Genetic features of hearing loss associated with ear anomalies: PDS and EYA1 mutation analysis

Mutation analysis of the PDS gene and the EYA1 gene, which are reported to be responsible for hearing loss associated with ear anomalies, was performed in 24 deaf patients with various middle and inner ear anomalies. The present study was done to clarify the spectrum of middle and inner ear malformations covered by these two genes. PDS mutations were found only in patients with enlarged vestibular aqueducts and EYA1 mutations were detected only in patients with ear pits and cervical fistulae, indicating that these two genes are associated with particular forms of middle and inner ear malformation. The genetic approach provides a strong tool for the diagnosis of hearing loss associated with ear anomalies. Received: January 16, 2001 / Accepted: June 12, 2001     

Molecular screening of patients with nonsyndromic hearing loss from Nanjing city of China

Hearing loss is the most frequent sensory disorder involving a multitude of factors,and at least 50% of cases are due to genetic etiology.To further characterize the molecular etiology of hearing loss in the Chinese population,we recruited a total of 135 unrelated patients with nonsyndromic sensorineural hearing loss (NSHL) for mutational screening of GJB2,GJB3,GJB6,SLC26A4,SLC26A5 IVS2-2A>G and mitochondrial 12SrRNA,tRNA Ser(UCN) by PCR amplification and direct DNA sequencing.The carrier frequencies of deafness-causing mutations in these patients were 35.55% in GJB2,3.70% in GJB6,15.56% in SLC26A4 and 8.14% in mitochondrial 12SrRNA,respectively.The results indicate the necessity of genetic screening for mutations of these causative genes in Chinese population with nonsyndromic hearing loss.     

A large cohort study of GJB2 mutations in Japanese hearing loss patients

Tsukada K, Nishio S, Usami S, and the Deafness Gene Study Consortium. A large cohort study of GJB2 mutations in Japanese hearing loss patients. GJB2 is the gene most frequently associated with hereditary hearing loss, and the GJB2 mutation spectrums vary among different ethnic groups. In this study, the mutation spectrum as well as clinical features of patients with GJB2 mutations as found in more than 1000 Japanese hearing loss families are summarized. The present results show that the frequency of GJB2 mutations in the Japanese population with hearing loss is 14.2% overall and 25.2% in patients with congenital hearing loss. c.235delC was the most frequent allele (49.8%), was associated with a more severe phenotype, and was mainly found in patients who were diagnosed by the age of 3. In contrast, the second most frequent was p.V37I (16.5%), which has a milder phenotype and was mainly found in patients diagnosed at a higher age. Additional clinical features in hearing loss patients with GJB2 mutations in this study were the near absence of tinnitus, vestibular dysfunction and inner ear malformations.     

Spectrum and Frequency of SLC26A4 Mutations Among Czech Patients with Early Hearing Loss with and without Enlarged Vestibular Aqueduct (EVA)

Mutations in SLC26A4 cause Pendred syndrome (PS) – hearing loss with goitre – or DFNB4 – non‐syndromic hearing loss (NSHL) with inner ear abnormalities such as Enlarged Vestibular Aqueduct (EVA) or Mondini Dysplasia (MD). We tested 303 unrelated Czech patients with early hearing loss (298 with NSHL and 5 with PS), all GJB2‐negative, for SLC26A4 mutations and evaluated their clinical and radiological phenotype. Among 115 available HRCT/MRI scans we detected three MD (2.6%), three Mondini‐like affections (2.6%), 16 EVA (13 bilateral – 19.2% and 15.6% respectively) and 61 EVA/MD‐negative scans (73.4%). We found mutation(s) in 26 patients (8.6%) and biallelic mutations in eight patients (2.7%) out of 303 tested. In 18 of 26 (69%) patients, no second mutation could be detected even using MLPA. The spectrum of SLC26A4 mutations in Czech patients is broad without any prevalent mutation. We detected 21 different mutations (four novel). The most frequent mutations were p.Val138Phe and p.Leu445Trp (18% and 8.9% of pathogenic alleles respectively). Among 13 patients with bilateral EVA, six patients (50%) carry biallelic mutations. In EVA ‐negative patients no biallelic mutations were found but 4.9% had monoallelic mutations. SLC26A4 mutations are present mostly in patients with EVA/MD and/or progressive HL and those with affected siblings.     

