RESUMEN
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.
Asunto(s)
Antioxidantes , Melatonina , Mitocondrias/efectos de los fármacos , Estrés Oxidativo/efectos de los fármacos , Síndrome de Wolf-Hirschhorn/tratamiento farmacológico , Síndrome de Wolf-Hirschhorn/metabolismo , Animales , Antioxidantes/farmacología , Antioxidantes/uso terapéutico , Proteínas de Unión al Calcio/genética , Proteínas de Transporte de Catión/genética , Línea Celular , Respiración de la Célula/efectos de los fármacos , Embrión de Mamíferos , Fibroblastos , Silenciador del Gen , Melatonina/farmacología , Melatonina/uso terapéutico , Ratones , Fosforilación Oxidativa/efectos de los fármacos , Oxígeno/metabolismo , Especies Reactivas de Oxígeno/metabolismo , Superóxido Dismutasa/metabolismo , Síndrome de Wolf-Hirschhorn/genéticaRESUMEN
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.
Asunto(s)
Enfermedad de Graves , Hipertiroidismo , Adulto , Niño , Femenino , Mutación de Línea Germinal , Humanos , Hipertiroidismo/congénito , Hipertiroidismo/tratamiento farmacológico , Hipertiroidismo/genética , Lactante , Recién Nacido , Receptores de Tirotropina/genéticaRESUMEN
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.
Asunto(s)
Saltamontes , Animales , ADN Mitocondrial/genética , Saltamontes/genética , Filogenia , Filogeografía , Análisis de Secuencia de ADNRESUMEN
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.
Asunto(s)
Virus de la Fiebre Hemorrágica de Crimea-Congo , Fiebre Hemorrágica de Crimea/transmisión , Garrapatas/virología , Tortugas/virología , Zoonosis/virología , Animales , Vectores Arácnidos/virología , ADN Viral , Reservorios de Enfermedades/virología , Virus de la Fiebre Hemorrágica de Crimea-Congo/genética , Virus de la Fiebre Hemorrágica de Crimea-Congo/aislamiento & purificación , Filogenia , Reacción en Cadena de la Polimerasa , Turquía/epidemiología , Tortugas/parasitologíaRESUMEN
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.
Asunto(s)
Cromograninas/genética , Subunidades alfa de la Proteína de Unión al GTP Gs/genética , Mutación , Receptores de Tirotropina/genética , Análisis de Secuencia de ADN , Nódulo Tiroideo/genética , Nódulo Tiroideo/metabolismo , Proteínas Portadoras/genética , Análisis Mutacional de ADN , Nucleótidos de Guanina , Secuenciación de Nucleótidos de Alto Rendimiento , Humanos , Yodo/metabolismo , Prevalencia , Sensibilidad y Especificidad , Programas InformáticosRESUMEN
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.
Asunto(s)
Asma/epidemiología , Asma/genética , Predisposición Genética a la Enfermedad/epidemiología , Peptidil-Dipeptidasa A/genética , Polimorfismo Genético , Adulto , Distribución por Edad , Asma/fisiopatología , Estudios de Casos y Controles , Distribución de Chi-Cuadrado , Femenino , Frecuencia de los Genes , Genotipo , Humanos , Incidencia , Masculino , Persona de Mediana Edad , Peptidil-Dipeptidasa A/metabolismo , Reacción en Cadena de la Polimerasa , Probabilidad , Valores de Referencia , Pruebas de Función Respiratoria , Medición de Riesgo , Sensibilidad y Especificidad , Índice de Severidad de la Enfermedad , Distribución por Sexo , Turquía/epidemiologíaRESUMEN
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.
Asunto(s)
Alopecia Areata/etiología , Receptores beta de Hormona Tiroidea/genética , Síndrome de Resistencia a Hormonas Tiroideas/complicaciones , Alopecia Areata/genética , Niño , Análisis Mutacional de ADN , Salud de la Familia , Femenino , Bocio , Humanos , Masculino , Mutación Missense , Linaje , Síndrome de Resistencia a Hormonas Tiroideas/genética , Factores de Transcripción/genéticaRESUMEN
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.
Asunto(s)
Antibacterianos/farmacología , Daptomicina/farmacología , Glicopéptidos/farmacología , Disco Intervertebral/efectos de los fármacos , Adulto , Proliferación Celular/efectos de los fármacos , Proliferación Celular/fisiología , Células Cultivadas , Evaluación Preclínica de Medicamentos/métodos , Femenino , Humanos , Disco Intervertebral/citología , Disco Intervertebral/fisiología , MasculinoRESUMEN
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.
Asunto(s)
Biomarcadores de Tumor/genética , ADN Mitocondrial/genética , ADN de Neoplasias/genética , Mutación , Cáncer Papilar Tiroideo , Neoplasias de la Tiroides , Femenino , Humanos , Masculino , Pronóstico , Cáncer Papilar Tiroideo/genética , Neoplasias de la Tiroides/diagnóstico , Neoplasias de la Tiroides/genética , TurquíaRESUMEN
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.
