Is there are relationship between polymorphisms TSHRgene frequencies and genetic ancestry markers in patients with PrimaryCongenital Hypothyroidism?

A defective thyroid-stimulating hormone receptor (TSHR) gene is one of the main known genetic factors leading to congenital hypothyroidism (CH). However, the relationship between TSHR genotypes and phenotype and the underlying reason for the broad spectrum of phenotypes in the patients carrying TSHR gene defects have not yet been clearly established. This study aimed to investigate the genetics of patients with CH to identify TSHR defects and to explore the specific extrathyroidal defects and other phenotypic features in these patients to establish a genotype-phenotype correlation. Consanguineous families with primary CH and a history of non-autoimmune acquired hypothyroidism were included in this study. The causative variants in the TSHR gene were identified using exome sequencing. Multiple in silico analysis tools were employed to interpret the variants. Five TSHR variants including two novel variants were identified in patients with thyroid dysgenesis from five families. Some patients presented inter- and intra-familial variable expression and different ages of onset. The data suggest the possibility that the clinical phenotype of patients with CH caused by TSHR variants can be influenced by the coexistence of other gene defects. This study investigated the variants of the TSHR gene contributing to CH for the first time in Iran. Our study on multiplex consanguineous families could help provide further evidence for the elucidation of the oligogenic inheritance in CH, possibly leading to variable expressivity in patients with CH. These data could have implications for genetic diagnosis and counseling to identify deleterious variants for possible diagnostics, clinical management, and preventive aims.

Objective To explore the characterization of thyroid stimulating hormone receptor(TSHR) gene mutational spectrum in children with hyperthyroidism from Guangzhou. Methods Ninety children were diagnosed with hyperthyroidism from July 2009 to July 2014 in our institute. Their median age at diagnosis was(7.5±3.4) years, and there were 28 males and 62 females. Mutational analysis were performed by performing polymerase chain reaction(PCR) and DNA direct sequencing of exon 10 of TSHR gene. TSHR gene mutations from 50 unrelated healthy children were served as controls. The correlation between TSHR gene and hyperthyroidism in children was explored. Results A total of 3 mutations were identified in ninety children who were diagnosed with hyperthyroidism, one synonymous mutations(p.V614V), and two missense mutations(p.R707W and p. D727E). Mutation of p. V614V do not change amino acid and do not influence the structure and function of TSHR, no pathogenicity. p. R707W is a SNP associated with human cancers. The frequency of C allele of the D727E in children with hyperthyroidism was 86.7%, while 55.0% in the controls, significant different between the children with hyperthyroidism and the controls(P<0.01). In this study, a very high association between the D727E SNP and hyperthyroidism(OR=18.86, P<0.01) was found. Conclusion Three different mutations of TSHR gene exon 10 were identified in 90 children with hyperthyroidism, (c.1842A>G, p.V614V、c.2119C>T, p.R707W、c.2181G>C, p.D727E), there were association between p. D727E and hyperthyroidism, nor p. V614V and p. R707W. Finally, p. D727E may be correlated with hyperthyroidism in children. Key words: Hyperthyroidism; Children; Thyroid stimulating hormone receptor(TSHR); Gene; Mutation

