Prenatal diagnosis of mitochondrial disease: analysis of 28 pedigrees with mitochondrial DNA mutation

Abstract

Objective To perform a prenatal diagnosis for the second fetuses from 28 pedigrees with proband of mitochondrial disease due to mitochondrial DNA (mtDNA) mutation. Methods From April 2011 to November 2015, peripheral blood samples of 28 probands and their parents, urine samples of these probands and their mothers as well as amniotic fluid samples of the second fetuses from the 28 pedigrees were collected in Peking University First Hospital. DNA sequencing was used to identify mtDNA mutations. Polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) was performed to verify mutation sites, calculate mutation loads, and further confirm the diagnosis after birth. Microsatellite maker analysis was also performed on five short tandem repeats located in nuclear genes to exclude maternal contamination. Statistical analysis was carried out using independent t-test. Results In the 15 pedigrees carrying A3243G mutation, 13 mothers and nine fetuses carried A3243G mutation. Neither the other two mothers nor their fetuses were positive for A3243G mutation. Among the 12 pedigrees with T8993G mutation, there were eight mothers carrying T8993G mutation and all of their fetuses carried the same mutation; and the other four mothers and their fetuses were negative for T8993G mutation. T10191C mutation was only found in one proband and the second fetus of that pedigree, but not in the mother. None of the fathers had mtDNA mutation. Results of PCR-RFLP were consistent with those of DNA sequencing. Short tandem repeat analysis demonstrated that amniocyte samples were from fetuses without maternal contamination. No mtDNA mutations were found in the six newborns who were negative for mtDNA mutations in prenatal diagnosis. The mean mutation load in urine samples of the six mothers without A3243G mutation in amniocytes was significantly lower than that of the nine mothers with A3243G mutation [(10.1±4.8) % vs (28.2±15.1) %, t=2.290, P=0.043]. Conclusions The lower the mtDNA mutation load in maternal urine samples, the less the possibility she bears a child with mtDNA mutation. However, prenatal diagnosis of mitochondrial disease is necessary. Key words: Mitochondrial diseases; Pedigree; Genes, Mitochondrial; Mutation; Prenatal diagnosis