Mitochondrial DNA 4977bp Deletion Mutation in Peripheral Blood Reflects Atrial Remodeling in Patients with Non-Valvular Atrial Fibrillation

Purpose

Recently, mitochondrial DNA 4977bp deletion (mtDNA4977^-mut), a somatic mutation related to oxidative stress, has been shown to be associated with atrial fibrillation (AF). We hypothesized that patient age, as well as electroanatomical characteristics of fibrillating left atrial (LA), vary depending on the presence of mtDNA4977^-mut in peripheral blood among patients with non-valvular AF.

Materials and Methods

Analyzing clinical and electroanatomical characteristics, we investigated the presence of the mtDNA4977^-mut in peripheral blood of 212 patients (51.1±13.2 years old, 83.5% male) undergoing catheter ablation for non-valvular AF, as well as 212 age-matched control subjects.

Results

The overall frequency of peripheral blood mtDNA4977^-mut in patients with AF and controls was not significantly different (24.5% vs. 19.3%, p=0.197). When the AF patient group was stratified according to age, mtDNA4977^-mut was more common (47.4% vs. 20.0%, p=0.019) in AF patients older than 65 years than their age-matched controls. Among AF patients, those with mtDNA4977^-mut were older (58.1±11.9 years old vs. 48.8±11.9 years old, p<0.001). AF patients positive for the mtDNA mutation had greater LA dimension (p=0.014), higher mitral inflow peak velocity (E)/diastolic mitral annular velocity (Em) ratio (p<0.001), as well as lower endocardial voltage (p=0.035), and slower conduction velocity (p=0.048) in the posterior LA than those without the mutation. In multivariate analysis, E/Em ratio was found to be significantly associated with the presence of mtDNA4977^-mut in peripheral blood.

Conclusion

mtDNA4977^-mut, an age-related somatic mutation detected in the peripheral blood, is associated with advanced age and electro-anatomical remodeling of the atrium in non-valvular AF.     

Extremely discrepant mutation spectrum of <Emphasis Type="Italic">SLC26A4</Emphasis> between Chinese patients with isolated Mondini deformity and enlarged vestibular aqueduct

Background

Mutations in SLC26A4 cause Pendred syndrome (hearing loss with goiter) or DFNB4 (non-syndromic hearing loss with inner ear malformation, such as enlarged vestibular aqueduct or Mondini deformity). The relationship between mutations in SLC26A4 and Mondini deformity without enlarged vestibular aqueduct has not been studied in any Chinese deaf population. The purpose of this study was to assess whether mutations in the SLC26A4 gene cause Mondini deformity without an enlarged vestibular aqueduct (isolated Mondini deformity) in a Chinese population.

Methods

In total, 144 patients with sensorineural hearing loss were included and subjected to high-resolution temporal bone CT. Among them, 28 patients with isolated Mondini dysplasia (MD group), 50 patients with enlarged vestibular aqueduct with Mondini dysplasia (EVA with MD group), 50 patients with enlarged vestibular aqueduct without Mondini dysplasia (EVA group), and 16 patients with other types of inner ear malformations (IEM group) were identified. The coding exons of SLC26A4 were analyzed in all subjects.

Results

DNA sequence analysis of SLC26A4 was performed in all 144 patients. In the different groups, the detection rate of the SLC26A4 mutation differed. In the isolated MD group, only one single allelic mutation in SLC26A4 was found in one patient (1/28, 3.6%). In the EVA with MD group, biallelic and monoallelic SLC26A4 mutations were identified in 46 patients (46/50, 92.0%) and three patients (3/50, 6.0%), respectively. Also, in the EVA group, biallelic and monoallelic SLC26A4 mutations were identified in 46 patients (46/50, 92.0%) and three patients (3/50, 6.0%), respectively. These percentages were identical to those in the EVA plus MD group. Only two patients carried monoallelic mutations of the SLC26A4 gene in the IEM group (2/16, 12.5%). There were significant differences in the frequency of SLC26A4 mutation among the groups (P < 0.001). The detection rate of SLC26A4 mutation in the isolated MD group was significantly lower than in the EVA group (with or without MD; P < 0.001), and there was no significant difference in the detection rate of SLC26A4 between the MD group and IEM group (P > 0.5).