To explore the association between the thyroid stimulating hormone receptor (TSHR) gene methylation and human papillary thyroid cancer (PTC), as well as PTC related clinicopathological indicators. We searched PubMed, Embase, Medline, and Web of Science databases through computer for articles published in English on association between methylation of TSHR gene and PTC. Articles published in Chinese were searched in China National Knowledge Infrastructure (CNKI), WanFang, China Biology Medicine (CBM) disc, and WeiPu databases. Database search took place in the 4th week of October. Totally 914 samples from 14 case-control studies were included in our meta-analysis. The methylation rate of TSHR gene in PTC group was significantly greater than that in control group (OR = 6.45, 95% CI 3.03, 13.71, P < 0.001). The subgroup analysis results showed the incidence of TSHR gene methylation was higher in autologous controls (OR = 16.39, 95% CI 8.83, 30.42, P < 0.001), Asian races (OR = 8.26, 95% CI 3.54, 19.23, P < 0.001), and Chinese (OR = 11.40, 95% CI 5.56, 23.39, P < 0.001). Hierarchical analysis of PTC related clinicopathological indicators showed that TSHR gene methylation rate are higher in PTC patients over 45 years (OR = 1.65, 95% CI 1.07, 2.55, P < 0.05) and lymph node metastasis (OR = 5.36, 95% CI 1.54, 18.67, P < 0.01). In addition, the occurrence of TSHR gene methylation had also been shown to be related to the clinical stage (OR = 0.23, 95% CI 0.07, 0.70, P < 0.05) and size (OR = 0.19, 95% CI 0.11, 0.32, P < 0.01) of tumors. The result of sensitivity analysis showed the combined results of the studies included in the meta-analysis were fairly stable. Begg's and Egger's tests also suggested that there was no significance publication bias (P > 0.1). The rate of TSHR gene methylation is higher in PTC and it may be associated with the pathogenesis of human PTC, suggesting that TSHR gene may be a candidate marker for PTC diagnosis. In addition, the occurrence of TSHR gene methylation in PTC patients is closely related to age, lymph node metastasis, clinical stage, and tumor size, suggesting that TSHR gene may be used as an index to judge the severity of PTC.

The molecular biology of follicular cell growth in thyroid nodules is still poorly understood. Because gain-of-function (activating) mutations of the thyroid-stimulating hormone receptor (TShR) and/or Gs alpha genes may confer TSh-independent growth advantage to neoplastic thyroid cells, we searched for somatic mutations of these genes in a series of hyperfunctioning and nonfunctioning follicular thyroid adenomas specifically selected for their homogeneous gross anatomy (single nodule in an otherwise normal thyroid gland). TShR gene mutations were identified by direct sequencing of exons 9 and 10 of the TShR gene in genomic DNA obtained from surgical specimens. Codons 201 and 227 of the Gs alpha gene were also analyzed. At histology, all hyperfunctioning nodules and 13 of 15 nonfunctioning nodules were diagnosed as follicular adenomas. Two nonfunctioning thyroid nodules, although showing a prevalent microfollicular pattern of growth, had histological features indicating malignant transformation (a minimally invasive follicular carcinoma and a focal papillary carcinoma). Activating mutations of the TShR gene were found in 12 of 15 hyperfunctioning follicular thyroid adenomas. In one hyperfunctioning adenoma, which was negative for TShR mutations, a mutation in codon 227 of the Gs alpha gene was identified. At variance with hyperfunctioning thyroid adenomas, no mutation of the TShR or Gs alpha genes was detected in nonfunctioning thyroid nodules. In conclusion, our findings clearly define a different molecular pathogenetic mechanism in hyperfunctioning and nonfunctioning follicular thyroid adenomas. Activation of the cAMP cascade, which leads to proliferation but maintains differentiation of follicular thyroid cells, typically occurs in hyperfunctioning thyroid adenomas. Oncogenes other than the TShR and Gs alpha genes are probably involved in nonfunctioning follicular adenomas.

Five patients, four brothers and their paternal aunt, presented with a history of overt hyperthyroidism and goiter. Hyperthyroidism in this family was remarkable for its poor response to carbimazole (30-50 mg/d). The thyroid ultrasound showed a diffusely enlarged gland in all the affected members, and thyroid stimulating antibodies (TSAB) were negative. Screening for germline mutations in thyroid stimulating hormone (TSH) receptor (TSHR) gene was performed by direct sequencing of genomic DNA extracted from peripheral blood leukocytes of all family members. The sequence analysis of all TSHR gene exons and intron borders revealed two genomic variants. The first was a single nucleotide polymorphism (SNP) within exon seven (Asn187Asn), whereas the other was located in intron seven (IVS7+68TG). All affected members, two asymptomatic brothers with sub-clinical hyperthyroidism, and their father were heterozygous for those two genomic variants. Anti-thyroid drug treatment for several months successfully relieved symptoms in one subject, whereas the remaining patients required total thyroidectomy to control their disease. This is the first Jordanian family with familial non-autoimmune hyperthyroidism, with mutations affecting the TSHR gene.

