TSHRV656F Activating Variant of the Thyroid Stimulating Hormone Receptor Gene in Neonatal Onset Hyperthyroidism: A Case Review

An activating variant of the thyroid stimulating hormone receptor (TSHR) gene is one of the rare causes of neonatal hyperthyroidism. This disorder may occur as a result of an autosomal dominant inheritance or sporadically through de novo variation. Here we present a case of neonatal onset congenital non-autoimmune hyperthyroidism (NAH) with a sporadic germline activating TSHRV656F variant. A female infant with tachycardia, who was transferred due to hyperthyroidism in the first week of life, displayed no other symptoms or signs. The patient's mother did not have Graves' disease, and TSHR stimulating antibodies were not present in the mother or baby. Imaging showed thyroid gland hyperplasia and left ventricular hypertrophy, the patient was subsequently put on methimazole treatment. After six months undergoing treatment, a heterozygous p.Val656Phe (V656F) (c.1966G>T) variant was detected on exon 10 of the TSHR gene. The variant was not identified in the mother and father, so the case was assumed to be sporadic. In conclusion, although the literature describes V656F variant as a somatic variant in children and adults with toxic thyroid nodule(s) that results in the structural activation of the TSH receptor, no previous cases of neonatal hyperthyroidism due to TSHRV656F variant have been reported. This study is the first case review that highlights the relationship between TSHRV656F variant and neonatal onset NAH.

Phylogenetic and phylogeographic analysis of Myrmeleotettix maculatus (Orthoptera: Acrididae: Gomphocerinae) species group in Anatolia.

Six Anatolian and one European populations of the Myrmeleotettix maculatus species group, which contains M. maculatus and M. ethicus species, have been studied by using molecular genetics methods with mitochondrial COI gene. Myrmeleotettix ethicus is an Anatolian endemic species with local distribution whereas M. maculatus is distributed in western Palearctic. The phylogenetic analysis (ML and BI analyses) of the M. maculatus species group in Anatolia reveals that it consistently recovered two well-supported main clades and four different lineages. Molecular time estimates suggest that the diversification of the M. maculatus species group took place between the Late Tortonian (around 8-9 My) and the Middle of Pliocene-Pleistocene (around 4.3 My-present) periods and the current distribution of the genetic diversity has been affected by the uplifting of the Central Anatolian plateau, the termination of the Messinian salinity crisis, and the Quaternary climatic changes.

Sensitive Sequencing Analysis Suggests Thyrotropin Receptor and Guanine Nucleotide-Binding Protein G Subunit Alpha as Sole Driver Mutations in Hot Thyroid Nodules.

Background: Constitutively activating mutations in the thyrotropin receptor (TSHR) and the guanine nucleotide-binding protein G subunit alpha (GNAS) are the primary cause of hot thyroid nodules (HTNs). The reported prevalence of TSHR and GNAS mutations in HTNs varies. Previous studies show TSHR mutations in 8-82% of HTNs and GNAS mutations in 8-75% of HTNs. With sensitive and comprehensive targeted next-generation sequencing (tNGS), we re-evaluated the prevalence of TSHR and GNAS mutations in HTNs. Methods: Samples from three previous studies found to be TSHR and GNAS mutation negative were selected and re-evaluated using high-resolution melting (HRM) PCR. Remaining mutation negative samples were further reanalyzed by tNGS with a sequencing depth between 3000 × and 10,000 × . Our tNGS panel covered the entire TSHR coding sequence along with mutation hot spots in GNAS. Sequencing reads were aligned to reference and variants were called using Torrent Suite software v5.8. Results: In total, 154 of 182 previously mutation negative HTNs were positive for TSHR or GNAS mutations, resulting in an 85% prevalence of TSHR and GNAS mutations in HTNs, 79% and 6%, respectively. In a subset of 25 HTNs with multiple samples per nodule, and analyzed by tNGS at high sequencing depth, TSHR mutations were detected in 23 (92%) HTNs and 1 GNAS mutation was detected in 1 (4%) HTN, 96% mutation positive HTNs in this subset. Conclusions: Owing to the higher sensitivity of tNGS as compared with denaturing gradient gel electrophoresis and HRM-PCR, TSHR or GNAS mutations could be detected in 85% of HTNs. The detection of TSHR and GNAS mutations occurred in 96% of HTNs in a sample set with multiple samples per nodule analyzed by tNGS. Taken together with the fact that no other driver mutations could be identified by whole exome sequencing, our study strongly supports the hypothesis that TSHR and GNAS mutations are the main somatic mutations leading to HTNs.