Conclusion

Although mutations in the SLC26A4 gene were frequently found in Chinese EVA patients with and without MD, there was no evidence to show a relationship between isolated MD and the SLC26A4 gene in the Chinese population examined. Hearing impairment in patients with isolated MD may be caused by factors other than mutations in the SLC26A4 gene.

     

Autosomal recessive otofaciocervical syndrome type 2 with novel homozygous small insertion in PAX1 gene

Otofaciocervical syndrome (OTFCS) is described as a single gene disorder of both autosomal dominant and autosomal recessive inheritance. The major clinical features of OTFCS include ear malformations (external/middle/inner ear), facial dysmorphism, shoulder girdle abnormalities, vertebral anomalies, and mild intellectual disability. The autosomal recessive form of OTFCS syndrome (OTFCS2) has been recently reported to be caused due to homozygous mutations in PAX1 gene. Here we report a third family of OTFCS2 phenotype wherein whole exome sequencing identified a novel homozygous small insertion in PAX1 as the underlying genetic cause.     

SNaPshot reveals high mutation and carrier frequencies of 15 common hearing loss mutants in a Chinese newborn cohort

Genetic causes account for more than half of congenital hearing loss cases. The most frequent mutations found in non‐syndromic hearing loss patients occur in GJB2 and SLC26A4. Mitochondrial genome mutations are also prevalent. However, the frequency of common hearing loss mutations in the Chinese population has not yet been well estimated. Here, we implemented the SNaPshot genotyping method to investigate the carrier frequency of 15 commonly reported hearing loss mutations in GJB2, SLC26A4 and the mitochondrial genome based on a cohort of 5800 neonates in China. Up to 15.9% (923/5800) of the newborns carry at least one mutant allele. The top three were GJB2‐c.109G>A, GJB2‐c.235delC, and SLC26A4‐c.919A>G, with notably high carrier frequencies of 1/10, 1/53 and 1/62 respectively, and mt‐7444G>A with 1/141 was the most frequent allele in the mitochondrial genome. In this cohort, 0.48% (28/5800) of neonates were genetically diagnosed with hearing loss, from which seven cases failed an OAE test. This is the first epidemiological study of non‐syndromic hearing loss in Chinese newborns indicating a notably high carrier frequency (1 per 6.3 newborns) among these 15 mutant alleles. Our carrier frequency data also aid in effective risk assessment and genetic counseling for hearing loss patients in the Chinese population.     

Prevalence of Mutations in Deafness‐Causing Genes in Cochlear Implanted Patients with Profound Nonsyndromic Sensorineural Hearing Loss in Shandong Province,China

The mutations of GJB2, SLC26A4, and mtDNA12SrRNA are the most common inherited causes of nonsyndromic sensorineural hearing loss (NSHL) in China, yet previous genetic screenings were mainly carried on patients with moderate‐to‐profound impairment. We aimed to detect the mutation frequencies in NSHL population within a more specified range of severity. Patients with profound NSHL who had undergone cochlear implantation in the Shandong Provincial Hospital (Shandong, China) were recruited. The majority (n = 472) were between 0.7 and 6 years old, and the remaining (n = 63) were between 6 and 70 years old. In total, 115 mutation alleles of the three genes were screened with SNP scan assay. Of the patients, 19.44% (104/535) were found to have GJB2 mutations, and the most common allele was c.235delC, followed by c.299_300delAT and c.109G>A. SLC26A4 mutations were detected in 13.46% patients (72/535), and the most common allele was c.919‐2A>G (IVS7‐2A>G), followed by c.1174A>T and c.2168A>G. Seven patients (1.31%) carried mutations in mtDNA12SrRNA, with the alleles of m.1555A>G and m.1494C>T. We found the allele frequency of c.109G>A (GJB2) was relatively lower in the profound NSHL population in comparison to the moderate‐to‐profound ones, and the c.1174A>T (SLC26A4) relatively higher. It suggests those mutations may be connected with the degree of deafness, which needs more observations and analyses to support.     

