The Relationship Between Phenotype And Genotype For Late-onset Progressive Hearing Loss

Objective: Hearing loss is the most common perception disorder in theworld. The main factors is thought to be hereditary, and mutated genesassociated with deafness are not uncommon in the normal group. That may beone of the important causes of high incidence of the deafness. Tough lots ofgenes were found associated with hereditary deafness, there are still lots ofdeaf patients unable to determine their causes. In this study, we have recruited5Chinese families with late-onset hearing loss. We analyzed the relationshipbetween phenotype and genotype of the five families and found new mutatedgenes to enrich the hereditary deafness pathogenic gene pool.Methods: We collect the five family’s information and numbered:HBZ-01, HBZ-02, HBX, HBD, HBM. Genomic DNA was extracted fromperipheral blood leukocytes using a commercially available DNA extractionkit （Qiagen Inc, Valencia, CA） according to the manufacturer’s instructions.Mutation screening was conducted using polymerase chain reaction （PCR）amplification and the exons were directly sequenced. Common hearing-relatedgenes including gap junction beta2（GJB2）, SLC26A4, mitochondrial DNA12S rRNA, GJB3and GJB6were examined in the family. If the pathogenicmutations were identified, the resultant sequence data were compared with theevaluation samples for alignment with the National Center for BiotechnologyInformation reference （NCBI） sequence. If not, according to the phenotype ofthe affected individuals and the gene mutations about dominant hereditaryhearing loss which were discovered previously, screening the may candidategenes, if positive, similar with previous. If still not, the next generation ofdeafness high-throughput genome sequencing was conducted using DNAsamples of the family. Candidate mutation was confirmed by Sangersequencing. Results:1In the family of HBZ-01, the reported onset of hearing problems inmost cases （excluding II-5） was in the second decade of life with subsequentgradual progression from mild to severe. The hearing loss in these casesaffected high frequencies initially and then developed to the middle and lowfrequencies. Except for individual II-5（26years） who was affected withprelingual deafness and no define nosogenesis. Pathogenic mutations in theGJB2gene were identified. The affected mother showed a heterozygousG→A transition at nucleotide232, resulting in an alanine to threoninesubstitution at codon78（p.A78T）, and the normal hearing father had ac.35insG insertion mutation. The three affected children displayedheterozygosity for the GJB2mutations, showing a previously unreportedcombination of c.35insG and c.232G>A.2In the family of HBZ-02, the clinical history and audiological findingsof hearing loss in the family members revealed a form of post-lingual bilateralSNHI. The reported onset of hearing problems in most cases was in the5thdecade of life, with the loss progressing over time. The hearing loss in thesecases affected high frequencies initially, expanding to all frequencies at a laterstage. Most of affected subjects have tinnitus and some of them experiencedvestibular disorder. Finally, two novel COCH mutations detected in thisfamily are heterozygous c.226G>A in exon4and c.1055C>G mutation inexon11. The heterozygous of c.226G>A which resulting in a alanine tothreonine substitution at residue76of the COCH protein （p.A76T）. Thisalanine residue is evolutionarily conserved in several species and50unrelatedrandom individuals of Chinese origin were analyzed. None of them showedthis mutation. The c.1055C>G resulting in a threonine to serine substitution atresidue352of the COCH protein （p.T352S）. Due to this mutation was foundin normal individuals in this family, so the p.T352S variation is suggested as apolymorphism.3In the family of HBX, we first discovered the EYA4gene mutation inChinese, and the discovery is a new mutation. The mutated gene is c.544₅45insA in exon8, which resulting in a frameshift mutation （p.E182X）.Unlike previous reported families, the family’s hearing curve is not typicalvalley type but flat or steep fall. The onset of hearing loss between20⁴0years, most of affected subjects have tinnitus and no experienced vestibulardisorder.4The rest of the two families （HBM, HBD） found no clear pathogenicgenes. The genetic still has limitation, there are a large number of deafnessgenes are not found.Conclusions:1Due to heterogeneity （1） individually rare mutations collectively play asubstantial role in causing complex illnesses;（2） the same gene may harbormany different rare severe mutations in unrelated affected individuals;（3） thesame mutation may lead to different clinical phenotypes in differentindividuals;（4） mutations in different genes in the same or related pathwaysmay lead to the same disorder.2Phenotypes can guide the diagnose of genotypes.3To date, there are lots of deafness genes among the general deafpopulation are unknown, hereditary deafness gene pool still to be enriched.