Crimean-Congo hemorrhagic fever virus in tortoises and Hyalomma aegyptium ticks in East Thrace, Turkey: potential of a cryptic transmission cycle.

BACKGROUND: Recent reports have demonstrated the presence of Crimean-Congo hemorrhagic fever virus (CCHFV) genomic material in Hyalomma aegyptium ticks feeding primarily on tortoises belonging to the genus Testudo. This raises the question if these ticks and their hosts play a role in the natural transmission dynamics of CCHFV. However, the studies are limited, and assessing the relevance of H. aegyptium in perpetuating the virus in nature, and a potential spillover to humans remains unknown. This study aimed to detect CCHFV in H. aegyptium ticks and their tortoise hosts in the East Thrace region of Turkey, where H. aegyptium is the most common human-biting tick and where a high density of tortoises of the genus Testudo can be found. METHODS: During the study period, 21 blood samples from different tortoises (2 T. hermanni and 19 T. graeca), 106 tick pools (containing 448 males, 152 females, 93 nymphs and 60 larvae) collected from 65 tortoises (5 T. hermanni and 60 T. graeca), 38 adult unfed questing ticks (25 males and 13 females, screened individually) and 14 pools (containing 8 nymphs and 266 larvae) of immature unfed questing ticks collected from the ground were screened for CCHFV genome by nested PCR and partial genomes sequenced. RESULTS: As a result of the screening of these 179 samples, 17 (9.5%) were detected as positive as follows: 2 of 21 blood samples (9.52%), 13 (containing 18 nymphs in 3 pools, and 52 males and 8 females in 10 pools) of 106 tick pools from tortoises (12.26%), and 2 of 38 adult questing ticks (5.26%). No positive result was determined in 14 pools of immature questing ticks. CONCLUSIONS: Previous studies have shown that reptiles can participate in the transmission of arthropod-borne viruses, but they may contribute to different aspects of the disease ecology and evolution of tick-borne viral pathogens. Our results indicate the presence of CCHFV in questing and feeding H. aegyptium ticks as well as tortoise hosts. This may indicate that CCHFV circulates in a cryptic transmission cycle in addition to the primary transmission cycle that could play a role in the natural dynamic of the virus and the transmission to humans.

Melatonin reverses the oxidative stress and mitochondrial dysfunction caused by LETM1 silencing.

LETM1 is a mitochondrial inner-membrane protein, which is encoded by a gene present in a locus of 4p, which, in turn, is deleted in the Wolf-Hirschhorn Syndrome, and is assumed to be related to its pathogenesis. The cellular damage caused by the deletion is presumably related to oxidative stress. Melatonin has many beneficial roles in protecting mitochondria by scavenging reactive oxygen species, maintaining membrane potential, and improving functions. The aim of this study was to investigate the effects of melatonin administration to LETM1-silenced mouse embryonic fibroblast cells as a cellular model for LETM1 deficiency. We transfected mouse embryonic fibroblast cells with a pair of siRNA against LETM1 and monitored the oxidative stress and mitochondrial functions with or without melatonin addition. MnSOD expression and aconitase activity decreased and oxidized protein levels increased in LETM1-silenced cells. LETM1 suppression did not alter the expression of OXPHOS complexes, but the oxygen consumption rates decreased significantly; however, this change was not related to complex I but instead involved complex IV and complex II. Melatonin supplementation effectively normalized the parameters studied, including the oxygen consumption rate. Our findings identified a novel effect of LETM1 deficiency on cellular respiration via complex II as well as a potential beneficial role of melatonin treatment. On the other hand, these effects may be specific to the cell line used and need to be verified in other cell lines.

