Maria Izabel Chiamolera

Adult exposure to bisphenol A (BPA) in Wistar rats reduces sperm quality with disruption of the hypothalamic–pituitary–testicular axis

Reproductive physiology involves complex biological processes that can be disrupted by exposure to environmental contaminants. The effects of bisphenol A (BPA) on spermatogenesis and sperm quality is still unclear. The objective of this study was to investigate the reproductive toxicity of BPA at dosages considered to be safe (5 or 25mg BPA/kg/day). We assessed multiple sperm parameters, the relative expression of genes involved in the central regulation of the hypothalamic-pituitary-testicular axis, and the serum concentrations of testosterone, estradiol, LH and FSH. BPA exposure reduced sperm production, reserves and transit time. Significant damage to the acrosomes and the plasma membrane with reduced mitochondrial activity and increased levels of defective spermatozoa may have compromised sperm function and caused faster movement through the epididymis. BPA exposure reduced the serum concentrations of testosterone, LH and FSH and increased the concentration of estradiol. The relative gene expression revealed an increase in gonadotropin releasing hormone receptor (Gnrhr), luteinizing hormone beta (Lhb), follicle stimulating hormone beta (Fshb), estrogen receptor beta (Esr2) and androgen receptor (Ar) transcripts in the pituitary and a reduction in estrogen receptor alpha (Esr1) transcripts in the hypothalamus. In this study, we demonstrated for the first time that adult male exposure to BPA caused a reduction in sperm production and specific functional parameters. The corresponding pattern of gene expression is indicative of an attempt by the pituitary to reestablish normal levels of LH, FSH and testosterone serum concentrations. In conclusion, these data suggest that at dosages previously considered nontoxic to reproductive function, BPA compromises the spermatozoa and disrupts the hypothalamic-pituitary-gonadal axis, causing a state of hypogonadotropic hypogonadism.

Fundamentally Distinct Roles of Thyroid Hormone Receptor Isoforms in a Thyrotroph Cell Line Are due to Differential DNA Binding

Thyroid hormones have a profound influence on human development and disease. The hypothalamic-pituitary-thyroid axis involves finely tuned feedback mechanisms to maintain thyroid hormone (TH) levels. Despite the important role of TH-negative feedback in regulating this axis, the mechanism by which this occurs is not clearly defined. Previous in vivo studies suggest separate roles for the two thyroid hormone receptor isoforms, THRA and THRB, in this axis. We performed studies using a unique pituitary thyrotroph cell line (TαT1.1) to determine the relative roles of THRA and THRB in the regulation of Tshb. Using chromatin immunoprecipitation assays, we found that THRB, not THRA, bound to the Tshb promoter. By selectively depleting THRB, THRA, or both THRA and THRB in TαT1.1 cells, we found that simultaneous knockdown of both THRB and THRA abolished T(3)-mediated down-regulation of Tshb at concentrations as high as 100 nm T(3). In contrast, THRA knockdown alone had no effect on T(3)-negative regulation, whereas THRB knockdown alone abolished T(3)-mediated down-regulation of Tshb mRNA levels at 10 nm but not 100 nm T(3) concentrations. Interestingly, chromatin immunoprecipitation assays showed that THRA becomes enriched on the Tshb promoter after knockdown of THRB. Thus, a likely mechanism for the differential effects of THR isoforms on Tshb may be based on their differential DNA-binding affinity to the promoter.

Clinical and Molecular Analysis of Thyroid Hypoplasia: A Population-Based Approach in Southern Brazil

BACKGROUND Congenital hypothyroidism (CH) is mainly due to developmental abnormalities leading to thyroid dysgenesis (TD). TD encompasses very distinct morphologic subtypes of disease. This study examined and compared the phenotype in TD variants and searched for genetic alterations in sporadic thyroid hypoplasia (TH), the most misdiagnosed form of CH. This was a longitudinal study over a 14-year period (1990-2004). METHODS A continuous series of 353 children with TD was identified using thyroid function tests [thyroxine (T4) and TSH], scintigraphy, and ultrasound as diagnostic tools. Individual phenotypes were analyzed in 253 children with TD. Mutations in the most likely candidate genes were studied in 35 cases of TH. RESULTS The overall birth prevalence of permanent CH was 1:4795. Ectopy represented 37% of all cases of permanent primary CH, dyshormonogenesis 28%, agenesis 24%, hypoplasia 10%, and hemiagenesis 1%. The lowest screening T4 level and the highest TSH level were in the agenetic group, followed by TH. The TH group had an improvement in the thyroid function showing less-severe phenotype with aging. In the molecular analysis, one patient was identified with a mutation in the PAX8 gene (155G>C; R52P); four patients had a heterozygous G>C substitution in position -569; two patients showed a (234C>A; P52T) or (2181C>G; D727E) polymorphic variants of the TSH-R gene; and one patient presented a novel heterozygous nonsynonymous substitution, 293G>A; S98N, in the NKX2.5 gene. CONCLUSIONS The prevalence of CH was within the previously reported range of 1:3000-4000. Ectopy was the most common etiology. Clinical analysis revealed distinct hormonal patterns in TH subgroup when compared with other variants of TD, with genetic abnormalities identified only in few cases in the TSH-R, PAX8, and NKX2.5 genes.