UBASH3A gene polymorphisms and expression profile in rheumatoid arthritis

Objectives: Recent evidence has demonstrated that UBASH3A play a pivotal role in multiple autoimmune diseases. In this study, we explored the association between UBASH3A gene single-nucleotide polymorphisms (SNPs) and rheumatoid arthritis (RA) in a Chinese Han population. We also comparatively evaluated the UBASH3A expression profile in peripheral blood mononuclear cells (PBMCs) from patients with RA and healthy controls.

Methods: Four UBASH3A polymorphisms (rs1893592, rs11203203, rs2277798, and rs3788013) were studied in 553 patients with RA and 587 controls in a Chinese population. Genotyping was performed using the Fluidigm 192.24 Dynamic Array Integrated Fluidic Circuit (IFC). For gene expression study, UBASH3A mRNA levels of 30 RA patients and 31 healthy individuals were assessed by real-time quantitative polymerase chain reaction (RT-qPCR). Data were analyzed by SPSS 19.0 software.

Results: A significant association between rs1893592 polymorphism and RA was found under all genetic models (all p<.05). We also discovered a significant association between rs3788013 polymorphism and RA in the allele and genotype distributions, as well as the recessive model (all p<.05). Moreover, we found the genotype distribution and allele frequency of rs1893592 were significantly associated with RF phenotype in the RA patients (χ²?=?6.786, p=.034; χ²?=?4.534, p=.033; respectively). We also found the genotype distribution and allele frequency of rs2277798 were significantly associated with anti-CCP phenotype in the RA patients (χ²?=?7.873, p=.020; χ²?=?4.473, p=.034; respectively). However, we did not detect any significant associations between rs11203203 and RA susceptibility and autoantibody profiles (all p>.05). The mRNA expression of UBASH3A was increased in PBMCs of patients with RA when compared to healthy controls (p=.001).

Conclusions: Our observations suggested that the dysregulation of UBASH3A might be associated with the pathogenesis of RA, and UBASH3A gene polymorphisms (rs1893592 and rs3788013) might contribute to RA susceptibility in Chinese Han population.     

Hearing impairment in Estonia: An algorithm to investigate genetic causes in pediatric patients

PurposeThe present study was initiated to establish the etiological causes of early onset hearing loss (HL) among Estonian children between 2000–2009.MethodsThe study group consisted of 233 probands who were first tested with an arrayed primer extension assay, which covers 199 mutations in 7 genes (GJB2, GJB6, GJB3, SLC26A4, SLC26A5 genes, and two mitochondrial genes – 12S rRNA, tRNA^Ser(UCN)). From probands whose etiology of HL remained unknown, DNA analysis of congenital cytomegalovirus (CMV) infection and G-banded karyotype and/or chromosomal microarray analysis (CMA) were performed.ResultsIn 110 (47%) cases, the etiology of HL was genetic and in 5 (2%) congenital CMV infection was diagnosed. We found mutations with clinical significance in GJB2 (100 children, 43%) and in 2 mitochondrial genes (2 patients, 1%). A single mutation in SLC26A4 gene was detected in 5 probands (2.2%) and was considered diagnostic. In 4 probands a heterozygous IVS2-2A>G change in the SLC26A5 gene was found. We did not find any instances of homozygosity for this splice variant in the probands. CMA identified in 4 probands chromosomal regions with the loss of one allele. In 2 of them we were able to conclude that the found abnormalities are definitely pathogenic (12q13.3-q14.2 and 17q22-23.2 microdeletion), but the pathogenity of 2 other findings (3p26.2 and 1p33 microdeletion) remained unknown.ConclusionThis practical diagnostic algorithm confirmed the etiology of early onset HL for 115 Estonian patients (49%). This algorithm may be generalized to other populations for clinical application.     

