Deliang Huang

GJB2 mutation spectrum in 2063 Chinese patients with nonsyndromic hearing impairment

BackgroundMutations in GJB2 are the most common molecular defects responsible for autosomal recessive nonsyndromic hearing impairment (NSHI). The mutation spectra of this gene vary among different ethnic groups.MethodsIn order to understand the spectrum and frequency of GJB2 mutations in the Chinese population, the coding region of the GJB2 gene from 2063 unrelated patients with NSHI was PCR amplified and sequenced.ResultsA total of 23 pathogenic mutations were identified. Among them, five (p.W3X, c.99delT, c.155_c.158delTCTG, c.512_c.513insAACG, and p.Y152X) are novel. Three hundred and seven patients carry two confirmed pathogenic mutations, including 178 homozygotes and 129 compound heterozygotes. One hundred twenty five patients carry only one mutant allele. Thus, GJB2 mutations account for 17.9% of the mutant alleles in 2063 NSHI patients. Overall, 92.6% (684/739) of the pathogenic mutations are frame-shift truncation or nonsense mutations. The four prevalent mutations; c.235delC, c.299_c.300delAT, c.176_c.191del16, and c.35delG, account for 88.0% of all mutantalleles identified. The frequency of GJB2 mutations (alleles) varies from 4% to 30.4% among different regions of China. It also varies among different sub-ethnic groups.ConclusionIn some regions of China, testing of the three most common mutations can identify at least one GJB2 mutant allele in all patients. In other regions such as Tibet, the three most common mutations account for only 16% the GJB2 mutant alleles. Thus, in this region, sequencing of GJB2 would be recommended. In addition, the etiology of more than 80% of the mutant alleles for NSHI in China remains to be identified. Analysis of other NSHI related genes will be necessary.

Comprehensive molecular etiology analysis of nonsyndromic hearing impairment from typical areas in China

BackgroundEvery year, 30,000 babies are born with congenital hearing impairment in China. The molecular etiology of hearing impairment in the Chinese population has not been investigated thoroughly. To provide appropriate genetic testing and counseling to families, we performed a comprehensive investigation of the molecular etiology of nonsyndromic deafness in two typical areas from northern and southern China.MethodsA total of 284 unrelated school children with hearing loss who attended special education schools in China were enrolled in this study, 134 from Chifeng City in Inner Mongolia and the remaining 150 from Nangtong City in JiangSu Province. Screening was performed for GJB2, GJB3, GJB6, SLC26A4, 12S rRNA, and tRNAser(UCN)genes in this population. All patients with SLC26A4 mutations or variants were subjected to high-resolution temporal bone CT scan to verify the enlarged vestibular aqueduct.ResultsMutations in the GJB2 gene accounted for 18.31% of the patients with nonsyndromic hearing loss, 1555A>G mutation in mitochondrial DNA accounted for 1.76%, and SLC26A4 mutations accounted for 13.73%. Almost 50% of the patients with nonsyndromic hearing loss in these typical Chinese areas carried GJB2 or SLC26A4 mutations. No significant differences in mutation spectrum or prevalence of GJB2 and SLC26A4 were found between the two areas.ConclusionIn this Chinese population, 54.93% of cases with hearing loss were related to genetic factors. The GJB2 gene accounted for the etiology in about 18.31% of the patients with hearing loss, SLC26A4 accounted for about 13.73%, and mtDNA 1555A>G mutation accounted for 1.76%. Mutations in GJB3, GJB6, and mtDNA tRNAser(UCN)were not common in this Chinese cohort. Conventionally, screening is performed for GJB2, SLC26A4, and mitochondrial 12S rRNA in the Chinese deaf population.

