mTOR/NF-κB signaling pathway protects hippocampal neurons from injury induced by intermittent hypoxia in rats.

OBJECTIVE: To expound the roles of mTOR and NF-kB signaling pathway in intermittent hypoxia (IH)-induced damage of hippocampal neurons. METHODS: For rat experiments, mTOR inhibitor (Rapamycin, Rapa) and NF-κB signaling inhibitor (ammonium pyrrolidine dithiocarbamate, PDTC) were applied to inhibit mTOR and NF-κB signaling, respectively. For neuron experiments, hippocampal neurons from rat were successfully cultured. Different concentrations of Rapa and PDTC were added to the cultured hippocampal neurons. Rat or primary hippocampal neurons were exposed to normoxic or IH conditions after administration of Rapa and PDTC. The effects of Rapa and PDTC administration on learning and memory ability of rats and hippocampal injury after IH exposure were assayed by Morris water maze and H&E staining. Electron microscope was utilized to examine primary hippocampal neuron ultrastructure changes after IH exposure and Rapa or PDTC administration. The expressions of NF-κB-p65, IκBα, IKKß, BDNF, TNF-α, IL-1ß, PSD-95 and SYN in hippocampal neurons were examined. RESULTS: Compared with normal control rats or neurons, IH-treated group had elevated expression levels of NF-kB, TNF-α and IL-1ß and suppressed expression level of BDNF, PSD-95 and SYN. These results were reversed upon pre-treatment with Rapa and PDTC. Furthermore, IκBα and IKKß expressions were down-regulated in IH group. No notable difference was manifested in PDTC pre-treatment group, while a prominent increase was shown after Rapa pre-administration. CONCLUSION: The administration of PDTC and Rapa could prevent IH-induced hippocampal neuron impairment, indicating that inhibition of the mTOR and NF-κB pathway may likely act as a therapeutic target for obstructive sleep apnea.

miR-325-3p Protects Neurons from Oxygen-Glucose Deprivation and Reoxygenation Injury via Inhibition of RIP3.

OBJECTIVE: Recent reports have corroborated that micro-RNAs (miRs) are related to the pathological changes of cerebral ischemia-reperfusion (CIR) induced injury. This work aimed to unearth the role and potential mechanism of miR-325-3p in regulating neuronal survival in CIR injury. METHODS: To conduct this investigation, we established an in vitro model of CIR injury by subjecting neurons to oxygen-glucose deprivation and reoxygenation (OGD/R). Gain and loss of function of miR-325-3p and receptor-interacting serine-threonine kinase 3 (RIP3) in neurons were performed to observe its effect on cell apoptosis and the release of lactate dehydrogenase. The levels of miR-325-3p and RIP3 in neurons were detected by qRT-PCR. Western blot was employed to inspect the levels of caspase3, Bax, and Bcl-2, as well as p38 and JNK phosphorylation. The relationship between miR-325-3p and RIP3 was detected by TargetScan and validated by dual-luciferase reporter assay. RESULTS: Firstly, miR-325-3p expression was obviously downregulated while RIP3 expression was upregulated in neurons following OGD/R treatment. Overexpressed miR-325-3p or downexpressed RIP3 ameliorated OGD/R-induced neuronal injury. Besides, RIP3 was a direct target mRNA of miR-325-3p. Additionally, Western blot revealed the mitogen-activated protein kinase (MAPK) pathway was involved in the regulation of miR-325-3p on OGD/R-induced neuronal injury. Furthermore, miR-325-3p was verified to hinder OGD/R-induced neuronal injury through downregulating RIP3. CONCLUSION: This study demonstrated that miR-325-3p targets RIP3 to inactivate the MAPK pathway, thereby protecting neurons against OGD/R-induced injury.

Genetic Polymorphisms and Susceptibility to Sudden Sensorineural Hearing Loss: A Systematic Review.

BACKGROUND: Recently, genetic factors have been considered as an important risk factor for sudden sensorineural hearing loss (SSNHL). Many studies analyzed the association between SSNHL and polymorphisms. However, most of them gave inconclusive results. Key Message: We performed a systematic review to find out the association between polymorphisms and susceptibility to SSNHL. Finally, 47 studies involving 5,230 SSNHL patients and 68 genes were included for analysis and discussion of results. Polymorphisms in 26 genes have been suggested to be correlated with the susceptibility to SSNHL. SUMMARY: Although a great number of studies support that polymorphisms in genes are associated with susceptibility to SSNHL, we need large multicenter studies, which evaluate multiple single nucleotide polymorphisms in SSNHL patients, to find real genetic risk factors for susceptibility to SSNHL. This is very helpful in designing more effective prevention and treatment strategies for patients with SSNHL.

