Graves' ophthalmopathy: low-dose dexamethasone reduces retinoic acid receptor-alpha gene expression in orbital fibroblasts

ABSTRACT Objective: Graves' ophthalmopathy (GO) is an autoimmune disease that leads to ocular proptosis caused by fat accumulation and inflammation, and the main treatment is corticosteroid therapy. Retinoid acid receptor-alpha (RARα) seems to be associated with inflammation and adipocyte differentiation. This study aimed to assess the effect of glucocorticoid treatment on orbital fibroblasts of GO patient treated or not with different glucocorticoid doses. Materials and methods: Orbital fibroblasts collected during orbital decompression of a female patient with moderately severe/severe GO were cultivated and treated with 10 nM and 100 nM dexamethasone (Dex). rRARα gene expression in the treated and untreated cells was then compared. Results: Fibroblast RARα expression was not affected by 100 nM Dex. On the other hand, RARα expression was 24% lower in cells treated with 10 nM Dex (p < 0.05). Conclusions: Orbital fibroblasts from a GO patient expressed the RARα gene, which was unaffected by higher, but decreased with lower doses of glucocorticoid.

Gene expression of estrogen receptor-alpha in orbital fibroblasts in Graves’ ophthalmopathy

Graves’ ophthalmopathy (GO) is one of the most severe clinical manifestations of Graves’ disease (GD), and its treatment might involve high-dose glucocorticoid therapy. The higher incidence of GO among females, and the reported association between polymorphisms of estrogen receptor (ER) and GD susceptibility have led us to question the role of estrogen and its receptor in GO pathogenesis. We, thus, assessed estrogen receptor-alpha (ERA) gene expression in cultures of orbital fibroblasts from a patient with GO before (controls) and after treatment with 10 nM and 100 nM dexamethasone (DEX). Orbital fibroblasts showed ERA gene expression. In the cells treated with 10 nM and 100 nM DEX, ERA gene expression was, respectively, 85% higher and 74% lower, than in the control group. We concluded that ERA gene expression is found in the orbital fibroblasts of patient with GO, which may be affected by glucocorticoids in a dose-related manner. Arch Endocrinol Metab. 2015;59(3):273-6.

Triiodothyronine modulates the expression of leptin and adiponectin in 3T3-L1 adipocytes / Triiodotironina modula a expressão de leptina e adiponectina em adipócitos 3T3-L1

Objective To study the effect of different doses of triiodothyronine on gene expression of the adipokines leptin and adiponectin, at different times, and to evaluate the difference in expression between the two adipokines in each group. Methods 3T3-L1 adipocytes were incubated with triiodothyronine at physiological dose (10nM) and supraphysiological doses (100nM or 1,000nM), or without triiodothyronine (control, C) for 0.5, 6, or 24 hours. Leptin and adiponectin mRNA was detected using real-time polymerase chain reaction (RT-PCR). One-way analyses of variance, Tukey’s test or Student’s t test, were used to analyze data, and significance level was set at 5%. Results Leptin levels decreased in the 1,000nM-dose group after 0.5 hour. Adiponectin levels dropped in the 10nM-dose group, but increased at the 100nM dose. After 6 hours, both genes were suppressed in all hormone concentrations. After 24 hours, leptin levels increased at 10, 100 and 1,000nM groups as compared to the control group; and adiponectin levels increased only in the 100nM group as compared to the control group. Conclusion These results demonstrated fast actions of triiodothyronine on the leptin and adiponectin expression, starting at 0.5 hour, at a dose of 1,000nM for leptin and 100nM for adiponectin. Triiodothyronine stimulated or inhibited the expression of adipokines in adipocytes at different times and doses which may be useful to assist in the treatment of obesity, assuming that leptin is increased and adiponectin is decreased, in obesity cases. .