Congenital Hypothyroidism affects between 1:3000 and 1:4000 newborn infants in iodine-sufficient regions. Some studies have shown that mutations and polymorphisms in the TSH receptor gene are responsible for this disease. In the present study, mutations of exon 10 of the TSH receptor gene were investigated in Congenital Hypothyroidism patients. In the present study a sample of 90 Brazilian patients with primary congenital hypothyroidism was analyzed. Genomic DNA was isolated from peripheric blood samples. Exon 10 of the TSH receptor gene was amplified by PCR, and amplicons were automatically sequenced. Three nucleotide alterations were identified: c.1377G>A (A459A), c.1935G>A (L645L), and c.2181C>G (D727E). A459A polymorphism was also described previously in patients with thyroid cancer. The nucleotide alteration L645L was found in a single patient. This is the first time the L645L mutation has been described. D727E polymorphism showed high frequency (allele frequency 10%) in present study when compared to others reports.

A germline mutation of the thyroid-stimulating hormone receptor (TSHR) gene has been reported to be associated with thyrotoxicosis and mitral valve prolapse syndrome (MVPS) in a Chinese family. The role of TSHR genetic variants in MVPS has not been well studied. This study investigated the possible relationship between the polymorphisms of codon 727 and 52 of the TSHR gene and MVPS among the Chinese population in Taiwan. We studied 100 patients with MVPS diagnosed by echocardiography and 100 ageand sex-matched normal control subjects. The polymorphisms of codon 727 and 52 of the TSHR gene were identified by polymerase chain reaction-based restriction analysis. There was no significant difference in either the genotype distribution or allelic frequencies between MVPS cases and controls for either TSHR gene D727E polymorphism ( = 0.51 and 0.45, respectively ) or P52T polymorphism (P = 0.60 and 0.31, respectively). The MVPS patients were divided into 2 subgroups: those with Graves' disease, and those without the thyroid disorder, and there were no statistical differences from the controls for both the TSHR gene D727E and P52T polymorphisms. Further categorization of the MVPS patients into mild and severe subgroups also revealed no statistical difference from controls for either the TSHR gene D727E or P52T polymorphisms. These findings suggest that the codon 727 and 52 polymorphisms of the TSHR gene are not the suitable genetic markers of MVPS in Taiwan Chinese.

The aim of the study was to investigate whether the mutations of the GNAS gene and thyroid-stimulating hormone receptor (TSHR) gene were a potential molecular biological mechanism for subclinical toxic multinodular goiter (sTMG) and to evaluate the association of these mutations with the clinicopathological features of these disorders.Forty-four patients with sTMG and 20 controls(multinodular goiter) from Heilongjiang province of China who underwent subtotal thyroidectomy were recruited. Genes' mutations were analyzed by direct DNA sequencing of the polymerase chain reaction-amplified the parts of exons. In sTMG group, three mutations at GNAS gene were identified in seven patients (15.9%). Six mutations at TSHR gene were identified in 14 patients (31.8%). Mutation positivity of TSHR gene had statistically significant by comparison of two groups. In sTMG group, the mutation positivity of patients with serum TSH level below 0.1 microU/ml and above 0.01 microU/ml is obviously different (P < 0.05) at TSHR gene. However, these statistically significant differences were both not being seen at GNAS gene, and patients with nodules before universal salt iodization (USI)and after USI (P > 0.05). Mutation positivity of TSHR gene has a relation with sTMG. It is more probable that serum TSH level play an important role in mutagenesis.