Phenotype and genotype in females with POU3F4 mutations

X‐linked deafness is a rare cause of hereditary isolated hearing impairment estimated as at least 1% or 2% of the non‐syndromic hearing loss. To date, four loci for DFN have been identified and only one gene, POU3F4 responsible for DFN3, has been cloned. In males, DFN3 is characterized by a progressive deafness associated with perilymphatic gusher at stapes surgery and with a characteristic inner ear malformation. The phenotype of eight independent females carrying POU3F4 anomalies is defined, and a late‐onset hearing loss is found in three patients. Only one has an inner ear malformation. No genotype/phenotype correlation is identified.     

Chondrocyte‐specific Smad4 gene conditional knockout results in hearing loss and inner ear malformation in mice

Smad4 is the central intracellular mediator of transforming growth factor‐β (TGF‐β) signaling, which plays crucial roles in tissue regeneration, cell differentiation, embryonic development, and regulation of the immune system. Conventional Smad4 gene knockout results in embryonic lethality, precluding its use in studies of the role of Smad4 in inner ear development. We used chondrocyte‐specific Smad4 knockout mice (Smad4^Co/Co) to investigate the function of Smad4 in inner ear development. Smad4^Co/Co mice were characterized by a smaller cochlear volume, bone malformation, and abnormalities of the osseous spiral lamina and basilar membrane. The development of the hair cells was also abnormal, as evidenced by the disorganized stereocilia and reduced density of the neuronal processes beneath the hair cells. Auditory function tests revealed the homozygous Smad4^Co/Co mice suffered from severe sensorineural hearing loss. Our results suggest that Smad4 is required for inner ear development and normal auditory function in mammals. Developmental Dynamics, 2009. © 2009 Wiley‐Liss, Inc.     

Association between the interleukin-1B polymorphism at rs16944 T>C and diabetic retinopathy

Diabetic retinopathy (DR) is a common microvascular complication of diabetes and the leading cause of blindness at working age. DR is considered to be a chronic low-grade inflammatory subclinical disease, and its pathogenesis is related to genetic and environmental factors. Interleukin (IL)-1 is an important inflammatory cytokine. An association between DR and the rs16944 (IL-1B-511) T>C gene polymorphism has not been reported. The aim of this study was to investigate the association between the rs16944 T>C gene polymorphism and DR in the Han population in southwest China. Participants in this study were 272 patients with DR, 274 patients with type 2 diabetes mellitus (T2DM), and 335 healthy controls (NC). The polymerase chain reaction-restriction fragment length polymorphism method was used to detect the rs16944 T>C genotype of participants. The distribution frequencies of the rs16944 T>C genotype and allele were significantly different among the three groups (p < .05). The distribution frequency of TT, CT, CC genotype (χ² = 9.893, p = .007; χ² = 6.567, p = .037) and each allele (χ² = 5.585, p = .018; χ² = 9.187, p = .002) in the DR group was significantly different from the NC and T2DM groups, respectively. Logistic regression analysis showed that the TT + CT genotype was a risk factor for DR, with an odds ratio of 1.731 (95% confidence interval 1.140–2.627, p = .01). The rs16944 T>C gene polymorphism may be associated with DR, and the TT+CT genotype may increase the risk of DR.     

Three novel GJB2 (connexin 26) variants associated with autosomal dominant syndromic and nonsyndromic hearing loss

Connexin 26 (Cx26), encoded by the GJB2 gene, is a key protein involved in the formation of gap junctions in epithelial organs including the inner ear and palmoplantar epidermis. Pathogenic variants in GJB2 are responsible for approximately 50% of inherited sensorineural deafness. The majority of these variants are associated with autosomal recessive inheritance; however, rare reports of dominantly co‐segregating variants have been published. Since we began offering GJB2 testing in 2003, only about 2% of detected GJB2 variants from our laboratory have been classified as dominant. Here we report three novel dominant GJB2 variants (p.Thr55Ala, p.Gln57_Pro58delinsHisSer, and p.Trp44Gly); two associated with syndromic sensorineural hearing loss and one with nonsyndromic hearing loss. In the kindred with the p.Thr55Ala variant, the proband and his father present with only leukonychia as a cutaneous finding of their syndromic hearing loss. This phenotype has been previously documented in conjunction with palmoplantar hyperkeratosis, but isolated leukonychia is a novel finding likely associated with the unique threonine to alanine change at codon 55 (other variants at this codon have been reported in cases of nonsyndromic hearing loss). This report contributes to the short list of GJB2 variants associated with autosomal dominant hearing loss, highlights the variability of skin and nail findings associated with such cases, and illustrates the occurrence of both syndromic and nonsyndromic presentations with changes in the same gene.     