BRAF V600E mutation in papillary thyroid cancer is correlated with adverse clinicopathological features but not with iodine exposure.

INTRODUCTION: BRAFV600E activating mutation is the most frequent genetic abnormality in the pathogenesis of papillary thyroid carcinoma. We aimed to evaluate the association between BRAFV600E mutation and well-established prognostic clinicopathological characteristics as well as iodine exposure. MATERIAL AND METHODS: From 2000 to 2012, the data of PTC patients admitted to Dr. Lutfi Kirdar Kartal Education and Research Hospital in Turkey were reviewed retrospectively. Clinicopathological parameters were collected. BRAFV600E mutation was analysed by DNA sequencing method in tumour specimens. We hypothesised thatBRAFV600E mutation prevalence is positively correlated with prolonged iodine exposure and expected to be higher in the second half of the recruitment period due to the increment in time spent from the iodisation process of the table salt in our country. Thus, iodine exposure was categorised as short-term (2000-2006) and long-term (2006-2012). RESULTS: A total of 197 patients were accrued. The study population predominantly consisted of conventional variant. A statistically significant relationship was observed betweenBRAFV600E mutation presence and age (p = 0.03), conventional variant PTC (p = 0.00002), T4 stage (p = 0.002), vascular invasion (p = 0.036), thyroid capsule invasion (p < 0.00001), extrathyroidal tissue invasion (p < 0.00001), and lymph node metastasis (p < 0.00001). When categorised as long-term and short-term, iodine exposure was not statistically significantly related withBRAFV600E mutation; however, there were far more PTC cases in the long-term group (86.3% vs. 13.7%). CONCLUSION: We revealed that BRAFV600E mutation is associated with adverse clinicopathological parameters. There appeared to be no relation between long-term iodine exposure and BRAFV600E.

The Mitochondrial DNA Control Region Might Have Useful Diagnostic and Prognostic Biomarkers for Thyroid Tumors.

The literature suggests that mitochondrial DNA (mtDNA) defects are associated with a large number of diseases including cancers. The role of mtDNA variations in thyroid cancer is a highly controversial topic. Therefore, we investigated the role of mt-DNA control region (CR) variations in thyroid tumor progression and the influence of mtDNA haplogroups on susceptibility to thyroid tumors. For this purpose, in total, 108 hot thyroid nodules (HTNs), 95 cold thyroid nodules (CTNs), 48 papillary thyroid carcinoma (PTC) samples with their surrounding tissues and 104 healthy control subjects' blood samples were screened for all mtDNA CR variations using Sanger sequencing. We found that MtDNA haplogroup U was significantly associated with susceptibility to benign thyroid entities. In addition, eight single nucleotide polymorphisms (SNPs) (T146C, G185A, C194T, C295T, G16129A, T16304C, A16343G and T16362C) in the mtDNA CR were associated with the occurrence of benign and malign thyroid nodules in the Turkish population. As compared with samples taken from a healthy Turkish population and HTNs, the frequency of C7 repeats in D310 polycytosine sequence was found to be higher in CTNs and the PTC samples. In addition, the frequency of somatic mutations in mtMSI regions including T16189C and D514 CA dinucleotide repeats were found to be higher in PTC samples than benign thyroid nodules. Conversely, the frequency of somatic mutations in D310 was found to be higher in HTNs than CTNs and PTCs. In conclusion, mtDNA D310 instability does not play a role in the tumorigenesis of PTC but the results indicate that it might be used as a diagnostic clonal expansion biomarker for premalignant thyroid tumor cells. In addition, D514 CA instability might be considered as a prognostic biomarker for benign to malign transformation in thyroid tumors.