Safety and Immunogenicity of a Nonadjuvant Human Papillomavirus Type 6 Virus-like Particle Vaccine in Recurrent Respiratory Papillomatosis.

OBJECTIVES: To assess the safety and immunogenicity of a nonadjuvant human papillomavirus (HPV) type 6 L1 virus-like particle (VLP) vaccine in recurrent respiratory papillomatosis (RRP) in local Chinese patients. METHODS: Patients with RRP who had undergone surgical treatment before intramuscular administration of an escalating dose of HPV type 6 L1 VLPs (1, 5, and 25 µg at 4 weekly intervals) as part of their treatment were followed up for more than 10 years. Efficacy was assessed by detecting the vaccine-induced type-specific antibody titer, calculating the intersurgical interval, and observing recurrence or remission of papillomas after receiving the vaccine. RESULTS: Nonadjuvant HPV vaccine was generally well tolerated, with no serious vaccine-related adverse episodes. It induced seroconversion for each vaccine-related HPV type. At week 12 (4 weeks after injecting 25 µg), the vaccine-induced type-specific antibody titer was significantly high. Analysis of all patients found a significant increase in the intersurgical interval and decrease in the scores. One patient (16.7%; female) experienced complete remission. Five patients (83.3%) (two males and three females) experienced partial remission. In total, complete or partial remission was achieved in six (100%) patients. CONCLUSIONS: Administration of nonadjuvant HPV type 6 L1 VLPs vaccine to RRP was generally well tolerated and highly immunogenic.

E26 Transformation-Specific Transcription Factor ETS2 as an Oncogene Promotes the Progression of Hypopharyngeal Cancer.

The E26 transformation-specific (ETS) family is one of the largest families of transcription factors. Upon activation by MAPK pathway, ETS participates in cell proliferation, differentiation, migration, apoptosis, and metastasis. However, the mechanism by which ETS is deregulated in cancer is unclear. In this study, the authors investigated the role of ETS factor, ETS2, in hypopharyngeal cancer pathogenesis in hypopharyngeal cancer tissues (N = 20) and corresponding non-neoplastic tissues (N = 20). The results showed that expression of ETS2 was increased in cancer tissues as compared with the expression in corresponding non-neoplastic tissues. Analysis of clinicopathological characteristics showed that increased level of ETS2 is associated with III-IV tumor node metastasis stage and lymph node metastasis. In addition, knockdown of ETS2 by siRNA in pharyngeal cancer cell line, FaDu, significantly decreased cell's vitality and colony-forming ability by inducing caspase-3-dependent apoptosis and cell cycle arrest. Furthermore, inhibition of ETS2 could abrogate the migration, invasion, and transforming growth factor-ß-induced epithelial mesenchymal transition through the upregulation of E-cadherin, zona occludens protein-1, together with downregulation of vimentin and α-sooth muscle actin. These functions of ETS2 could be associated with the activation of MAPK/p38/ERK/JNK signals. Taken together, the authors opined that ETS2 functions as an oncogene and plays a key role in the progression of hypopharyngeal cancer.

Psychosocial impact of dental aesthetics and desire for orthodontic treatment among Chinese undergraduate students.

OBJECTIVE: The objective of this study is to evaluate the psychosocial impact of dental aesthetics in undergraduate students in the People's Republic of China and to investigate the association between normal orthodontic treatment needs, psychosocial impact of dental aesthetics, and desire for orthodontic treatment. MATERIALS AND METHODS: A cross-sectional study was carried out in two universities in a city of the People's Republic of China with 374 young adults aged between 19 years and 24 years. The students answered a Chinese version of the Psychosocial Impact of Dental Aesthetics Questionnaire (PIDAQ) and addressed their desire for orthodontic treatment. Objective malocclusion severity was assessed with the Index of Orthodontic Treatment Need (IOTN). Statistical analysis was performed by the SPSS software (Version 15.0). RESULTS: There was no statistical sex difference in relation to the dental health component of IOTN (P=0.893) and PIDAQ scores (P=0.06), but it was found that the desire for orthodontic treatment was significantly stronger among females. The total and subscale PIDAQ scores and malocclusion severity differed significantly among the five grades of desire (P<0.01). Significant positive correlation was found among desire for orthodontic treatment, IOTN-dental health component grades, and total or subscale PIDAQ scores (P<0.01). High correlation was found between desire and PIDAQ score (r=0.93). CONCLUSION: The desire for orthodontic treatment is higher among female young adults who have the same orthodontic treatment needs compared to males. The desire for orthodontic treatment has high positive correlation with PIDAQ scores and increases with the increase in self-perceived psychosocial impacts of malocclusion and the needs for orthodontic treatment.