Asunto(s)
Carcinoma Papilar/genética , Radioisótopos de Yodo/uso terapéutico , Proteínas Proto-Oncogénicas B-raf/genética , Neoplasias de la Tiroides/genética , Adulto , Carcinoma Papilar/patología , Carcinoma Papilar/radioterapia , Análisis Mutacional de ADN , Femenino , Humanos , Masculino , Persona de Mediana Edad , Invasividad Neoplásica/genética , Proteínas Proto-Oncogénicas B-raf/efectos de la radiación , Neoplasias de la Tiroides/patología , Neoplasias de la Tiroides/radioterapiaRESUMEN
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.
Asunto(s)
Mutación de Línea Germinal/genética , Hipertiroidismo/genética , Receptores de Tirotropina/genética , Adolescente , Adulto , Anciano , Niño , AMP Cíclico/metabolismo , Análisis Mutacional de ADN , Femenino , Estudios de Seguimiento , Predisposición Genética a la Enfermedad , Humanos , Modelos Lineales , Masculino , Persona de Mediana Edad , Linaje , Tirotoxicosis/genéticaRESUMEN
The studies for the development of transgenic mice models which provide important profits for the studies concerning immunopathogenesis of hepatitis B virus (HBV) infections are in progress since 20 years. For this purpose different lineages bearing whole HBV genome or selected viral genes have been developed and their usage in clarifying the HBV replication and pathogenesis mechanisms have been emphasized. The aim of this study was to develop and breed a HBV carrier mice model. In the study the full HBV genome has been transferred to mouse embryos by microinjection procedure. Following transgenic manipulation, the HBV carriers among the daughter mice have been detected by molecular methods in which HBV-DNA replication and expression have been shown. The manipulations for transgene transfers have been performed in TUBITAK Marmara Research Center Transgene Laboratory, Gebze, Istanbul. The HBV-DNA carrier mice have been demonstrated by polymerase chain reaction (PCR) using the DNA samples obtained from tail tissues and also by dot-blot hybridization of the mice sera. Integrated HBV-DNA has been detected by applying in-situ hybridization to the liver tissue sections. HBV-DNA expression has been shown by reverse transcriptase PCR method with total RNA molecules that have been isolated from the liver tissues of the HBV-DNA carrier mice. HBsAg has been detected in the liver by immunohistochemical method, and HBsAg and HBeAg have additionally been demonstrated by ELISA. HBV genome, expression of the genome and the expression products have been determined in approximately 10% of the mice of which HBV-DNA have been transferred. By inbreeding heterozygote carrier mice, homozygote HBV transgenic mice line have been obtained. These HBV transgenic mice are the first lineages developed in our country. It is hopefully thought that this HBV carrier transgenic mouse model may contribute to the studies on the pathogenesis of HBV infections which are important health problems in the world as well as in Turkey.
Asunto(s)
Portador Sano/inmunología , Modelos Animales de Enfermedad , Virus de la Hepatitis B/genética , Hepatitis B/etiología , Ratones Transgénicos , Animales , Portador Sano/virología , ADN Viral/análisis , Genoma Viral/genética , Hepatitis B/inmunología , Ratones , MicroinyeccionesRESUMEN
OBJECTIVE: Differences in iodine intake could account for the variable prevalences reported for somatic TSH receptor (TSHR) mutations in toxic thyroid nodules (TTNs). However, this question has not been settled, since no study has yet determined the TSHR mutation prevalence in regions with different iodine supplies in the same population using the same methodology. Therefore, we studied the prevalence of somatic TSHR mutations in TTNs from patients living in iodine-deficient or -sufficient regions in Turkey. DESIGN AND METHODS: We screened 74 TTNs for somatic TSHR mutations. Exons 9 and 10 of the TSHR and 7 and 8 of the Gsalpha were screened by denaturing gradient gel electrophoresis. Determination of X-chromosome inactivation was used for clonality analysis. RESULTS: TSHR mutations were identified in 52 (70.2%) of 74 TTNs. A Gsalpha mutation was identified in one TTN. Three new TSHR mutations were detected (A627V, I640K, I486N). No significant difference between frequencies of TSHR mutations in iodine deficient/sufficient regions was found. The frequency of non-random X-chromosome inactivation was similar in iodine-sufficient or -deficient regions and in TSHR mutation positive or negative hot nodules. CONCLUSIONS: These findings suggest that TTNs in iodine deficient/sufficient areas predominantly arise from aberrant growth of a single cell. Our results suggest that neither the prevalence of TSHR mutations nor that of monoclonal TTNs is related to iodine supply.
Asunto(s)
Subunidades alfa de la Proteína de Unión al GTP Gs/genética , Yodo/provisión & distribución , Receptores de Tirotropina/genética , Nódulo Tiroideo/epidemiología , Tirotoxicosis/epidemiología , Adulto , Anciano , Análisis Mutacional de ADN , Femenino , Geografía , Humanos , Masculino , Persona de Mediana Edad , Mutación , Polimorfismo Conformacional Retorcido-Simple , Prevalencia , Nódulo Tiroideo/genética , Tirotoxicosis/genética , Turquía/epidemiologíaRESUMEN
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.