Evaluation of the Effect of Daptomycin, a Glycopeptide Agent, on Intact Intervertebral Disc Tissue.

AIM: To evaluate the effects of pre- and intra-operatively administered daptomycin (DAP) on the intact human primary intervertebral disc tissue cells. MATERIAL AND METHODS: Primary cell cultures were established using tissues obtained through decompressive laminectomy, traumatic intervertebral disc herniation excision, and posterior transpedicular stabilization. Non-drug-administered samples were used as a control group. The samples treated with DAP formed the study group. Molecular assays for proliferation and gene expression were performed. The obtained data were evaluated statistically, and results with a value of p < 0.05 were accepted as significant. RESULTS: While no reduction was observed in the proliferation, the gene expression of intact intervertebral disc tissue cells was time-dependently decreased compared to the control group, and these results were reported to be statistically significant. CONCLUSION: This study observed the effect that a pharmaceutical preparation, which was used on intervertebral disc tissue before and after the operation, had on normal, healthy, and intact tissue. It concludes that alterations in the expression of genes involved in the anabolic and/or catabolic process, even in adjacent healthy tissue, may slow down the healing process of the damaged tissue or cause undesired cell differentiation.

Recurrent EZH1 mutations are a second hit in autonomous thyroid adenomas.

Autonomous thyroid adenomas (ATAs) are a frequent cause of hyperthyroidism. Mutations in the genes encoding the TSH receptor (TSHR) or the Gs protein α subunit (GNAS) are found in approximately 70% of ATAs. The involvement of other genes and the pathogenesis of the remaining cases are presently unknown. Here, we performed whole-exome sequencing in 19 ATAs that were paired with normal DNA samples and identified a recurrent hot-spot mutation (c.1712A>G; p.Gln571Arg) in the enhancer of zeste homolog 1 (EZH1) gene, which codes for a catalytic subunit of the polycomb complex. Targeted screening in an independent cohort confirmed that this mutation occurs with high frequency (27%) in ATAs. EZH1 mutations were strongly associated with known (TSHR, GNAS) or presumed (adenylate cyclase 9 [ADCY9]) alterations in cAMP pathway genes. Furthermore, functional studies revealed that the p.Gln571Arg EZH1 mutation caused increased histone H3 trimethylation and increased proliferation of thyroid cells. In summary, this study revealed that a hot-spot mutation in EZH1 is the second most frequent genetic alteration in ATAs. The association between EZH1 and TSHR mutations suggests a 2-hit model for the pathogenesis of these tumors, whereby constitutive activation of the cAMP pathway and EZH1 mutations cooperate to induce the hyperproliferation of thyroid cells.

How different methodologies of harvesting and analysing the samples affect the test results in determining joint mediators.

Purpose. This study has researched the affect of different methodologies of harvesting and analysing the samples in determining the mediators emerging after the rat articular cartilage injury. Materials and Methods. One hundred and forty-four male wistar rats were divided into 2 groups. Synovial fluid samples were taken from all of the rats. We entered into the right knees of the rats in group I (n = 36) under anaesthesia and took cartilage tissue samples from their distal femur. Samples were taken as reference values for enzyme linked immunosorbent assay (ELISA) and histopathological evaluations. We entered into the right knees of the rats in group II (n = 108) and formed complete layer of cartilage injury in their medial femoral condyles. At the end of the 15th day, the rats were sacrificed after taking synovial fluid samples from their right knees creating defect in the rats in group II. The molecular markers in the synovial fluid and cartilage tissue samples which were taken from the experimental and control groups (MMP-9, MMP-13, TIMP-1, TNF- α , and NO) were analysed by direct or indirect methodologies. SPSS 18.0 Package program was used in the statistical evaluation. Students t-test where the measurement variables between the experimental and control groups were compared was applied. Receiver Operating Characteristics (ROC) curves were used in the determination of the diagnostic sufficiency from the tissue. Results. No difference was found between TIMP-1 (P = 0.67) and MMP-9 (P = 0.28) levels in synovial fluid and cartilage tissue. From the molecular markers, when MMP-9, MMP-13, NO, TIMP-1, TNF- α ', the area under ROC curve, and P values were examined, MMP-13 (P < 0.0001, 95% CI: 0.70-0.85), NO (P < 0.0001, 95% CI: 0.72-0.86), and TNF- α (P < 0.0001, 95% CI: 0.91-0.98) results were found to be statistically significant. Inferences. The indirect ELISA protocol which we apply for the cartilage tissue as an alternative to synovial lavage fluid is a reliable method which can be used in the determination of articular cartilage injury markers.