[Juvenile onset respiratory papillomatosis: risk factors for severity].

OBJECTIVE: This study was designed to explore the risk factors associated with severity of juvenile onset recurrent respiratory papillomatosis. METHOD: A retrospective study was conducted to study determinants of severe forms of juvenile recurrent onset respiratory papillomatosis. The patients were separated into different groups based on the onset age, the first recurrence of age, the first recurrence of period, gender and incision of tracheal respectively. The relationship among the lesion severity score,the involvement of the subregion, operation period and the next operation period were also explored. RESULT: It was observed that some children who recurred before 4 years old required more surgery, shorter operation period(the average, longest or shortest operation period) than those elder children, the differences were statistically (P=0. 029, 0. 003, 0. 010, 0. 039, respectively). The severity score of lesion was correlated positively with the involvement of the subregion and negatively with operation period (r=0. 914, -0. 451, respectively). Some children who diagnosed before 4 years old had to endure more severity score and shorter operation period than those older children, the differences were statistically (P= 0. 036, 0. 000, respectively). 8 cases accepted incision of tracheal, they accepted more surgery too. But the differences in the onset age, the first recurrence of age, and the operation period were not statistically. CONCLUSION: The results showed that the clinical course of juvenile onset recurrent respiratory papillomatosis was closely related to the first recurrence age and period, while the severity of disease was associated to the onset age and the involvement of the subregion.

[Levels of serum IL-4, IFN-gamma, IL-32 in children with juvenile-onset recurrent respiratory papillomatosis].

OBJECTIVE: To explore the relationship between serum IL-4, IFN-gamma, IL-10 levels and the aetiology of juvenile-onset recurrent respiratory papillomatosis. METHOD: Serum IL-4, IFN-gamma, IL-32 levels of 15 JORRP children were detected by use of enzyme-linked immunosorbent assay (ELISA) and compared with those of healthy control group. RESULT: Serum IL-4 levels were significantly higher in the JORRP children (P<0.01): (524.65 +/- 147.77)pg/ml in the JORRP children and (213.27 +/- 87.48) pg/ml in the healthy control group. Serum IFN-gamma levels were significantly lower in the JORRP children (P<0.01): (2.87 +/- 0.84) pg/ml in the JORRP children and (10.63 +/- 5.09) pg/ml in the healthy control group. Serum IL-32 levels were significantly lower in the JORRP children (P< 0.01): (2.47 +/- 1.60) pg/ml in the JORRP children and (9.08 +/- 2.66) pg/ml in the healthy control group. CONCLUSION: 1) While the concentration of Th2 like cytokine IL-4 in children with JORRP was higher than that in control group, the concentration of Th1 like cytokine IFN-gamma in children with JORRP was lower than that in controls, indicating that the polarization of Th1 /Th2 T cell in children with JORRP; 2) The polarization of Th1/Th2 T cell may cause the reduction of the serum IL-32 as a proinflammatory role in host immunity system that could not eradicate HPVs because of lacking enough inflammatory stimulation.

[Prevalence of common genetic mutations and clinical characteristics analysis in patients at different ages with nonsyndromic hearing impairment].

To evaluate the correlation between genetic mutations and the age in nonsyndromic hearing impairment (NSHI) and the clinical characteristics of NSHI, 215 patients with NSHI were enrolled between April 2006 and April 2012. All patients were divided into four groups according to ages of hearing loss onset and clinic presentation (0-3, 3-6, 6-18 and 18+ years). The mutations of GJB2 and mitochondria DNA (mtDNA) 1555G/C1494T were screened from peripheral blood samples in each age group. The prevalence of mutations and the age ratio were obtained. The study showed that 18.14% of all patients were found to have GJB2 mutations and 11.16% were found to have mtDNA A1555G/C1494T mutations. The prevalence of GJB2 mutation in adult group (5.26%) was lower than juvenile group who sought medical attention at 0-18 years of age (22.36%), while the prevalence of mtDNA A1555G/C1494T in adult group (31.48%) was higher than juvenile group (4.97%). Significant differences in the prevalence of GJB2 (χ2=7.108, P=0.008) and mtDNA A1555G/C1494T (χ2=20.852, P=0.000) were observed in both of two groups. The prevalence of GJB2 mutations between adult and juvenile groups according to ages of hearing loss onset was statistically significant different (0%, 20.10%, respectively, and P=0.023), while the prevalence of mtDNA A1555G/C1494T mutations was not different (14.29%, 11.34%, respectively, and P=0.698). The onset age of 66.67% of patients with GJB2 mutations was less than 1 year old, while the onset of patients with mtDNA A1555G/C1494T mutations could be found at any age group. Different standardizations of hearing loss could also show different results. These data strongly suggest that most of GJB2 mutations are found in congenital deafness and mtDNA A1555G/C1494T mutations mainly represent acquired deafness, which can be induced or aggravated by aminoglycoside antibiotics in all age groups and should be tested mainly ranging from 4 kHz to 8 kHz. Both newborn hearing screening and genetic testing are important to find early deafness.