Asunto(s)
Mutación , Complejo Represivo Polycomb 2/genética , Neoplasias de la Tiroides/genética , Adulto , Anciano , Dominio Catalítico , Diferenciación Celular , Proliferación Celular , Femenino , Subunidades alfa de la Proteína de Unión al GTP Gs/metabolismo , Regulación Neoplásica de la Expresión Génica , Células HEK293 , Humanos , Masculino , Persona de Mediana Edad , Receptores de Tirotropina/genética , Programas Informáticos , Glándula Tiroides/patologíaRESUMEN
Autonomously functioning thyroid nodules (AFTNs) can present as hyperfunctioning adenomas or toxic multinodular goiters. In the last decade, a large number of activating mutations have been identified in the thyrotropin receptor (TSHR) gene in autonomously functioning thyroid nodules. Most have been situated close to, or within the sixth transmembrane segment and third intracellular loop of the TSHR where the receptor interacts with the Gs protein. In this study we describe two novel mutations in the sixth transmembrane segment of the TSHR causing hyperfunctioning thyroid nodules. Genomic DNAs were isolated from four hyperfunctioning thyroid nodules, normal tissues and peripheral leukocytes of two patients with toxic multinodular goiter. After amplifying the related regions, TSHR and G(s)alpha genes were analyzed by single-strand conformation polymorphism (SSCP) analysis. The precise localization of the mutations was identified by automatic DNA sequence analysis. Functional studies were done by site-directed mutagenesis and transfection of a mutant construct into COS-7 cells. We identified two novel TSHR mutations in two hyperfunctioning thyroid nodules: Phe631Val in the first patient and Iso630Met in the second patient. Both mutant receptors display an increase in constitutive stimulation of basal cyclic adenosine monophosphate (cAMP) levels compared to the wild-type receptor. This confirms that these mutant receptors cause hyperfunctioning thyroid nodules.
Asunto(s)
Mutación , Receptores de Tirotropina/genética , Nódulo Tiroideo/etiología , Adulto , Femenino , Humanos , Persona de Mediana Edad , Polimorfismo Conformacional Retorcido-SimpleRESUMEN
In the last decade, studies were first done to determine the frequency of Gsalpha and later thyrotropin receptor (TSHR) mutations in benign autonomously functioning thyroid nodules (AFTN). Different frequencies ranging from 0% to 38% for GSp mutations and from 20% to 86% for TSHR mutations were found. There were only some limited case reports related to TSHR genetic alterations in malignant AFTN. Their role in autonomously functioning thyroid carcinomas is not well established. We present a patient who had thyroidectomy for toxic multinodular goiter and a papillary carcinoma was demonstrated histopathologically. Genomic DNA was isolated from two solid areas in the hot nodule and peripheral leukocytes of the patient. After amplifying the related regions, TSHR and GSalpha genes were analyzed by single-strand conformation polymorphism (SSCP) analysis. The precise localization of the mutations was identified by automatic DNA sequence analysis. An activating mutation of the TSHR gene (Leu 512 Arg) was found in the autonomously functioning papillary carcinoma. It is believed that this mutation causes constitutive activation of the cyclic adenosine monophosphate (cAMP) signal transduction pathway and thereby causes thyrotoxicosis and a hot thyroid nodule in an autonomously functioning papillary carcinoma.
Asunto(s)
Carcinoma Papilar/fisiopatología , Mutación , Receptores de Tirotropina/genética , Neoplasias de la Tiroides/fisiopatología , Secuencia de Aminoácidos , Arginina , Secuencia de Bases , Carcinoma Papilar/diagnóstico por imagen , Carcinoma Papilar/genética , Carcinoma Papilar/patología , Carcinoma Papilar/cirugía , Femenino , Bocio Nodular/cirugía , Humanos , Leucina , Polimorfismo Conformacional Retorcido-Simple , Cintigrafía , Neoplasias de la Tiroides/diagnóstico por imagen , Neoplasias de la Tiroides/genética , Neoplasias de la Tiroides/patología , Neoplasias de la Tiroides/cirugía , TiroidectomíaRESUMEN
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.
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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.
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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.
Asunto(s)
Encéfalo/anomalías , Labio Leporino/genética , Fisura del Paladar/genética , ADN/genética , Metilenotetrahidrofolato Reductasa (NADPH2)/genética , Padres , Polimorfismo Genético , Adolescente , Adulto , Alelos , Encéfalo/metabolismo , Niño , Preescolar , Labio Leporino/epidemiología , Labio Leporino/metabolismo , Fisura del Paladar/epidemiología , Fisura del Paladar/metabolismo , Femenino , Frecuencia de los Genes , Predisposición Genética a la Enfermedad , Genotipo , Humanos , Incidencia , Lactante , Recién Nacido , Masculino , Metilenotetrahidrofolato Reductasa (NADPH2)/metabolismo , Factores de Riesgo , Turquía/epidemiología , Adulto JovenRESUMEN
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.