Synergistic Effect of TGF-ß1 And BMP-7 on Chondrogenesis and Extracellular Matrix Synthesis: An In Vitro Study.

INTRODUCTION: The purpose of the present study seeks to determine the signal timing of BMP-7 and TGF-ß1 from a novel chitosan based hydrogel system that may affect chondrocyte proliferation resulting in the presence of a synergism seen conspicuously in consecutive controlled delivery. METHODS: Four groups of cultured chondrocytes were seeded on a novel designed chitosan based hydrogel. The hydrogel was left empty (control) in one group and loaded with BMP-7, TGF-ß1 and their combination in the other groups, respectively. Hydrogel structure was analyzed with scanning electron microscope. The release kinetics of Growth Factors (GFs) was determined with ELISA. Chondrocyte viability and toxicity after being tested with MTS and collagen type II synthesis, were quantified with western blotting. Canonical regression analysis was used for measuring statistical evaluation. RESULTS: Chitosan based hydrogel allowed controlled release of GFs in different time intervals for BMP-7 and TGF-ß1. Double peak concentration gradient was found to be present in the group loaded with both GFs. In this group, substantially higher chondrocyte growth and collagen synthesis were also detected. CONCLUSIONS: We concluded that, chitosan based hydrogel systems may be adjusted to release GFs consecutively during biodegradation at the layers of surface, which may increase the cell number and enhance collagen type II synthesis.

Association between C677T and A1298C MTHFR gene polymorphism and nonsyndromic orofacial clefts in the Turkish population: a case-parent study.

Two common MTHFR gene polymorphisms (C677T and A1298C) have been implicated in the etiology of nonsyndromic cleft lip/palate (nsCL/P). To investigate the genotype association among nsCL/P in the Turkish population, 56 case-parent trios were recruited into the study. Genotype frequencies were compared to two groups of controls from the same population. A total of 46 case-parent trios were included in transmission disequilibrium test (TDT) analysis. The mothers of the study group had a higher frequency of 677TT genotype, with a three-fold increased risk of having nsCL/P offspring (odds ratio [OR]: 3.14, p=0.03). The combined 677CT/1298AC genotype was also common among these mothers (28%), but it did not reach statistical significance (OR: 2.27, p=0.07). TDT analysis for (C677T) T allele transmission did not reveal a significant association. In conclusion, mothers carrying 677TT genotype or with 677CT/1298AC combined genotype have increased risk of having nsCL/P offspring; therefore, higher periconceptional folic acid supplementation should be advised for decreasing the recurrence risk.

A novel mutation in the MC2R gene causing familial glucocorticoid deficiency type 1.