[Analysis of genotype-phenotype correlation for GJB2 in 221 non-syndromic deafness probands and their pedigrees].

OBJECTIVE: To assess the possible genotype-phenotype correlation for GJB2. METHODS: Retrospectively analyzed GJB2 gene mutations with non-syndromic hearing impairment (NSHI) patients and their families audiological data. Individuals were grouped, according to non-truncated mutant (non-truncating, NT) and truncating mutations (truncating, T), into T/T group, T/NT group and NT/NT group. And according to whether they carry 235delC, grouped into 235delC/235delC group, 235delC/Non-235del group and Non-235delC/Non-235delC group. RESULTS: Grouped according to whether the truncation mutants:Fisher exact statistical analysis showed that the degree of hearing loss among the three groups did not meet the random distribution (P = 0.003) , T/T group was significantly higher than T/NT group (P = 0.000) and NT/NT group (P = 0.000) on the degree of hearing loss. Grouped according to whether they carry 235delC mutation: degrees of hearing loss among the three groups were statistically significant differences. Respectively pairwise comparisons (Fisher exact test) found 235delC/235delC group was significantly higher than 235delC/Non-235delC on the degree of hearing loss group (P = 0.001) and Non-235delC/Non-235delC group (P = 0.000), 235delC/Non-235delC group higher than Non-235delC/Non-235delC group (P = 0.033). In GJB2 mutations homozygous and compound heterozygous mutation genotype:G109A/G109A, 235delC/512insAACG, 299delAT/G109A and 235delC/G109A degree of hearing loss caused by genotype was significantly lower than 235delC/235delC group. CONCLUSIONS: 235delC homozygotes have significantly more hearing impairment, when compared with 235delC/non-235delC compound heterozygotes. People with two non-235delC mutations have even less hearing impairment. Patients with non-truncation mutants (G109A) suffer from lighter hearing loss than truncation mutations(235delC, 299delAT).

[Peri-operative management on juvenile recurrent respiratory papillomatosis].

OBJECTIVE: To investigate the safety of peri-operative management on children with juvenile recurrent respiratory papilloma (JORRP). METHODS: A retrospective analysis was conducted on preoperative assessment, anesthesia methods and options, operative procedure, and postoperative airway maintenance in 28 JORRP children aged from ten months to seven years old. A total of 148 times of surgery was performed on these 28 children. RESULTS: One hundred and nine JORRP children graded one and two-degree dyspnea underwent surgery within 24 hours and were intubated successfully in the first attempt after intravenous induction. Thirty-nine emergency operations were performed in the children graded three and four-degree dyspnea, 35 of them were intubated successfully in the first attempt after inhalation induction and 4 succeeded in the second attempt. No complications occurred in 129 JORRP children postoperatively, 17 children suffered from mild dyspnea and relieved after oxygen inhalation, 2 children were intubated and sent to intensive care unit because of postoperative hypoxemia. All JORRP children got through the peri-operative period safely. The quality of pronunciation in 101 children improved markedly and 35 suffered from slight hoarseness on the 1st postoperative day. Three children had the tracheal tube of tracheostomy removed after receiving five, four and three operations respectively. Nineteen children were followed up for 2 - 5 years. Among them, one child did not relapse 3 years after surgical management.One child suffered from laryngostenosis postoperatively. No death occurred. CONCLUSION: Complete preoperative preparation, rational anesthesia methods, careful operative procedure and airway maintenance after surgery could increase the safety for children with recurrent respiratory papilloma.

[Characterization of two Chinese families with aminoglycoside-induced and nonsyndromic hearing loss both carrying a mitochondrial 12S rRNA 1494C>T mutation].