Objective To investigate the association of single nucleotide polymorphisms in the rs2268458 locus of thyroid-stimulating hormone receptor (TSHR) gene with Graves disease (GD) and Hashimoto's thyroiditis (HT). Methods Part of the cases diagnosed through the epidemiological investigation project about thyroid diseases of Cangzhou City in 2016 was selected as the case group. The case group was subdivided into GD group and HT group according to the diagnosis. At the same time, healthy people with similar gender and age to the case group were selected as the control group. All subjects were from the Han nationality and were not related to each other. The genotypes and alleles of the TSHR gene rs2268458 (C/T) of all subjects were detected by restriction endonuclease (RFLP). And based on genotyping analysis of patient risk (odds ratio, OR) and 95% confidence interval (95%CI). Results There were 87 cases in GD group [aged (43.17 ± 12.56) years old], including 64 females and 23 males. There were 31 cases in HT group [aged (44.41 ± 16.51) years old], including 26 females and 5 males. In the control group, there were 147 cases [age (40.26 ± 9.31) years old], including 80 females and 67 males. Hardy-Weinberg equilibrium test was performed on each group. The results showed that P > 0.05, suggesting that the study samples were representative of the population. The results of genetic analysis showed that in females, the C allele frequency of GD patients was significantly higher than that of the control group [χ2 = 4.632, 36.7% (47/128) vs 25.0% (40/160), P 0.05). In males, there was no statistically significant difference in genotype (CC, CT, TT) frequency distribution and allele between groups (P > 0.05). Conclusion Among women in Cangzhou, the single nucleotide polymorphism of rs2268458 in TSHR gene is associated with the susceptibility to GD, but not to HT, and C genotype increases the risk of GD by dominant inheritance. Key words: Receptors, thyrotropin; Graves disease; Thyroiditis; Polymorphism, single nucleotide; Alleles

Objective : This study was aimed to investigate the clinical and biochemical response to levothyroxine (L-T4) doses in hypothyroid patients in correlation with genetic variation in thyroid stimulating hormone receptor ( TSHR ) gene and thyroid hormone receptor ( THR α) gene. Design : This cross-sectional correlation study includes 228 patients with primary hypothyroidism who were using L-T4 replacement therapy. Thyroid function test was performed using standard techniques. Genotyping of rs939348 at THRα gene, and rs2268458 and rs2239610 at TSHR gene was performed using the polymerase chain reaction-based restriction fragment length polymorphism assay (PCR-RFLP). Patient history of illness, medication and compliance data was collected using patients’ medical files. Results: The THRα rs939348 polymorphism was associated with L-T4 replacement doses in hypothyroid patient and central obesity. No significant correlation between the examined SNPs on TSHR and L-T4 doses or the different clinical and biochemical parameters. Finally, L-T4 dose was associated with lower BMI, waist circumference, and TSH, and higher free T4 (fT4) among hypothyroid patients. Conclusions: Whereas the two tested TSHR polymorphisms were not associated with the dose of T4, the THRα rs939348 polymorphism was associated with L-T4 dose and central obesity among hypothyroid patients. T4 dose is also associated with multiple beneficial effects among hypothyroid patients.

Toxic adenoma and toxic multinodular goiter (TMNG) are common causes of hyperthyroidism in iodine-deficient regions, but they are relatively rare in iodine-sufficient regions, including Japan. Constitutive activating mutations of the thyroid stimulating hormone receptor (TSHR) gene and adenylate cyclase-stimulating G α protein (GNAS) gene are frequent in these thyrotoxic disorders. Here we report two cases of rare TSHR gene mutations in Japanese thyrotoxicosis patients. In Case 1, we observed multiple toxic nodules with thyrotoxicosis, and in Case 2, we detected a solitary toxic nodule in an 8-year-old girl. In both cases, ultrasonography showed thyroid nodules and scintigraphy revealed increased uptake. Total thyroidectomy was performed for Case 1 and a hemi-thyroidectomy was performed for Case 2. Genetic analysis of the resected tissues revealed an I568F mutation in Case 1 and a S281I mutation in the TSHR gene in Case 2. The I568F mutation was located in the second extracellular loop, and the S281I mutation was located in the N-terminal extracellular domain of the TSH receptor. In Case 1, the mutation was restricted to the largest nodule, and was not detected in other functioning nodules or non-nodule thyroid tissue. Bi-allelic expression of the TSHR gene was confirmed by reverse transcription-polymerase chain reaction in both tumors. Both the I568F and S281I mutations were studied previously in vitro, and were revealed to cause basal activation of the protein kinase A pathway. Case 1 represents the second reported case of an I568F mutation and Case 2 represents the third reported case of an S281I mutation.

