RESUMO
OBJECTIVE: Nodular goiter may increase the risk of thyroid cancer, but the genetic factors contributing to nodular goiter are not well understood. There is an overexpression of H19 lncRNA in goiter tissue and its target remains unknown. In this study, we attempted to identify a new target for H19 in the context of goiter development. METHODS: Using interaction energy calculations, the interaction between NKX2-1 mRNA and H19 lncRNA was examined. Putative microRNAs were found at the H19 lncRNA target site with the highest affinity for NKX2-1. RNAseq data was analyzed to determine the tissue specificity of gene expression. Samples were taken from 18 goiter and 18 normal tissues during thyroidectomy. The expression of NKX2-1 was determined by RT-qPCR using specific primers. RESULTS: The interaction between NKX2-1 and H19 was characterized by six local base-pairing connections, with a maximum energy of -20.56â¯kcal/moL. Specifically, the sequence that displayed the highest affinity for binding with H19 overlapped with the binding site of has-miR-1827 to NKX2-1. It was found that NKX2-1 is exclusively co-expressed with H19 in normal thyroid tissue. As compared to adjacent normal tissues, nodular goiter tissues have a significant overexpression of NKX2-1 (relative expressionâ¯=â¯1.195, pâ¯=⯠0.038). CONCLUSION: NKX2-1 has been identified as the putative target of H19 lncRNA, which is overexpressed in nodular goiter tissues significantly.
Assuntos
Bócio Nodular , RNA Longo não Codificante , Fator Nuclear 1 de Tireoide , Humanos , RNA Longo não Codificante/genética , RNA Longo não Codificante/metabolismo , Bócio Nodular/genética , Fator Nuclear 1 de Tireoide/genética , Fator Nuclear 1 de Tireoide/metabolismo , Feminino , Masculino , Adulto , MicroRNAs/genética , MicroRNAs/metabolismo , Pessoa de Meia-Idade , RNA Mensageiro/metabolismo , RNA Mensageiro/genética , Estudos de Casos e ControlesRESUMO
Objective: Mutations in DICER1 are found in differentiated thyroid carcinoma (DTC) and in multinodular goiter (MNG) at a younger age with other tumors, which characterizes DICER1 syndrome. DICER1 is one driver to DTC; however, it is also found in benign nodules. We speculated that patients with mutations in DICER1 may present long-lasting MNG. Our aim was to investigate the frequency of DICER1 variants in patients with MNG. Subjects and methods: Patients who submitted to total thyroidectomy due to large MNG with symptoms were evaluated. DICER1 hotspots were sequenced from thyroid nodule samples. To confirm somatic mutation, DNA from peripheral blood was also analyzed. Results: Among 715 patients, 154 were evaluated with 56.2 ± 12.3 years old (28-79) and the thyroid volume was 115.7 ± 108 mL (16.2-730). We found 11% with six DICER1 variations in a homo or heterozygous state. Only rs12018992 was a somatic DICER1 variant. All remaining variants were synonymous and likely benign, according to the ClinVar database. The rs12018992 was previously described in an adolescent with DTC, measuring 13 mm. There were no significant differences according to gender, familial history of goiter, age, thyroid volume, TSH and TI-RADS classification between DICER1 carriers. Free T4 were lower in patients with DICER1 polymorphisms (13.77 ± 1.8 vs. 15.44 ± 2.4 pmol/L, p = 0.008), regardless of TSH levels. Conclusion: We conclude that germline DICER1 variants can be found in 11% of large goiters but no second-hit somatic mutation was found. DICER1 is one driver to thyroid lesion and a second-hit event seems unnecessary in the MNG development.