OBJECTIVE: To evaluate the effect of mitochondrial DNA(mtDNA) secondary mutations, haplotypes, GJB2 gene mutations on phenotype of 1494C>T mutation, and to study the molecular pathogenic mechanism of maternally transmitted aminoglycoside-induced and nonsyndromic hearing loss. METHODS: Two Chinese Han pedigrees of maternally transmitted aminoglycoside induced and nonsyndromic hearing loss were collected. The two probands and their family members underwent clinical, genetic and molecular evaluations including audiological examinations and mutational analysis of mitochondrial genome and GJB2 gene. RESULTS: Clinical evaluation revealed wide range of severity, age-at-onset and audiometric configuration of hearing impairment in matrilineal relatives in both families, for which the penetrance of hearing loss was respectively 42.9% and 28.6% when aminoglycoside-induced deafness was included. When the effect of aminoglycosides was excluded, the penetrances of hearing loss were 14.3% and 14.3%. Sequence analysis of mitochondrial genomes identified a known 12S rRNA 1494C>T mutation, in addition with distinct sets of mtDNA polymorphisms belonging to Eastern Asian haplogroups C4a1a and B4b1c, respectively. CONCLUSION: Mitochondrial 12S rRNA 1494C>T mutation probably underlie the deafness in both families. Lack of significant mutation in the GJB2 gene ruled out involvement of GJB2 in the phenotypic expression. However, aminoglycosides and other nuclear modifier genes may still modify the phenotype of the 1494C>T mutation in these families. The B4b1c is a newly identified haplogroup in aminoglycoside-induced and nonsyndromic hearing loss family carrying the 1494C>T mutation. The 1494C>T mutation seems to have occurred sporadically through evolution.

[Maternally inherited aminoglycoside-induced and nonsyndromic hearing loss in five Han Chinese pedigrees].

OBJECTIVE: To study the effect of the mitochondrial 12S rRNA mutations on aminoglycoside-induced and nonsyndromic hearing loss, to carry out the clinical and molecular characterization of five Han Chinese pedigrees with maternally transmitted aminoglycoside-induced and nonsyndromic hearing loss. METHODS: Five pedigrees of maternally transmitted aminoglycoside-induced and nonsyndromic hearing loss were collected, genomic DNA was extracted, and complete mitochondrial genomes and the gap junction protein beta 2 (GJB2) gene were amplified and sequenced. RESULTS: Clinical evaluation revealed a wide range of severity, age-at-onset and audiometric configuration of hearing impairment in the matrilineal relatives in these families. The penetrance rates of hearing loss in these pedigrees were 17.6%, 50.0%, 66.7%, 31.3% and 23.1%, with an average of 37.7%, when aminoglycoside-induced deafness was included. Sequence analysis of the complete mitochondrial genomes in these pedigrees identified the known 1555A>G mutation and distinct sets of mitochondrial DNA(mtDNA) polymorphisms belonging to Eastern Asian haplogroups D4b2b, B4c1b1, F3, C1 and D5a, respectively. Of these variants, ND1 L89T and CO3 A200T mutations resided at the highly conservative regions. However, there were no functionally significant mutations in tRNAs and rRNAs or secondary known mutations. No hearing loss related GJB2 gene mutation was observed. CONCLUSION: The lack of significant mutation in the ruled out the possible involvement of GJB2 in the phenotypic expression of the 1555A>G mutation in those affected subjects. However, aminoglycosides, mtDNA variations and other nuclear modifier genes may play an important role in the phenotypic manifestation of the 1555A>G mutation in these Chinese families.

Frequency and spectrum of mitochondrial 12S rRNA variants in 440 Han Chinese hearing impaired pediatric subjects from two otology clinics.