Familial glucocorticoid deficiency (FGD) or hereditary unresponsiveness to adrenocorticotropin (ACTH) is an autosomal recessive disorder characterized by isolated glucocorticoid deficiency associated with normal mineralocorticoid secretion. Mutations in genes encoding either ACTH receptor or melanocortin 2 receptor accessory protein are responsible for the disease in about 50% of cases, named FGD type 1 and type 2, respectively. Patients may present with hyperpigmentation, recurrent infections, failure to thrive, hypoglycemic seizures, and coma in infancy or early childhood. Here we report the case of a 17-day-old newborn diagnosed with FGD type 1 who presented with hyperbilirubinemia and hyperpigmentation, a sign which was erroneously assumed to be due to prolonged phototherapy by the referring physician. Hormone analysis showed low cortisol and high ACTH levels with normal serum electrolytes and renin-aldosterone axis. Genetic analysis revealed a novel homozygous melanocortin 2 receptor mutation p.Leu225Arg in the patient. The healthy parents were heterozygous for the mutation.

Shared sporadic and somatic thyrotropin receptor mutations display more active in vitro activities than familial thyrotropin receptor mutations.

BACKGROUND: Germline thyrotropin receptor (TSHR) mutations are associated with sporadic congenital nonautoimmune hyperthyroidism and familial nonautoimmune hyperthyroidism. Somatic TSHR mutations are associated with toxic thyroid nodules (TTNs). The objective of the study was to define a relation of the clinical appearance and the in vitro activity (IVA) of the TSHR mutations described by several authors for these thyroid disorders. METHODS: We analyzed the IVAs published as linear regression analysis (LRA) of the constitutive activity as a function of the TSHR expression and the basal cyclic adenosine monophosphate (cAMP) values to determine differences between exclusively somatic, exclusively familial, and shared sporadic and somatic TSHR-mutations. Further, we investigated correlations of the LRAs/basal cAMP values with clinical activity characteristics (CACs) of TTNs, such as largest diameter of the TTN and the age of the patient at thyroid surgery. RESULTS: Shared sporadic and somatic mutations showed higher median LRA (14.5) and higher median basal cAMP values (fivefold) than exclusively familial mutations (6.1, p = 0.0002; 2.9-fold, p < 0.0001, respectively). Moreover, mutations shared between sporadic congenital nonautoimmune hyperthyroidism and toxic thyroid nodules (TTNs) showed higher median LRA/basal cAMP values (p < 0.0001) than exclusively somatic mutations in TTNs (5.1; 3.89-fold, respectively). Exclusively somatic mutations and exclusively familial mutations showed no significant difference in their median LRA values (p = 0.786) but a significant difference for basal cAMP values (p = 0.0006). The two examined CACs showed no correlation with the IVA characterized by LRA/basal cAMP values or with the presence or absence of a TSHR-mutation. CONCLUSIONS: This systematic analysis of published constitutively activating TSHR-mutations, their CACs, and their IVA provides evidence for higher IVA of shared sporadic and somatic TSHR mutations as compared with familial TSHR mutations. CACs of somatic TSHR mutations in TTNs did not have a clear association with the IVA as characterized by LRA or basal cAMP values.

Genetics and phenomics of inherited and sporadic non-autoimmune hyperthyroidism.

TSH receptor (TSHR) germline mutations occur as activating mutations in familial non-autoimmune hyperthyroidism (FNAH) or sporadic non-autoimmune hyperthyroidism (SNAH). Up to date 17 constitutively activating TSHR mutations have been reported in 24 families with FNAH. The diagnosis of FNAH should be considered in cases with a positive family history, early onset of hyperthyroidism, goiter, absence of clinical stigmata of autoimmunity and recurrent hyperthyroidism. Moreover, 14 subjects with sporadic non-autoimmune hyperthyroidism and 10 different TSH receptor germline mutations have been reported. The main characteristic of SNAH is a negative family history. Additional consequences of prolonged neonatal hyperthyroidism (mental retardation, speech disturbances and craniosynostosis) have often been reported in SNAH. No genotype-phenotype relationship has been reported in patients with germline TSHR mutations. There is no association of in vitro activities determined by linear regression analysis (LRA) and several clinical indicators of hyperthyroidism activity for SNAH. However, the comparison of the LRA values of sporadic TSHR mutations with LRA values of familial TSHR mutations does show a significantly higher median LRA value for sporadic as compared to familial autosomal dominant hyperthyroidism. This finding is in line with the clinical impression of a more active clinical course in patients with SNAH. However, additional genetic, constitutional or environmental factors are most likely responsible for the phenotypic variations of the disease and the lack of correlation between in vitro activities of the TSHR mutations and the severity of hyperthyroidism.