Distinct and novel SLC26A4/Pendrin mutations in Chinese and U.S. patients with nonsyndromic hearing loss

Mutations of the human SLC26A4/PDS gene constitute the most common cause of syndromic and nonsyndromic hearing loss. Definition of the SLC26A4 mutation spectrum among different populations with sensorineural hearing loss is important for development of optimal genetic screening services for congenital hearing impairment. We screened for SLC26A4 mutations among Chinese and U.S. subjects with hearing loss, using denaturing HPLC (DHPLC) and direct DNA sequencing. Fifty-two of 55 Chinese subjects with deafness accompanied by enlargement of the vestibular aqueduct (EVA) exhibited at least one mutant SLC26A4 allele, whereas SLC26A4 mutations were found in only 2 of 116 deaf Chinese patients without EVA. The spectrum of SLC26A4 mutations differed among Chinese and U.S. subjects and included 10 previously unreported SLC26A4 variants: 4 in the Chinese population (p.E303Q, p.X329, p.X467, p.X573) and 6 in the U.S. population (p.V250A, p.D266N, p.F354S, p.D697A, p.K715N, p.E737D). Among the seven novel in-frame missense mutations, five encoded SLC26A4 proteins with substantially reduced Cl(-)/anion exchange activity as expressed and measured in Xenopus oocytes, but four of these were sufficiently active to allow study of anion selectivity. The only mutant polypeptide exhibiting complete loss of anion exchange function, p.E303Q, was expressed at or near the oocyte surface at near-wild-type levels. Two variants, p.F354S and p.E737D, displayed selective reduction in relative rate of Cl(-)/HCO(3)(-) exchange compared with similarly measured rates of Cl(-)/Cl(-) and Cl(-)/I(-) exchange. Our data show that mutation analysis of the SLC26A4 gene is of high diagnostic yield among subjects with deafness and bilateral EVA in both China and the U.S. However, the pathogenicity of monoallelic SLC26A4 gene variants in patients with hearing loss remains unclear in many instances.

The prevalence of the 235delC GJB2 mutation in a Chinese deaf population

Purpose: Mutations in the GJB2 gene are the most frequently found mutations in patients with nonsyndromic hearing impairment in populations studied to date. However, the prevalence of mutations varies among different ethnic groups. In most areas of China, genetic testing for nonsyndromic hearing impairment is currently not available because of the lack of information regarding the molecular cause of nonsyndromic hearing impairment. The purpose of this study is to determine the prevalence of a common GJB2 mutation, 235delC, in Chinese deaf children.Methods: We collected DNA specimens from 3004 patients with nonsyndromic hearing impairment from 26 regions of China; 368 Han Chinese and 98 Uigur controls, and screened for the 235delC mutation. The coding exon of the GJB2 gene was polymerase chain reaction amplified, followed by restriction enzyme digestion with ApaI and analysis by agarose gel.Results: Overall, 488 patients (16.3%) were determined to carry at least one 235delC mutant allele, with 233 (7.8%) homozygotes and 255 (8.5%) heterozygotes. Therefore, within the subpopulations examined, the frequency varies from 0% to 14.7% for 235delC homozygotes and from 1.7% to 16.1% for heterozygotes. On the basis of this survey of the patient cohort as stated, Chinese patients with nonsyndromic hearing impairment appear to have a relatively higher 235delC frequency than that of other Asian populations.Conclusion: These results demonstrate that an easy and fast genetic testing method for this well-known GJB2 gene mutation can be made available for at least 2 million Chinese patients and family members with nonsyndromic hearing impairment. By screening for the common GJB2 235delC mutation, the molecular cause in as high as 15% of patients with nonsyndromic hearing impairment in certain regions of China can be identified. In addition, patients who are negative for the 235delC mutation would be candidates for further mutational analysis of GJB2 or other deafness-related genes.

Molecular Etiology of Hearing Impairment in Inner Mongolia: mutations in SLC26A4 gene and relevant phenotype analysis