Genetic Polymorphism (rs329498) in the Pellino-1 Gene as Possible Predisposal Factor for Systemic Lupus Erythematosus in a Chinese Population

Objective: The purpose of this study was to analyze the association of two single nucleotide polymorphisms (SNPs) in Peli-1 gene with systemic lupus erythematosus (SLE) in a Chinese population.

Methods: We conducted a case–control study and a total of 738 SLE patients and 827 healthy controls were finally recruited. Peli-1 rs329498 and rs10496105 polymorphisms were specified from genomic DNA using TaqMan genotyping assay on Fluidigm 192.24 system.

Results: Allele contrast showed the minor allele C was associated with decreased risk for SLE when compared with the A allele (OR = 0.851, 95% CI = 0.737–0.983, p = 0.028). Significant difference was observed in genotype distribution of rs329498 polymorphism between lupus nephritis (LN) patients and non-LN patients (χ² = 8.18, p = 0.017). Furthermore, we also found a decreased frequency of the minor allele C in LN patients (29.2%) than in non-LN patients (37.7%) (χ² = 8.67, p = 0.003). Moreover, a significant difference was also detected under a dominant model with regard to the distribution of genotype frequencies between LN patients and non-LN patients (CC + AC vs. AA: OR = 0.632, 95% CI = 0.451–0.884, p = 0.007). Clinical features analysis showed a significant difference in the distribution of genotypic frequencies between patients with malar rash and patients without this feature (χ² = 6.63, p = 0.036). Unfortunately, we failed to find any significant results between Peli-1 gene rs10496105 and SLE susceptibility.

Conclusions: Our observations suggested that Peli-1 gene polymorphism rs329498 might contribute to SLE susceptibility in Chinese Han Population. Likewise, the rs329498 SNP was also associated with the clinical features LN and malar rash in SLE patients.     

The role of Zic genes in inner ear development in the mouse: Exploring mutant mouse phenotypes

Background: Murine Zic genes (Zic1–5) are expressed in the dorsal hindbrain and in periotic mesenchyme (POM) adjacent to the developing inner ear. Zic genes are involved in developmental signaling pathways in many organ systems, including the ear, although their exact roles haven't been fully elucidated. This report examines the role of Zic1, Zic2, and Zic4 during inner ear development in mouse mutants in which these Zic genes are affected. Results: Zic1/Zic4 double mutants don't exhibit any apparent defects in inner ear morphology. By contrast, inner ears from Zic2^kd/kd and Zic2^Ku/Ku mutants have severe but variable morphological defects in endolymphatic duct/sac and semicircular canal formation and in cochlear extension in the inner ear. Analysis of otocyst patterning in the Zic2^Ku/Ku mutants by in situ hybridization showed changes in the expression patterns of Gbx2 and Pax2. Conclusions: The experiments provide the first genetic evidence that the Zic genes are required for morphogenesis of the inner ear. Zic2 loss‐of‐function doesn't prevent initial otocyst patterning but leads to molecular abnormalities concomitant with morphogenesis of the endolymphatic duct. Functional hearing deficits often accompany inner ear dysmorphologies, making Zic2 a novel candidate gene for ongoing efforts to identify the genetic basis of human hearing loss. Developmental Dynamics 243:1487–1498, 2014. © 2014 Wiley Periodicals, Inc.     