Paralisia periódica hipocalêmica tirotóxica, uma urgência endócrina: revisão do quadro clínico e genético de 25 pacientes

Thyrotoxic hypokalemic periodic paralysis (THPP) is a medical emergency characterized by acute attacks of weakness, hypokalemia, and thyrotoxicosis that resolve with the treatment of hyperthyroidism. Attacks are transient, self-limited, associated with hypokalemia and resemble those of familial hypokalemic periodic paralysis (FHPP), an autosomal dominant neurological channelopathy. This study reviews the clinical features and genetic findings of THPP in 25 Brazilian patients. Most patients had weight loss, taquicardia, goiter, tremor, and ophthalmopathy. Most often attacks arose during the night and recovered spontaneously but some patients evolved to total quadriplegia, and few experienced cardiac arrhythmias. All patients had suppressed TSH and elevated T4 and most had positive anti-thyroid antibodies, indicating autoimmunity thyrotoxic etiology. Potassium was low in all patients during the crisis. Prophylactic potassium therapy has not been shown to prevent attacks; however it was useful for curbing the paralysis during the crisis. We identified the mutation R83H in the KCNE3 gene in one sporadic case, and M58V in the KCNE4 gene in one case with family history. Furthermore, we identified other genetic polymorphisms in the CACNA1S, SCN4A, KCNE1, KCNE2, KCNE1L, KCNJ2, KCNJ8 e KCNJ11 genes. We conclude that THPP is the most common treatable cause of acquired periodic paralysis; therefore, it must be included in the differential diagnosis of acute muscle weakness.

Comparison of practical methods for urinary glycosaminoglycans and serum hyaluronan with clinical activity scores in patients with Graves' ophthalmopathy

objective  Immunosuppressive treatment of Graves’ opthalmopathy (GO) should be restricted to patients with active eye disease, but assessing disease activity is difficult. Several methods to evaluate GO activity have been introduced, but none of them is satisfactory. Glycosaminoglycans (GAGs) are complex polysaccharides that participate on the pathogenesis of GO and attempts to correlate its local increase to urinary GAGs (uGAGs) or serum hyaluronan (sHA) have been made, but the available techniques are labourious, time‐consuming and difficult for routine use. The aim of the present study is to develop practical and simple methods for uGAGs and sHA and compare them to the activity and severity of thyroid‐associated ophthalmopathy.

Daily exposure to silver nanoparticles during prepubertal development decreases adult sperm and reproductive parameters

Abstract As silver nanoparticles (AgNPs) have antimicrobial properties and potentiate the activity of some antibiotics, they are broadly used in both medical and nonmedical applications. In this study, prepubertal male Wistar rats were orally treated with 15 or 30 µg/kg/day AgNPs from postnatal day 23 (PND23) to PND58 and sacrificed at PND102. The acrosome integrity, plasma membrane integrity, mitochondrial activity and morphological alterations of the sperm were analyzed. Sexual partner preference, sexual behavior and the serum concentrations of FSH, LH, testosterone and estradiol were also recorded. The results were evaluated following the appropriate statistical analyses, and differences among the groups were considered significant when p < 0.05. AgNPs reduced the acrosome and plasma membrane integrities, reduced the mitochondrial activity and increased the abnormalities of the sperm in both treatment groups. AgNP exposure also delayed the onset of puberty, although no changes in body growth were observed in either treatment group. The animals did not show changes in sexual behavior or serum hormone concentrations. This study shows for the first time that prepubertal exposure to AgNPs causes alterations in adult sperm parameters. Importantly, the sperm appeared to be more sensitive to the toxic effects of AgNPs and demonstrated adverse effects following exposure to lower doses. Consequently, the effects of AgNPs on sperm should be considered in order to establish safety limits for the use of these particles.

Ion channelopathies in endocrinology: recent genetic findings and pathophysiological insights

Ion channels serve diverse cellular functions, mainly in cell signal transduction. In endocrine cells, these channels play a major role in hormonal secretion, Ca(2+)-mediated cell signaling, transepithelial transport, cell motility and growth, volume regulation and cellular ionic content and acidification of lysosomal compartments. Ion channel dysfunction can cause endocrine disorders or endocrine-related manifestations, such as pseudohypoaldosteronism type 1, Liddle syndrome, Bartter syndrome, persistent hyperinsulinemic hypoglycemia of infancy, neonatal diabetes mellitus, cystic fibrosis, Dents disease, hypomagnesemia with secondary hypocalcemia, nephrogenic diabetes insipidus and, the most recently genetically identified channelopathy, thyrotoxic hypokalemic periodic paralysis. This review briefly recapitulates the membrane action potential in endocrine cells and offers a short overview of known endocrine channelopathies with focus on recent progress regarding the pathophysiological mechanisms and functional genetic defects.

