Study On The Mitochondrial DNA Mutation And The Phenotype Diversity Of Nonsyndromic Hearing Loss

Objective:1. In order to investigate the possible role of mitochondrial DNA mutations in NSHL, we analyze the incidence, character of three types of mitochondrial DNA mutations in the nonsyndromic hearing loss (NSHL) patients and their clinical data, healthy control subjects were also analyzed simultaneously.2. To study the correlation between the number of mtDNA copies containing mtDNA A1555G mutation site and the phenotype, and further elucidate the molecular genetic basis of the phenotype diversity of NSHL.Methods:1. PCR-RFLP, directional sequencing of PCR products were applied to detect the mutations of mtDNA 1555, mtDNA 3243 and mtDNA 7445 sites in 486 patients with nonsyndromic hearing loss and 100 normal controls.2. Real time-Amplification refractory mutation system-quantitative PCR was established to detect the number of mtDNA copies containing mild type and mutant type mtDNA 1555.3. SPSS 10.0 package was used to analyze the correlation between the number of mtDNA copies containing mtDNA A1555G mutation site and the phenotype.Results:1. Among the total 486 patients, the mutation rate of the mtDNA A1555G is 9.05%(44/486), 32 of 44 cases are homozygosis, 12 of 44 cases are heterozygosis. mtDNA A3243G, mtDNA A7445G mutation were not detected. The same mutation was not detected in the control subjects.2. Among the 113 people of 7 pedigrees, 52 people were found to harbor the mtDNA A1555G mutation, all of them were from the maternal side. 45 of 52 cases are homozygosis, 7 of 52 cases are heterozygosis. The clinical phenotype of these 52 people were from normal to serious hearing loss. These pedigrees were hypersensitivity to AmAn ototoxicity. 3. The results of mtDNA 1555 copy number detected by real time- Amplification Refractory Mutation System-quantitative PCR are stable and accurate.4.There was not significant correlation between mtDNA 1555 homogenicity mutation copies and phenotype (R=0.001,P=0.997) in diverging group, there was significant correlation between mtDNA 1555 heterogenicity mutation copies and phenotype (R=0.771,P=0.003) in diverging group, there was significant correlation between mtDNA 1555 homogenicity mutation copies and phenotype (R=0.341,P=0.022) in family group, there was significant correlation between mtDNA 1555 heterogenicity mutation copies and phenotype (R=0.85,P=0.015) in family group.Conclusions:1. The mutation rate of the mtDNA A1555G is high in the Chinese NSHL patients, the mutation type conclude homozygosis and heterozygosis. We found a NSHL family carring mtDNA G7444A mutation.2. The results of mtDNA 1555 copy number detected by real time-Amplification Refractory Mutation System-quantitative PCR are stable and accurate.3. There is significant correlation between mtDNA A1555G copies and the severity of hearing loss.