Assuntos
Adenocarcinoma , RNA Helicases DEAD-box , Ribonuclease III , Neoplasias da Glândula Tireoide , Adolescente , Adulto , Idoso , Humanos , Pessoa de Meia-Idade , Adenocarcinoma/diagnóstico por imagem , Adenocarcinoma/genética , RNA Helicases DEAD-box/genética , Bócio Nodular/genética , Bócio Nodular/diagnóstico , Prevalência , Ribonuclease III/genética , Neoplasias da Glândula Tireoide/diagnóstico por imagem , Neoplasias da Glândula Tireoide/genética , Neoplasias da Glândula Tireoide/patologia , TireotropinaRESUMO
INTRODUCTION: The goiter, a neglected heterogeneous molecular disease, remains a major indication for thyroidectomies in its endemic regions. OBJECTIVES: This study analyzed differential gene expression in surgical specimens diagnosed with multi nodular and compared the data to that of thyroid tissue without multinodular goiter from patients undergoing thyroidectomy in Manaus-AM, Brazil using RNA-seq technology. METHODOLOGY: The transcriptome information of the surgical specimen fragments with and without multinodular goiter was accessed by Illumina HiSeq 2000 New Generation Sequencing (NGS) using the RNA-seq NEBNext® Ultra™ RNA Library Prep Kit for Illumina®-#E7530L protocol and differential gene expression analysis. RESULTS: Differences were found between the gene expression profiles of the diseased tissues and those of the healthy control tissues; at least 70 genes were differentially expressed. The HOTS gene was expressed only in multinodular goiter tissues (p < 0.05). CONCLUSION: These results demonstrate that the gene expression profile of multinodular goiter is pro-tumoral and that HOTS can play a central role in multinodular goiter development.
Assuntos
Bócio Nodular , Biblioteca Gênica , Bócio Nodular/genética , Bócio Nodular/cirurgia , Humanos , Tireoidectomia/métodos , TranscriptomaRESUMO
Deregulation of VEGFA (Vascular Endothelial Growth Factor A) and NFE2L2 (Nuclear Factor (Erythroid-derived 2)-Like 2), involved in angiogenesis and oxidative stress, can lead to thyroid cancer progression. MiR-17-5p and miR-612 are possible regulators of these genes and may promote thyroid disorders. In order to evaluate the involvement of VEGFA, NFE2L2, hsa-miR-17-5p, and hsa-miR-612 in thyroid pathology, we examined tissue samples from colloid goiter, papillary thyroid cancer (PTC), and a normal thyroid. We found higher levels of VEGFA and NFE2L2 transcripts and the VEGFA protein in goiter and PTC samples than in normal tissue. In the goiter, miR-612 and miR-17-5p levels were lower than those in PTC. Tumors, despite showing lower VEGFA mRNA expression, presented higher VEGFA protein levels compared to goiter tissue. In addition, NRF2 (Nuclear Related Transcription Factor 2) protein levels in tumors were higher than those in goiter and normal tissues. Inhibition of miR-17-5p resulted in reduced NFE2L2 expression. Overall, both transcript and protein levels of NFE2L2 and VEGFA were elevated in PTC and colloid goiter. Hsa-miR-612 showed differential expression in PTC and colloid goiter, while hsa-miR-17-5p showed differential expression only in colloid goiter, suggesting that hsa-miR-17-5p may be a positive regulator of NFE2L2 expression in PTC.
Assuntos
Biomarcadores Tumorais/metabolismo , Bócio Nodular/patologia , MicroRNAs/genética , Fator 2 Relacionado a NF-E2/metabolismo , Câncer Papilífero da Tireoide/patologia , Fator A de Crescimento do Endotélio Vascular/metabolismo , Apoptose , Biomarcadores Tumorais/genética , Estudos de Casos e Controles , Proliferação de Células , Feminino , Regulação Neoplásica da Expressão Gênica , Bócio Nodular/genética , Bócio Nodular/metabolismo , Humanos , Masculino , Pessoa de Meia-Idade , Fator 2 Relacionado a NF-E2/genética , Prognóstico , Câncer Papilífero da Tireoide/genética , Câncer Papilífero da Tireoide/metabolismo , Neoplasias da Glândula Tireoide/genética , Neoplasias da Glândula Tireoide/metabolismo , Neoplasias da Glândula Tireoide/patologia , Células Tumorais Cultivadas , Fator A de Crescimento do Endotélio Vascular/genéticaRESUMO