Two SOX11 variants cause Coffin–Siris syndrome with a new feature of sensorineural hearing loss

Coffin-Siris syndrome (CSS, OMIM#135900) is a rare congenital disorder associated with neurodevelopmental and dysmorphic features. The primary cause of CSS is pathogenic variants in any of 9 BAF chromatin-remodeling complex encoding genes or the genes SOX11 and PHF6. Herein, we performed whole-exome sequencing (WES) and a series of analyses of growth-related, auditory, and radiological findings in two probands with syndromic sensorineural hearing loss and inner ear malformations who exhibited distinctive facial features, intellectual disability, growth retardation, and fifth finger malformation. Two de novo variants in the SOX11 gene (c.148A>C:p.Lys50Asn; c.811_814del:p.Asn271Serfs*10) were detected in these probands and were identified as pathogenic variants as per ACMG guidelines. These probands were diagnosed as having CSS based upon clinical and genetic findings. This is the first report of CSS caused by variants in SOX11 gene in Chinese individuals. Deleterious SOX11 variants can result in sensorineural hearing loss with inner ear malformation, potentially extending the array of phenotypes associated with these pathogenic variants. We suggest that both genetic and clinical findings be considered when diagnosing syndromic hearing loss.     

Development of a new multiplex quantitative real‐time PCR assay for the detection of the mtDNA4977 deletion in coronary artery disease patients: A link with telomere shortening

Mitochondrial DNA (mtDNA) and telomere shortening have been proposed as important contributors to vascular disease and atherogenesis. The role of mitochondrial and telomere alterations has been examined frequently, but usually separately. Recently, an integrated model in which DNA damage and metabolic pathways intersect in age‐associated cardiovascular disease has been proposed. In this study we developed a fast and reliable real‐time PCR‐based procedure to investigate relative quantification of the 4,977 bp mitochondrial DNA deletion (also indicated as “mtDNA⁴⁹⁷⁷ deletion”), employing TaqMan probes with a multiplex approach. As a validation of the assay, a nested PCR coamplification was performed. Telomere shortening was evaluated by a real‐time monochrome multiplex PCR technique employing a SybrGreen‐based analysis. The study of mtDNA⁴⁹⁷⁷ deletion and telomere shortening was carried out in atrial biopsies from 11 patients undergoing coronary artery (n = 5) and valve surgery (n = 6). The relative quantifications showed that the amount of mtDNA⁴⁹⁷⁷ deletion was greater in tissue of patients with coronary artery disease (CAD) (P = 0.01) and that telomere length (expressed as telomere length relative to a single copy reference gene) was significantly shorter in tissue of CAD patients, compared to patients without CAD (P = 0.03). Moreover, most conventional risk factors were significantly more frequent in CAD patients, smoking and dyslipidemia having the strongest association with the degree of mtDNA⁴⁹⁷⁷deletion and a significant correlation with telomere attrition (P = 0.02 and P = 0.006, respectively). In conclusion, the present study suggests that mtDNA⁴⁹⁷⁷ deletion and telomere shortening may represent additional and synergic major risk factors for the pathogenesis of CAD and its complications. Environ. Mol. Mutagen. 54:299–307, 2013. © 2013 Wiley Periodicals, Inc.     

Upper limb deficiencies and associated malformations: A population-based study

As part of an ongoing analysis of limb deficiencies occurring among 1,213,913 consecutive livebirths in British Columbia during the years 1952–1984, all cases with deficiencies of the upper limbs were analysed with a view to identifying associated patterns of anomalies. This analysis resulted in seven subgroups. For each subgroup, incidence figures for cases with and without additional anomalies were calculated separately. The proportion of cases with additional anomalies varied markedly by subgroup. For example, 89% of cases with longitudinal defects of the radius had additional malformations, while only 28% of cases with transverse defects of the radius had other organ anomalies (χ² = 40.55; P < 0.001, one degree of freedom). A preponderance of males was found among the cases with associated defects, particularly in the group with longitudinal defects of the radius (28 males, 14 females; χ² = 14.10; P < 0.001). Clustering of specific patterns of associated malformations is described within subgroups. © 1992 Wiley-Liss, Inc.