Diagnóstico de metástases de carcinoma papilífero de tiróide através da dosagem de tiroglobulina no líquido obtido da lavagem da agulha utilizada na punção aspirativa

The widespread use of neck ultrasonography (US) during the follow-up of patients with papillary thyroid carcinoma (PTC) has led to the discovery of small cervical lymph nodes (LN). Although US has a high sensitivity for diagnosing LN, fine needle aspiration biopsy (FNA) and measurement of thyroglobulin in fine needle aspirates (FNA-Tg) have proven to be invaluable tools. The aim of this study is to determine the sensitivity of the combined use of neck US, FNA biopsy and FNA-Tg for diagnosis of cervical lymph nodes. We have studied 32 patients with 44 LN detected by US, 19 classified as inflammatory and 25 as suspicious. 15 of those 25 suspicious LN had high FNA-Tg (13 of the 15 had positive cytology and 2 indeterminate). All of these 15 LN (11 patients) were proven to be PTC metastasis by histopathology. All 19 inflammatory LN and those 10/25 suspicious LN, had cytology negative for malignancy and undetectable FNA-Tg. We conclude that fine needle aspiration biopsy and FNA-Tg combined with neck US are essential for detecting positive cervical lymph nodes due to its high sensitivity and specificity and it should be considered the standard for investigating locally recurrent disease in patients with PTC.

Perinatal exposure to glyphosate-based herbicide alters the thyrotrophic axis and causes thyroid hormone homeostasis imbalance in male rats

Glyphosate-based herbicides (GBHs) are widely used in agriculture. Recently, several animal and epidemiological studies have been conducted to understand the effects of these chemicals as an endocrine disruptor for the gonadal system. The aim of the present study was to determine whether GBHs could also disrupt the hypothalamic-pituitary-thyroid (HPT) axis. Female pregnant Wistar rats were exposed to a solution containing GBH Roundup®Transorb (Monsanto). The animals were divided into three groups (control, 5mg/kg/day or 50mg/kg/day) and exposed from gestation day 18 (GD18) to post-natal day 5 (PND5). Male offspring were euthanized at PND 90, and blood and tissues samples from the hypothalamus, pituitary, liver and heart were collected for hormonal evaluation (TSH-Thyroid stimulating hormone, T3-triiodothyronine and T4-thyroxine), metabolomic and mRNA analyses of genes related to thyroid hormone metabolism and function. The hormonal profiles showed decreased concentrations of TSH in the exposed groups, with no variation in the levels of the thyroid hormones (THs) T3 and T4 between the groups. Hypothalamus gene expression analysis of the exposed groups revealed a reduction in the expression of genes encoding deiodinases 2 (Dio2) and 3 (Dio3) and TH transporters Slco1c1 (former Oatp1c1) and Slc16a2 (former Mct8). In the pituitary, Dio2, thyroid hormone receptor genes (Thra1 and Thrb1), and Slc16a2 showed higher expression levels in the exposed groups than in the control group. Interestingly, Tshb gene expression did not show any difference in expression profile between the control and exposed groups. Liver Thra1 and Thrb1 showed increased mRNA expression in both GBH-exposed groups, and in the heart, Dio2, Mb, Myh6 (former Mhca) and Slc2a4 (former Glut4) showed higher mRNA expression in the exposed groups. Additionally, correlation analysis between gene expression and metabolomic data showed similar alterations as detected in hypothyroid rats. Perinatal exposure to GBH in male rats modified the HPT set point, with lower levels of TSH likely reflecting post-translational events. Several genes regulated by TH or involved in TH metabolism and transport presented varying degrees of gene expression alteration that were probably programmed during intrauterine exposure to GBHs and reflects in peripheral metabolism. In conclusion, the role of GBH exposure in HPT axis disruption should be considered in populations exposed to this herbicide.

The absence of mutations in homeobox candidate genes HOXA3, HOXB3, HOXD3 and PITX2 in familial and sporadic thyroid hemiagenesis

Abstract Background: The molecular mechanisms leading to the formation of the two thyroid symmetrical lobes, which are impaired in thyroid hemiagenesis (TH), are little known. Objective: The aim of this work was to search for mutations in thyroid developmental candidate genes HOXA3, HOXB3, HOXD3 and PITX2. Methods: Total DNA from peripheral blood was extracted and then the entire coding region of all these genes was amplified by polymerase chain reaction and direct sequencing. Results: Herein we describe familial cases of TH in two generations (proband and his father), in addition to other two sporadic cases. We have found polymorphisms in the HOXB3 (rs2229304), HOXD3 (rs34729309, rs1051929, c.543-199G>T and c.543-34G>A; and a new synonymous variant, NP_008829.3:p.314;C>G) and PITX2 (c.45+76C>T) genes, but no deleterious mutations. Conclusion: These results suggest the existence of other left-right thyroid asymmetry candidate genes in humans such as classical Mendelian mutation-causing disease, as well as other etiopathogenic mechanisms such as epigenetic modifications, especially for sporadic hemiagenesis.