BackgroundThe molecular etiology of hearing impairment in Chinese has not been thoroughly investigated. Study of GJB2 gene revealed that 30.4% of the patients with hearing loss in Inner Mongolia carried GJB2 mutations. The SLC26A4 gene mutations and relevant phenotype are analyzed in this study.MethodsOne hundred and thirty-five deaf patients were included. The coding exons of SLC26A4 gene were sequence analyzed in 111 patients, not including 22 patients carrying bi-allelic GJB2 mutations or one patient carrying a known GJB2 dominant mutation as well as one patient with mtDNA 1555A>G mutation. All patients with SLC26A4 mutations or variants were subjected to high resolution temporal bone CT scan and those with confirmed enlarged vestibular aqueduct and/or other inner ear malformation were then given further ultrasound scan of thyroid and thyroid hormone assays.ResultsTwenty-six patients (19.26%, 26/135) were found carrying SLC26A4 mutation. Among them, 17 patients with bi-allelic SLC26A4 mutations were all confirmed to have EVA or other inner ear malformation by CT scan. Nine patients were heterozygous for one SLC26A4 mutation, including 3 confirmed to be EVA or EVA and Mondini dysplasia by CT scan. The most common mutation, IVS7-2A>G, accounted for 58.14% (25/43) of all SLC26A4 mutant alleles. The shape and function of thyroid were confirmed to be normal by thyroid ultrasound scan and thyroid hormone assays in 19 of the 20 patients with EVA or other inner ear malformation except one who had cystoid change in the right side of thyroid. No Pendred syndrome was diagnosed.ConclusionIn Inner Mongolia, China, mutations in SLC26A4 gene account for about 12.6% (17/135) of the patients with hearing loss. Together with GJB2 (23/135), SLC26A4 are the two most commonly mutated genes causing deafness in this region. Pendred syndrome is not detected in this deaf population. We established a new strategy that detects SLC26A4 mutations prior to the temporal bone CT scan to find EVA and inner ear malformation patients. This model has a unique advantage in epidemiologic study of large deaf population.

SLC26A4 c.919-2A>G varies among Chinese ethnic groups as a cause of hearing loss

Purpose: Mutations in the SLC26A4 gene are second only to GJB2 mutations as a currently identifiable genetic cause of sensorineural hearing loss. In most areas of China, genetic testing for sensorineural hearing loss is unavailable because of limited knowledge of the mutation spectrum. Although SLC26A4 c.919-2A>G (IVS7–2A>G) is a common mutation among some Asian populations, the mutation prevalence among various ethnic groups within China has not been studied.Methods: DNA specimens from 3271 subjects with moderate to profound sensorineural hearing loss from 27 regions of China were genotyped for the c.919-2A>G mutation by polymerase chain reaction/restriction-fragment-length polymorphism. Normal hearing controls from Han (n = 185) and Uigur (n = 152) populations were also tested.Results: Overall, 408 subjects with sensorineural hearing loss (12.5%) carried at least one c.919-2A>G allele, with 158 (4.8%) homozygotes and 250 (7.6%) heterozygotes. Within the subpopulations examined, the rate varies from 0% to 12.2% for c.919-2A>G homozygotes and from 0% to 17.6% for heterozygotes. Based on this cohort, Chinese subjects with sensorineural hearing loss seem to have a relatively higher c.919-2A>G frequency than that of other Asian populations.Conclusion: These results demonstrate that a simple and efficient genetic test for the c.919-2A>G mutation alone would identify the molecular cause in up to 8–12% of individuals with sensorineural hearing loss in a few eastern and central regions of China. Those who are negative for the c.919-2A>G mutation would be candidates for further mutational analysis of SLC26A4 or other deafness-related genes. This would greatly improve genetic diagnosis and counseling for a huge number of Chinese individuals and family members with sensorineural hearing loss in China, and many more ethnic Chinese in other countries, which might be up to one million.

Prevalence of the GJB2 IVS1+1G >A mutation in Chinese hearing loss patients with monoallelic pathogenic mutation in the coding region of GJB2.

BackgroundMutations in the GJB2 gene are the most common cause of nonsyndromic recessive hearing loss in China. In about 6% of Chinese patients with severe to profound sensorineural hearing impairment, only monoallelic GJB2 mutations known to be either recessive or of unclear pathogenicity have been identified. This paper reports the prevalence of the GJB2 IVS1+1G>A mutation in a population of Chinese hearing loss patients with monoallelic pathogenic mutation in the coding region of GJB2.MethodsTwo hundred and twelve patients, screened from 7133 cases of nonsyndromic hearing loss in China, with monoallelic mutation (mainly frameshift and nonsense mutation) in the coding region of GJB2 were examined for the GJB2 IVS1+1G>A mutation and mutations in the promoter region of this gene. Two hundred and sixty-two nonsyndromic hearing loss patients without GJB2 mutation and 105 controls with normal hearing were also tested for the GJB2 IVS1+1G>A mutation by sequencing.ResultsFour patients with monoallelic mutation in the coding region of GJB2 were found carrying the GJB2 IVS1+1G>A mutation on the opposite allele. One patient with the GJB2 c.235delC mutation carried one variant, -3175 C>T, in exon 1 of GJB2. Neither GJB2 IVS1+1G>A mutation nor any variant in exon 1 of GJB2 was found in the 262 nonsyndromic hearing loss patients without GJB2 mutation or in the 105 normal hearing controls.ConclusionTesting for the GJB2 IVS 1+1 G to A mutation explained deafness in 1.89% of Chinese GJB2 monoallelic patients, and it should be included in routine testing of patients with GJB2 monoallelic pathogenic mutation.