Cases of borderline in vitro constitutive thyrotropin receptor activity: how to decide whether a thyrotropin receptor mutation is constitutively active or not?

BACKGROUND: Previous in vitro data for several constitutively activating thyrotropin receptor (TSHR) mutations reported divergent results for the constitutive activity of the same mutations. Moreover, several case reports have highlighted the difficulties in determining whether a TSHR mutation is constitutively active or not. Retrospectively, this has repeatedly been the case for mutants with only a slight increase of basal cAMP activity. We re-examined 10 previously described TSHR germline mutations with minor increases of basal cAMP activity and analyzed the influences of the cell line and vector system on the basal receptor activity. METHODS: TSHR mutations were characterized by determination of cell surface expression, cAMP accumulation, and linear regression analysis of constitutive activity. RESULTS: Re-examination of the previously described constitutively active TSHR germline mutations did not show constitutive activity for R310C and N670S as tested in COS-7 cells and confirmed constitutive activity for the other eight mutations. However, mutant N670S showed a slight but significant increase of basal activity measured by linear regression analysis when analyzed in HEK(GT) cells transiently transfected with pcDNA but not with the pSVL vector. This was not the case for R310C. CONCLUSIONS: Our findings indicate that current methods to precisely classify mutants with only a slight increase of the basal activity as constitutively active are limited. The results concerning the level of the basal activity can be influenced by the vector and/or the cell system. A comprehensive clinical characterization of the respective patients appears as a necessary and promising adjunct for the activity classification of these borderline mutations.

Angiotensin converting enzyme gene polymorphism in Turkish asthmatic patients.

Asthma is a chronic inflammatory disease of the airways. Several candidate genes have been identified with a potential role in the pathogenesis of asthma, including the angiotensin converting enzyme (ACE) gene. We aimed to investigate the frequency of an ACE gene polymorphism in Turkish asthmatic patients and to determine its impact on clinical parameters and disease severity. Ninety-seven asthmatic patients (M/F 25/72, mean age 39 +/- 13 years) and 96 healthy subjects (M/F 26/70, mean age 38 +/- 12 years) were included. At baseline, all participants completed a questionnaire on demographics, symptoms, triggering factors, severity of asthma, and the presence of atopism. Blood samples were obtained from all patients and genomic DNA was isolated. The frequency of the ACE genotypes (I = insertion and D = deletion) among asthmatics and controls were compared: asthmatics showed a 40.2% prevalence of the DD genotype (n = 39), ID was 45.4% (n = 44), and II was 14.4% (n = 14.4). In the control subjects, the frequency of DD was 18.8% (n = 18), ID was 50% (n = 48) and II was 31.3% (n = 30). The DD ACE genotype was significantly more frequent in asthmatics compared with controls (p < 0.001). Asthmatics with the ID ACE genotype showed a higher frequency of drug allergies, although this was not statistically significant (p = 0.08). Asthmatics with the DD genotype appeared to have a higher incidence of asthmatic episode exacerbations due to viral infections, but again this was not statistically significant (p = 0.08). Patients with mild or moderate-severe asthma had similar frequencies of these mutations. We found a higher frequency of the ACE DD gene mutation in Turkish asthmatic patients compared with non-asthmatics, suggesting that this ACE gene polymorphism may be a risk factor for asthma but does not increase the severity of the disease.

Alopecia: association with resistance to thyroid hormones.