BACKGROUND: Mutations in the SLC26A4 gene are associated with Pendred syndrome and autosomal recessive non-syndromic deafness (DFNB4). Both disorders have similar audiologic characteristics: bilateral hearing loss, often severe or profound, which may be associated with abnormalities of the inner ear, such as dilatation of the vestibular aqueduct or Mondini dysplasia. But, in Pendred syndrome (OMIM #274600), with autosomal recessive inheritance, besides congenital sensorineural deafness, goiter or thyroid dysfunctions are frequently present. The aim of this study was to determine whether mutations in SLC26A4 are a frequent cause of hereditary deafness in Brazilian patients. METHODS: Microsatellite haplotypes linked to SLC26A4 were investigated in 68 families presenting autosomal recessive non-syndromic deafness. In the probands of the 16 families presenting segregation consistent with linkage to SLC26A4, Sanger sequencing of the 20 coding exons was performed. In an additional sample of 15 individuals with suspected Pendred syndrome, because of the presence of hypothyroidism or cochleovestibular malformations, the SLC26A4 gene coding region was also sequenced. RESULTS: In two of the 16 families with indication of linkage to SLC26A4, the probands were found to be compound heterozygotes for probably pathogenic different mutations: three novel (c.1003 T > G (p. F335 V), c.1553G > A (p.W518X), c.2235 + 2 T > C (IVS19 + 2 T > C), and one already described, c.84C > A (p.S28R). Two of the 15 individuals with suspected Pendred syndrome because of hypothyreoidism or cochleovestibular malformations were monoallelic for likely pathogenic mutations: a splice mutation (IVS7 + 2 T > C) and the previously described c.1246A > C (p.T416P). Pathogenic copy number variations were excluded in the monoallelic cases and in those with normal results after Sanger sequencing. Additional mutations in the SLC26A4 gene or other definite molecular cause for deafness were not identified in the monoallelic patients, after exome sequencing. CONCLUSIONS: Biallelic pathogenic mutations in SLC26A4 explained ~ 3% of cases selected because of autosomal recessive deafness. Monoallelic mutations were present in ~ 13% of isolated cases of deafness with cochleovestibular malformations or suspected Pendred syndrome. These data reinforce the importance of mutation screening of SLC26A4 in Brazilian subjects and highlight the elevated frequency of monoallelic patients.
Assuntos
Bócio Nodular/genética , Perda Auditiva Neurossensorial/genética , Mutação , Análise de Sequência de DNA/métodos , Transportadores de Sulfato/genética , Brasil , Análise Mutacional de DNA , Feminino , Haplótipos , Humanos , Masculino , Repetições de Microssatélites , LinhagemRESUMO
ABSTRAC This article presents the results of a comprehensive analysis of the combined influence of genetic polymorphisms associated with various links of apoptosis regulation (BCL-2, CTLA-4 and APO-1/Fas) on the development of nodular goiter with autoimmune thyroiditis and thyroid adenoma in the studied population. The analysis was performed using the Multifactor Dimensionality Reduction (MDR) method by calculating the prediction potential. Graphic models of gene-gene interaction with the highest cross-validation consistency created by the MDR method showed complex "synergistic or independent" impact of polymorphic loci of the CTLA-4 (+49G/A), Fas (-1377G/A) and BCL-2 (63291411 A>G) genes on the onset of thyroid pathology in general, or its individual types (nodular goiter with autoimmune thyroiditis and thyroid adenoma) in the population of Northern Bukovyna.
RESUMEN Este artículo presenta los resultados de un análisis exhaustivo de la influencia combinada de polimorfismos genéticos asociados a diversos enlaces en la regulación de la apoptosis (BCL-2, CTLA-4 y APO-1/FAS) sobre el desarrollo de bocio nodular con tiroiditis autoinmune y adenoma tiroideo en la población estudiada. Para ello, se utilizó el método de reducción de dimensionalidad multifactorial (MDR) mediante el cálculo de los potenciales de predicción. Los modelos gráficos de interacción gen-gen con la mayor consistencia de validación cruzada creada por el método MDR mostraron un complejo impacto «sinérgico o independiente¼ de los loci polimórficos de los genes CTLA-4 (+49G/A), FAS (-1377G/A) y BCL-2 (63291411A>G) en el inicio de la patología tiroidea en general, o sus tipos individuales (bocio nodular con tiroiditis autoinmune y adenoma tiroideo) en la población de Bucovina septentrional.