Common Molecular Etiologies Are Rare in Nonsyndromic Tibetan Chinese Patients with Hearing Impairment

Background Thirty thousand infants are born every year with congenital hearing impairment in mainland China. Racial and regional factors are important in clinical diagnosis of genetic deafness. However, molecular etiology of hearing impairment in the Tibetan Chinese population living in the Tibetan Plateau has not been investigated. To provide appropriate genetic testing and counseling to Tibetan families, we investigated molecular etiology of nonsyndromic deafness in this population. Methods A total of 114 unrelated deaf Tibetan children from the Tibet Autonomous Region were enrolled. Five prominent deafness-related genes, GJB2, SLC26A4, GJB6, POU3F4, and mtDNA 12S rRNA, were analyzed. Inner ear development was evaluated by temporal CT. A total of 106 Tibetan hearing normal individuals were included as genetic controls. For radiological comparison, 120 patients, mainly of Han ethnicity, with sensorineural hearing loss were analyzed by temporal CT. Results None of the Tibetan patients carried diallelic GJB2 or SLC26A4 mutations. Two patients with a history of aminoglycoside usage carried homogeneous mtDNA 12S rRNA A1555G mutation. Two controls were homozygous for 12S rRNA A1555G. There were no mutations in GJB6 or POU3F4. A diagnosis of inner ear malformation was made in 20.18% of the Tibetan patients and 21.67% of the Han deaf group. Enlarged vestibular aqueduct, the most common inner ear deformity, was not found in theTibetan patients, but was seen in 18.33% of the Han patients. Common molecular etiologies, GJB2 and SLC26A4 mutations, were rare in the Tibetan Chinese deaf population. Conclusion The mutation spectrum of hearing loss differs significantly between Chinese Tibetan patients and Han patients. The incidence of inner ear malformation in Tibetans is almost as high as that in Han deaf patients, but the types of malformation vary greatly. Hypoxia and special environment in plateau may be one cause of developmental inner ear deformity in this population.

A de novo GJB2 (Connexin 26) Mutation, R75W, in a Chinese Pedigree With Hearing Loss and Palmoplantar Keratoderma

A de novo GJB2 (Connexin 26) Mutation, R75W, in a Chinese Pedigree With Hearing Loss and Palmoplantar Keratoderma Yongyi Yuan, Deliang Huang, Fei Yu, Xiuhui Zhu, Dongyang Kang, Huijun Yuan, Dongyi Han, and Pu Dai* Department of Otolaryngology, PLA General Hospital, Beijing, People’s Republic of China Department of Otolaryngology, Chifeng Second Hospital, Chifeng City (Inner Mongolia), People’s Republic of China

Rapid screening for the mitochondrial DNA C1494T mutation in a deaf population in China using real-time quantitative PCR.

Abstract Conclusion: Real-time quantitative polymerase chain reaction (qPCR) with a TaqMan minor groove binding (MGB) probe is useful for large-scale screening for the C1494T mutation. The mitochondrial DNA(mtDNA) C1494T mutation has a low carrier frequency in Chinese patients with nonsyndromic hearing loss. Objective: To develop a simple, rapid, and reliable real-time qPCR assay based on TaqMan technology using a new MGB probe for detecting the mtDNA C1494T mutation directly, and to investigate the carrier frequency in nonsyndromic deaf Chinese subjects. Methods: A TaqMan-MGB probe was constructed. Peripheral blood samples were collected from 3133 nonsyndromic deaf patients and genomic DNA was extracted. A real-time qPCR using MGB probes (wild-type) in a single tube was used to detect the mtDNA C1494T mutation. The results were then compared to the DNA sequence of the PCR products. Results: A total of 13 of 3133 (0.4%) Chinese nonsyndromic hearing loss patients were C1494T-positive. The results of the TaqMan-MGB probe method were consistent with those of sequencing.