UNLABELLED: Resistance to thyroid hormone (RTH) syndrome is caused by thyroid hormone beta receptor (TRbeta) mutations. Goiter, learning disabilities, psychological abnormalities, sinus tachycardia, hearing deficits, short stature, and growth delay are among the most common symptoms in patients with RTH. Alopecia areata (AA) is an autoimmune disease of the hair follicle, frequently associated with other autoimmune disorders. In some cases local alopecia of different genetic backgrounds could be misdiagnosed as AA. We describe here clinical, biochemical and genetic features of a family having RTH syndrome, caused by a novel TRbeta mutation, coexistent with alopecia. Mutational analyses of the TRbeta gene and the hairless gene (HR) in genomic DNA were performed. The index patient is a 9-4/12 year-old boy with RTH due to a novel heterozygous missense mutation of the TRbeta gene (I353V), and diffuse, patchy alopecia without autoimmune thyroid disease. This mutation was also detected in his father and elder brother, who also have local alopecia. One of his paternal aunts and paternal grandmother have local alopecia and they have previously been operated for goiter. Although they refused any genetic analysis, the pre-operative medical report of the paternal aunt was compatible with RTH. A second paternal aunt has alopecia totalis universalis but has no RTH mutation in genomic DNA. Genomic DNA sequence of the HR gene of the family (index patient, two brothers, father, mother and second paternal aunt) was normal as well. CONCLUSION: We speculate that RTH due to a novel I353V TRbeta mutation could be causally related to different phenotypic expressions of alopecia in this family, either by a direct effect of unresponsiveness to T3 of the hair follicle or by the modulated action of the HR gene.

MTHFR, prothrombin and Factor V gene variants in Turkish patients with coronary artery stenosis

Many epidemiological studies have reported an association between hemostatic factors and risk of both coronary and peripheral artery diseases. Using polymerase chain reaction and restriction fragment length polymorphism (PCR-RFLP) analysis, we investigated the association between coronary artery disease and polymorphisms in the methylenetetrahydrofolate reductase (MTHFR C677T and A1298C), prothrombin (G20210A), and factor V (A4070G) genes. We screened these gene variants in 174 subjects who had undergone coronary angiography - 115 patients with patent coronary artery disease (grade 3 vessel disease, i.e., significant coronary stenosis), and 59 healthy controls with grade 0 vessel disease. The analysis of our data did not show any statistically significant association between coronary artery disease (CAD) and the investigated polymorphisms.

A new silent germline mutation of the TSH receptor: coexpression in a hyperthyroid family member with a second activating somatic mutation.

BACKGROUND: Up to date, three thyroid-stimulating hormone receptor (TSHR) germline variants have been reported for which no functional consequences have been detected by in vitro characterizations. However, familial nonautoimmune hyperthyroidism and hot nodules are clearly associated with constitutively activating TSHR germline mutations. We describe a family with a new TSHR germline mutation that is associated with euthyroidism in 13 family members and hyperthyroidism in 1 family member. METHODS: Mutation analysis of the TSHR gene was performed by denaturing gradient gel electrophoresis. TSHR constructs were characterized by determination of cell surface expression, 3'-5'-cyclic adenosine monophosphate (cAMP) accumulation, and constitutive cAMP activity. RESULTS: A novel TSHR germline mutation (N372T) was found in a man who presented with thyrotoxicosis. The mutation was also detected in 13 family members, all of whom were euthyroid. Interestingly, an additional constitutively active somatic mutation (S281N) was identified on the second parental TSHR allele of the hyperthyroid index patient. Linear regression analysis showed a lack of constitutive activity for N372T. Moreover, coexpression studies of N372T with S281N did not reveal any evidence for a functional influence of N372T on the constitutively active mutation (CAM). CONCLUSIONS: N372T is unlikely to cause altered thyroid function. This is consistent with the finding that only the index patient with the additional somatic mutation S281N was hyperthyroid.