BACKGROUND: Aminoglycoside ototoxicity is one of the common health problems. Mitochondrial 12S rRNA mutations are one of the important causes of aminoglycoside ototoxicity. However, the incidences of 12S rRNA mutations associated with aminoglycoside ototoxicity are less known. METHODS: A total of 440 Chinese pediatric hearing-impaired subjects were recruited from two otology clinics in the Ningbo and Wenzhou cities of Zhejiang Province, China. These subjects underwent clinical, genetic evaluation and molecular analysis of mitochondrial 12S rRNA. Resultant mtDNA variants were evaluated by structural and phylogenetic analysis. RESULTS: The study samples consisted of 227 males and 213 females. The age of all participants ranged from 1 years old to 18 years, with the median age of 9 years. Ninety-eight subjects (58 males and 40 females) had a history of exposure to aminoglycosides, accounting for 22.3% cases of hearing loss in this cohort. Molecular analysis of 12S rRNA gene identified 41 (39 known and 2 novel) variants. The incidences of the known deafness-associated 1555A > G, 1494C > T and 1095T > C mutations were 7.5%, 0.45% and 0.91% in this entire hearing-impaired subjects, respectively, and 21.4%, 2% and 2% among 98 subjects with aminoglycoside ototoxicity, respectively. The structural and phylogenetic evaluations showed that a novel 747A > G variant and known 839A > G, 1027A > G, 1310C > T and 1413T > C variants conferred increased sensitivity to aminoglycosides or nonsyndromic deafness as they were absent in 449 Chinese controls and localized at highly conserved nucleotides of this rRNA. However, other variants were polymorphisms. Of 44 subjects carrying one of definite or putative deafness-related 12S rRNA variants, only one subject carrying the 1413T > C variant harbored the 235DelC/299DelAT mutations in the GJB2 gene, while none of mutations in GJB2 gene was detected in other 43 subjects. CONCLUSIONS: Mutations in mitochondrial 12S rRNA accounted for ~30% cases of aminoglycoside-induced deafness in this cohort. Our data strongly support the idea that the mitochondrial 12S rRNA is the hot spot for mutations associated with aminoglycoside ototoxicity. These data have been providing valuable information and technology to predict which individuals are at risk for ototoxicity, to improve the safety of aminoglycoside antibiotic therapy, and eventually to decrease the incidence of deafness.

[Genetic analysis of family constellation for cochlear implant recipients].

GJB2, SLC26A4 (PDS) and mitochondrial DNA (mtDNA) have been associated with sensorineural hearing loss. In the present study, the clinical, genetic and molecular analysis of 14 cochlear implant recipients and their parents was studied from April 2006 to September 2007. Of the 14 subjects, 35.7% had gene mutations; 28.6% had homozygous GJB2 235delC mutation, whose parents carried heterozygous GJB2 235delC mutation; and 7.1% had mtDNA A1555G mutation, whose mother carried mtDNA A1555G mutation too. There was no SLC26A4 (PDS) mutation. These results strongly suggested that the mutation in GJB2 gene was a major cause of deafness in cochlear implant recipients and the mutation of mtDNA A1555G was another important cause. Genetic test of hot-spots and analysis of family constellation can offer an accurate genetic counseling to deaf family and reduce the incidence of hearing loss.

Mitochondrial ND5 T12338C, tRNA(Cys) T5802C, and tRNA(Thr) G15927A variants may have a modifying role in the phenotypic manifestation of deafness-associated 12S rRNA A1555G mutation in three Han Chinese pedigrees.

We report here on the clinical, genetic, and molecular characterization of three Han Chinese pedigrees with aminoglycoside-induced and nonsyndromic hearing loss. Clinical evaluation revealed the variable phenotype of hearing impairment including severity, age-at-onset, audiometric configuration in these subjects. The penetrance of hearing loss in WZD8, WZD9, and WZD10 pedigrees were 46%, 46%, and 50%, respectively, when aminoglycoside-induced deafness was included. When the effect of aminoglycosides was excluded, the penetrance of hearing loss in these pedigrees were 23%, 31%, and 37.5%, respectively. Mutational analysis of the complete mitochondrial genomes showed the homoplasmic A1555G mutation and distinct sets of mitochondrial DNA variants belonging to haplogroups D4b2b, B5b1, and F2, respectively. Of these, the tRNA(Cys) T5802C, tRNA(Thr) A15924C, and ND5 T12338C variants are of special interest as these variants occur at positions which are highly evolutionarily conserved nucleotides of tRNAs or amino acid of polypeptide. These homoplasmic mtDNA variants were absent among 156 unrelated Chinese controls. The T5802C and G15927A variants disrupted a highly conserved A-U or C-G base-pairing at the anticodon-stem of tRNA(Cys) or tRNA(Thr), while the ND5 T12338C mutation resulted in the replacement of the translation-initiating methionine with a threonine, and also located in two nucleotides adjacent to the 3' end of the tRNA(Leu(CUN)). Thus, mitochondrial dysfunctions, caused by the A1555G mutation, would be worsened by these mtDNA variants. Therefore, these mtDNA mutations may have a potential modifier role in increasing the penetrance and expressivity of the deafness-associated 12S rRNA A1555G mutation in those Chinese pedigrees.