Molecular Mechanism Of Reactive Oxygen Species Damage To Cochlea

The mitochondria are one of the major sources of endogenous reactiveoxygen species (ROS). Because there are a lack of the protection of histone-likeproteins and the systems of repairing DNA damage, mitochondrial DNA (mtDNA)is a sensitive target for ROS damage. Several primary studies suggested that micelacking ROS defensive enzymes such as superoxide dismutases (SODs) increasedROS to the vulnerability of cochlea. To investigate the molecular mechanism ofROS damage to mtDNA, We detected the cochlear mtDNA in Cu/ZnSOD gene(Sod1) Knockout mice and in wild-type mice (CD129/CD1) by nested polymerasechain reaction (PCR) and discussed from following four parts:Part one Three delstions are detectable in cochlearmitochondrial DNA of Cu/Zn superoxide dismutase geneknockout miceObjective To investigate the molecular mechanism of ROS damage tocochlea. Methods 21 cochleae of Cu/ZnSOD gene (Cu/Zn superoxide dismutasegene,Sod1) knockout mice including 14 cochleae of homozygous mutant mice(no measurable Cu/ZnSOD activity, Sod1-/-) and 7 cochleae of heterozygousMutant mice (50% reduction of Cu/ZnSOD activity Sod1+/-) and l6 cochleae ofwild-type mice (100% Cu/ZnSOD activity, Sod1+/+) were detected by nestedpolymerase chain reaction (PCR). Results 1. Three deletions were detectable incochlear mtDNA of Sod1 knockout mice. MtDNA3867bp deletion was observedeasily in Sod1-/- mice and Sod1+/+ mice, mtDNA3726bp deletion was the mostvisible in Sodl+/- mice, mtDNA236bp deletion was barely detected in all ofndce. 2. The imidene of mtDNA delehons and the ratio of deloted mtoNA/otalM in Sod1-/whce (100% and 6.284X 10', resPectively) were sighficanlyhigher than in Sodl+/+ mice (31.25% and 0.273 X 10', resPectively). There wasno difference betWen Sod1-/- mice and Sod1+/- mice, and between Sod1+/- ndceand Sod1+/+ ndce in the incidence of un deletions. Howmp the ratio ofdeleted theotal tntDN was higher in Sod1-/- comPared tO Sod1+/ mce,and in Sod1+/- mice comPared tO Sod1+/+ mice. Conclusions Sod1 deficencycan lead to three de1etions in the cochlear mtDN of ndce. MultiPle deletions ofmtoNA influence the functon of en-zyInes coding OXPHOS, it may p1ay antheorttal role in heedng loss. p'' Part tWo Deletions are easy datectable in cochlearwhochondrial DNA of 'Cu/Zn' superotide dismutase geneknockOut ndce -- Study of tissue sPecificity of ROS dtuage tomitochondrial DNAObjective To invest1gate the tissue sPecificity of ROS damage tondtochondrial DNA m and to deennin whether cochlear mtoNA is asenslere target fOr ROS daxnage. Methods 10 CchSOD gene (Cu/ZnsuPeroxide diSmuase gene, Sod1) bockoul mce and 16 wild-tyPe mice weredetected by nested polytnerase chain reaction RCR). Reults 1 Tbree deletionswere dctectable in various tissues of Sod1 bockoat mice. MtDNA3867bPdeletion and mtDNA3726bP was the most visible in vedous hssues,MNA4236bP deletion was barely 4eected in vedous organ. 2. Ther wereobvious differnces in the ratio of delcted theQtal mtoNA of various organs.Deleted mtDNA was most abundance in the liver and kidney, and less abUndan incochlea. heart and brain, and the most low abUndant in SPleen and dri. The ratioof various tissues were about 3~20 tirns in Sod1 bockout nde than in wild-tyPemice. Ih the cochea, the rat1o was about l5 tims. ConclIIstons Sod1 deficiencycan lead to mtDN deletions in vndous tissues, bul ROS damage tO on mtoNAon vedous tissues is sighficAn differellt, CQChlear im is a sensitive target forROS daxnage.Part three Cochlear mitochondrial DNA 3867bp deletion inaging miceObjectiVe To study the re1ationshiP between Cochlear mitochondrial DNA(IntoNA 3867bp deletion and presbycusis, and to deterInine the lOcation of- mtoNA de1etion in aging ndce. Msthods We deected the cochlear amNA in 2,7~1Q and 17~19 month-old mice by nested polyInerase chain reachon (PCR) andDNA Sequencing. Results 1. IntDN 3867bP deletions were deectable in thecochieae of aging mice.