Assuntos
Humanos , Feminino , Adulto , Pessoa de Meia-Idade , Idoso , Polimorfismo Genético/fisiologia , Tireoidite Autoimune/genética , Neoplasias da Glândula Tireoide/genética , Bócio Nodular/fisiopatologia , Bócio Nodular/genética , Apoptose/fisiologia , Receptor fas/análise , Genes bcl-2/genética , Redução Dimensional com Múltiplos Fatores/métodos , Abatacepte/análise , Bócio Nodular/etiologiaRESUMO
The D727E germline polymorphism in the thyroid-stimulating hormone receptor gene (TSHR) may cause genetic susceptibility to the development of goiter. Therefore, in this study we investigated allele frequencies and genotype distributions of the TSHR D727E polymorphism, their association with clinical parameters, and the development of goiter in the Turkish population. We investigated the TSHR D727E polymorphism in 123 patients and 97 healthy subjects using the polymerase chain reaction-restriction fragment length polymorphism technique. Peripheral blood was used for DNA extraction. Although no significant difference was found in TSHR D727E polymorphism frequencies between the patients with nodular goiters (26/123 patients, 21.1%) and the controls (12/97 patients, 12.4%) (P = 0.107), the frequency of the TSHR D727E polymorphism in the hyperthyroid+subclinical hyperthyroid patient groups (23%) was significantly higher than in the control subjects (12.4%) (P = 0.024). In this study, nodular goiter presented significantly earlier in GC genotype patients (mean age 35 years) than in CC genotype patients (mean age 42 years) in the hyperthyroid group (P = 0.009). More importantly, TSH levels in the GC variant controls were closely significant lower (1.26 ± 0.49) than in the CC variant controls (1.74 ± 0.84) (P = 0.053). The TSHR D727E polymorphism might be involved in the pathogenesis of toxic multinodular goiter (TMNG). Moreover, this polymorphism might be an indication of early-onset TMNG. However, development of MNG is multifactorial. Therefore, further case-control studies with larger populations are required to verify these observations.
Assuntos
Alelos , Predisposição Genética para Doença , Bócio Nodular/genética , Polimorfismo Genético , Receptores da Tireotropina/genética , Estudos de Casos e Controles , Frequência do Gene , Genótipo , Humanos , Reação em Cadeia da Polimerase , Polimorfismo de Fragmento de Restrição , TurquiaRESUMO
ABSTRACT Objective Pendred syndrome (PS) is an autosomal recessive disorder characterised by sensorineural hearing loss and thyroid dyshormonogenesis. It is caused by biallelic mutations in the SLC26A4 gene encoding for pendrin. Hypothyroidism in PS can be present from birth and therefore diagnosed by neonatal screening. The aim of this study was to examine the SLC26A4 mutation spectrum and prevalence among congenital hypothyroidism (CH) patients in the Guangxi Zhuang Autonomous Region of China and to establish how frequently PS causes hearing impairment in our patients with CH. Subjects and methods Blood samples were collected from 192 CH patients in Guangxi Zhuang Autonomous Region, China, and genomic DNA was extracted from peripheral blood leukocytes. All exons of the SLC26A4 gene together with their exon-intron boundaries were screened by next-generation sequencing. Patients with SLC26A4 mutations underwent a complete audiological evaluation including otoscopic examination, audiometry and morphological evaluation of the inner ear. Results Next generation sequencing analysis of SLC26A4 in 192 CH patients revealed five different heterozygous variations in eight individuals (8/192, 4%). The prevalence of SLC26A4 mutations was 4% among studied Chinese CH. Three of the eight were diagnosed as enlargement of the vestibular aqueduct (EVA), no PS were found in our 192 CH patients. The mutations included one novel missense variant p.P469S, as well as four known missense variants, namely p.V233L, p.M147I, p.V609G and p.D661E. Of the eight patients identified with SLC26A4 variations in our study, seven patients showed normal size/location of thyroid gland, and one patients showed a decreased size one. Conclusions The prevalence of SLC26A4 pathogenic variants was 4% among studied Chinese patients with CH. Our study expanded the SLC26A4 mutation spectrum, provided the best estimation of SLC26A4 mutation rate for Chinese CH patients and indicated the rarity of PS as a cause of CH.