We found the G132R heterozygous mutation of thyroid stimulating hormone receptor (TSHR) gene in a patient with recurrent hypokalemia. Because the patient had a medical history of hyperthyroidism, the mutation was suspected to be related to hyperthyroidism at first. Subsequently, the expression and function studies in vitro were conducted. Wide-type TSHR and mutant TSHR (mutTSHR) were constructed in the phage vector and pEGFP-C1 vector. After transfection, the samples were collected for detection of mRNA level, protein expression, cell activity and cAMP content. Compared with the wild-type TSHR, the mRNA level of the mutTSHR was not significantly different. But the protein expression, cell activity and cAMP content of the mutTSHR were significantly lower. So this indicated that the G132R mutation is a loss-of-function mutation. We identified the G132R monoallelic heterozygous mutation of TSHR gene in a patient with hyperthyroidism. Based on disease history of the patient, we speculated that the heterozygous mutation did not cause thyroid dysplasia or hypothyroidism for her. Our study enriched experiment content in vitro studies and clinical phenotype about the G132R mutation in TSHR gene.

Graves’ disease (GD) is a common thyroid disease, and Graves ophthalmopathy(GO) is the most common extra-thyroidal manifestation of GD. Genetic associations of the thyroid stimulating hormone receptor (TSHR) gene with GD and GO have been studied in different population groups for a long time. We aimed to obtain a more precise estimation of the effects of TSHR single nucleotide polymorphisms (SNPs) on GD/GO using a meta-analysis. Publications were searched on Pub Med and EMBASE up to December 30, 2015. Eight studies involving three SNPs (rs179247, rs12101255, and rs2268458), which included 4790 cases and 5350 controls, met the selection criteria. The pooled odds ratios (OR) and the 95% confidence intervals (CI) were estimated. SNPs rs179247 (dominant model [GG + GA vs. AA]: OR = 0.66, 95%CI: 0.61–0.73, P = 0.000, I2 = 0%) and rs12101255 (dominant model [TT + TC vs. CC]: OR = 1.67, 95%CI: 1.53–1.83, P = 0.000, I2 = 0%) were significantly associated with GD in all of the genetic models. TSHR rs12101255 and rs2268458 polymorphisms had no association between GO and GD (GD without GO). The results indicate that rs179247 and rs12101255 are likely to be genetic biomarkers for GD. Further studies with different population groups and larger sample sizes are needed to confirm the genetic associations of the TSHR gene with GD/GO.

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.

Congenital hypothyroidism affects 1/3000-4000 newborns. The causes of this group of disorders are still largely unknown. Although most cases are sporadic some families have several affected children and/or consanguineous parents, suggesting autosomal recessive inheritance. Furthermore, there is a murine strain (hyt) with congenital hypothyroidism and autosomal recessive inheritance, whose phenotype appears to be identical with the corresponding human disease. In the hyt mouse, the disease is caused by a mutation in the thyroid-stimulating hormone receptor (TSHR) gene, making this gene a likely candidate also for the human disease. The human TSHR gene was mapped on radiation hybrid panels and closely linked flanking markers D14S287 and D14S616 were identified. On the Genebridge 4 panel, D14S287 was found to be located 8.5 cR (corresponding to 2.3 cM) proximal to the TSHR gene, and D14S616 was found to be located 4.4 cR (1.2 cM) distal to the TSHR gene. These markers were analyzed in 23 families, most of them with two or more children affected by congenital hypothyroidism and some with appreciable consanguinity of the parents. Assuming homogeneity, the two-point lod score at theta = 0.1 was -4.8 for D14S287 and -5.8 for D14S616, and thus linkage to the TSHR gene was excluded. Even when the data were analyzed with allowance for heterogeneity, there was no evidence of linkage. Our conclusion is that if mutation of the TSHR gene causes familial congenital hypothyroidism in humans, it affects only a small proportion of the cases.