Analysis Of Mitochondrial DNA Mutation In Leber Hereditary Optic Neuropathy (LHON) In Two Chinese Pedigrees

BackgroundLeber hereditary optic neuropathy(LHON.OMIM:535000)is generally regarded as a maternally transmitted disorder presented predominantly in young men.The disease has been associated with many missense mutations in the mtDNA that can act autonomously or in association with each other to cause the disease.LHON patients present with mid-life,acute or subacute,painless,central vision loss leading to central scotoma.Neuroophthalmologic examination commonly reveals peripapillary telangiectasia,microangiopathy,disc pseudoedema,and vascular tortuosity;these features are observed in 58%of patients with the nucleotide pair(np)11778 mutation and occasionally in their asymptomatic maternal relatives. The mean age of onset has been variously reported from 27 to 34 years with a range of 1 to 70 years.The eyes can be affected simultaneously or sequentially,with an average interval between eyes being affected of about 2 months.In addition to visual loss,some patients developed kinds of complicated neurologic syndromes.While LHON is traditionally considered to be familial,many individuals represent isolated cases.The proportion of cases with family histories have been reported to be 43%for np 11778,to be 78%for np 3460,and to be 65%for np 14484.Since Wallace firstly discovered the mutation G11778A,LHON has been associated with three primary mitochondrial DNA mutations:G3460A/ND1,G11778A/ND4,and T14484C/ND6.Although the primary etiological factor of LHON is a mtDNA mutation,the presence of a primary mtDNA mutation does not necessarily lead to visual loss.The pathogenesis of LHON remains unclear.The marked incomplete penetrance and gender bias indicate that additional genetic(nuclear or mitochondfial)and epigenetic factors(smoking, drinking,et al.)may also be involved.ObjectivesTo investigate the mitochondrial factor related to LHON through sequencing and analysis the mitochondrial genome of Han Chinese patients with LHON.PatientsLHON probands and their maternal relatives in 2 Chinese families from Zhejiang Province and 10 matched healthy controls.Methods(1)The patients and matched healthy controls were collected,genomic DNA from peripheral venous blood was extracted using the phenol-chloroform method.(2)LHON patients in two Chinese families were selected,amplifying their whole sequence of mtDNA, and bidirection sequencing with the universal M13 primer.(4)DNA sequence analysis and variation identification by DNAssist and Chromas 2 software.Then compare with the authority databases such as mitomap.Results(1)Mutational analysis of mtDNA in these two Chinese pedigrees revealed the presence of one common LHON associated G11778A mutation.(2)There is one consistency mutation in pedigree 1:3497C＞T(Ala→Val)and 3571C＞T(Leu→Phe).Mutation 3571C＞T has not been reported.There were four consistency mutations in pedigree 2:10398A＞G(Thr→Ala),14502T＞C(Ile→Val),14766C＞T(Thr→Ile) and 15326A＞G(Thr→Ala).In addition,there were three mutations:73A＞G,94G＞A and 263A＞G belong to the polymorphic nucleotide positions from the mtDNA control region.Conclusions(1)mtDNA 11778G＞A(Arg→His)/ND4 mutation is still the most significant cause of LHON in these two pedigrees.(2)Mutational analysis of mtDNA in pedigree 1 revealed the presence of one consistency mutation:3497C＞T(Ala→Val)and 3571C＞T(Leu→Phe).They are both at mutation hot spot ND1.They may affect respiratory chain in coordination with primary mutation.Mutation 3571C＞T(Leu→Phe)has not been reported.Further research should be done.(3)There were four consistency mutations in pedigree 2:10398A＞G(Thr→Ala), 14502T＞C(Ile→Val),14766C＞T(Thr→Ile)and 15326A＞G(Thr→Ala).ND1,ND3,ND6 and CYB assist electron transmission and oxidative phosphorylation to a great extent in the respiratory chain.They are all nicotinamide adenine dinucleotide(NADH)subunits.NAD~+ accept 2H of most metabolite and form NADH,then these 2H were transmitted to ubiquinone by mitochondrial respiratory complexⅠ.We suggested that this mutation: 14502T＞C(Ile→Val)influence the mitochondrial respiratory complexⅠ,lead to the mitochondrial respiratory chain dysfunction resulting in optic atrophy of the patients.Further researches on the molecular consequence of these mutations are needed to confirm the mutation.