Assuntos
Humanos , Masculino , Feminino , Recém-Nascido , Proteínas de Membrana Transportadoras/genética , Testes Genéticos/métodos , Bócio Nodular/genética , Perda Auditiva Neurossensorial/genética , Mutação , Tiroxina/sangue , Aqueduto Vestibular/anormalidades , Tireotropina/sangue , China/epidemiologia , Prevalência , Estudos de Coortes , Triagem Neonatal/métodos , Transportadores de Sulfato , Bócio Nodular/epidemiologia , Perda Auditiva Neurossensorial/epidemiologiaRESUMO
OBJECTIVE: Pendred syndrome (PS) is an autosomal recessive disorder characterised by sensorineural hearing loss and thyroid dyshormonogenesis. It is caused by biallelic mutations in the SLC26A4 gene encoding for pendrin. Hypothyroidism in PS can be present from birth and therefore diagnosed by neonatal screening. The aim of this study was to examine the SLC26A4 mutation spectrum and prevalence among congenital hypothyroidism (CH) patients in the Guangxi Zhuang Autonomous Region of China and to establish how frequently PS causes hearing impairment in our patients with CH. SUBJECTS AND METHODS: Blood samples were collected from 192 CH patients in Guangxi Zhuang Autonomous Region, China, and genomic DNA was extracted from peripheral blood leukocytes. All exons of the SLC26A4 gene together with their exon-intron boundaries were screened by next-generation sequencing. Patients with SLC26A4 mutations underwent a complete audiological evaluation including otoscopic examination, audiometry and morphological evaluation of the inner ear. RESULTS: Next generation sequencing analysis of SLC26A4 in 192 CH patients revealed five different heterozygous variations in eight individuals (8/192, 4%). The prevalence of SLC26A4 mutations was 4% among studied Chinese CH. Three of the eight were diagnosed as enlargement of the vestibular aqueduct (EVA), no PS were found in our 192 CH patients. The mutations included one novel missense variant p.P469S, as well as four known missense variants, namely p.V233L, p.M147I, p.V609G and p.D661E. Of the eight patients identified with SLC26A4 variations in our study, seven patients showed normal size/location of thyroid gland, and one patients showed a decreased size one. CONCLUSIONS: The prevalence of SLC26A4 pathogenic variants was 4% among studied Chinese patients with CH. Our study expanded the SLC26A4 mutation spectrum, provided the best estimation of SLC26A4 mutation rate for Chinese CH patients and indicated the rarity of PS as a cause of CH.
Assuntos
Testes Genéticos/métodos , Bócio Nodular/genética , Perda Auditiva Neurossensorial/genética , Proteínas de Membrana Transportadoras/genética , Mutação , China/epidemiologia , Estudos de Coortes , Feminino , Bócio Nodular/epidemiologia , Perda Auditiva Neurossensorial/epidemiologia , Humanos , Recém-Nascido , Masculino , Triagem Neonatal/métodos , Prevalência , Transportadores de Sulfato , Tireotropina/sangue , Tiroxina/sangue , Aqueduto Vestibular/anormalidadesRESUMO
BACKGROUND DREAM (Downstream Regulatory Element Antagonistic Modulator) is a neuronal calcium sensor that was suggested to modulate TSH receptor activity and whose overexpression provokes an enlargement of the thyroid gland in transgenic mice. The aim of this study was to investigate somatic mutations and DREAM gene expression in human multinodular goiter (MNG). MATERIAL AND METHODS DNA and RNA samples were obtained from hyperplastic thyroid glands of 60 patients (54 females) with benign MNG. DREAM mutations were evaluated by PCR and direct automatic sequencing, whereas relative quantification of mRNA was performed by real-time PCR. Over- and under-expression were defined as a 2-fold increase and decrease in comparison to normal thyroid tissue, respectively. RQ M (relative quantification mean); SD (standard deviation). RESULTS DREAM expression was detected in all nodules evaluated. DREAM mRNA was overexpressed in 31.7% of MNG (RQ M=6.26; SD=5.08), whereas 53.3% and 15% had either normal (RQ M=1.16; SD=0.46) or underexpression (RQ M=0.30; SD=0.10), respectively. Regarding DREAM mutations analysis, only previously described intronic polymorphisms were observed. CONCLUSIONS We report DREAM gene expression in the hyperplastic thyroid gland of MNG patients. However, DREAM expression did not vary significantly, and was somewhat underexpressed in most patients, suggesting that DREAM upregulation does not significantly affect nodular development in human goiter.