positive for cytochrome-c oxidase (COX) activity. In contrast, her mother had developed a generalized myopathy without progressive external ophthalmoplegia (PEO), morphologically characterized by many COX-negative RRFs. Single-muscle fibe r PCR demonstrated in both patients significantly higher percentages of wil d-type mtDNA in normal fibers (daughter: 23.25 +/- 15.22; mother: 43.13 +/- 26.11) than in COX-positive RRFs (daughter: 11.25 +/- 5.22, P < 0.005; mot her: 9.12 +/- 5.9, P < 0.001) and in COX-negative RRFs (daughter: 8.9 +/- 4 .2, P < 0.001 mother: 4.8 +/- 2.8, P < 0.001). Wild-type mtDNA levels resul ted higher also in COX-positive vs. COX-negative RRFs (daughter: P < 0.05; mother: P < 0.001). Our data confirm a direct correlation between A3243G le vels and impairment of COX function at the single-muscle fiber level. Moreo ver, the evidence of a clinical myopathy in the patient with higher amounts of COX-negative RRFs bolsters the concept that a differential distribution of mutant mtDNAs at the cellular level may have effects on the clinical in volvement of individual tissues. However, the occurrence of a similar morph ological and biochemical muscle phenotype also in PEO3243 patients suggests that other genetic factors involved in the interaction between mitochondri al and nuclear DNA, rather than the stochastic distribution of mtDNA genome s during embryogenesis, are primarily implicated in determining the various clinical expressions of the A3243G of mtDNA. (C) 2000 Wiley-Liss, Inc.