Objetivo Examinar o efeito de diferentes doses de triiodotironina sobre a expressão gênica das adipocinas leptina e adiponectina, em diferentes períodos de tempo, além de avaliar a diferença de expressão entre as duas adipocinas em cada grupo. Métodos Adipócitos 3T3-L1 foram incubados com triiodotironina nas doses fisiológica (10nM) e suprafisiológicas (100nM ou 1.000nM), ou na ausência de triiodotironina (controle, C) durante 0,5, 6 ou 24 horas. O mRNA das adipocinas foi analisado em tempo real, utilizando a reação em cadeia de polimerase. Para as análises dos dados, foi utilizada a análise de variância, complementada com o teste de Tukey, ou o teste t de Student com 5% de significância. Resultados Os níveis de leptina diminuíram no grupo com dose de 1.000nM em 0,5 hora. A adiponectina também diminuiu no grupo com dose de 10nM, porém se elevou com a dose de 100nM. Após 6 horas, ambos os genes foram suprimidos em todas concentrações de hormônio. Em 24 horas, os níveis de leptina foram elevados em 10, 100 e 1.000nM, em relação ao grupo controle. No que concerne à adiponectina, observou-se aumento apenas no grupo cuja dose foi de 100nM, em comparação ao controle. Conclusão Foram demonstradas ações rápidas da triiodotironina sobre a expressão da leptina e da adiponectina, iniciando em 0,5 hora na dose de 1.000nM, para a primeira, e na dose de 100nM, para a segunda. A triiodotironina estimulou ou inibiu a expressão de adipocinas em adipócitos em diferentes tempos e doses, o que pode auxiliar no tratamento da obesidade, levando em consideração que, nesta, a leptina está aumentada e adiponectina, diminuída. .

Short-term effects of triiodothyronine on thyroid hormone receptor alpha by PI3K pathway in adipocytes, 3T3-L1 / Efeitos rápidos da triiodotironina no receptor de hormônio tireoidiano alfa pela via PI3K em adipócitos, 3T3-L1

Objective The present study aimed to examine the effects of thyroid hormone (TH), more precisely triiodothyronine (T3), on the modulation of TH receptor alpha (TRα) mRNA expression and the involvement of the phosphatidyl inositol 3 kinase (PI3K) signaling pathway in adipocytes, 3T3-L1, cell culture. Materials and methods: It was examined the involvement of PI3K pathway in mediating T3 effects by treating 3T3-L1 adipocytes with physiological (P=10nM) or supraphysiological (SI =100 nM) T3 doses during one hour (short time), in the absence or the presence of PI3K inhibitor (LY294002). The absence of any treatment was considered the control group (C). RT-qPCR was used for mRNA expression analyzes. For data analyzes ANOVA complemented with Tukey’s test was used at 5% significance level. Results T3 increased TRα mRNA expression in P (1.91±0.13, p<0.001), SI (2.14±0.44, p<0.001) compared to C group (1±0.08). This increase was completely abrogated by LY294002 in P (0.53±0.03, p<0.001) and SI (0.31±0.03, p<0.001). To examine whether TRα is directly induced by T3, we used the translation inhibitor cycloheximide (CHX). The presence of CHX completely abrogated levels TRα mRNA in P (1.15±0.05, p>0.001) and SI (0.99±0.15, p>0.001), induced by T3. Conclusion These results demonstrate that the activation of the PI3K signaling pathway has a role in T3-mediated indirect TRα gene expression in 3T3-L1 adipocytes. .

Objetivo O objetivo do presente estudo foi analisar os efeitos do hormônio tireoidiano (HT), triiodotironina (T3), na modulação da expressão de mRNA do receptor alfa (TRα) de HT e o envolvimento da via de sinalização da via fosfatidilinositol 3-quinase (PI3K) em adipócitos, 3T3-L1. Materiais e métodos: Foi examinado o envolvimento da via PI3K nos efeitos do T3 nos tratamentos de adipócitos, 3T3-L1, nas doses fisiológica (P=10nM) ou suprafisiológica (SI =100 nM) durante uma hora (tempo curto), na ausência ou na presença do inibidor da PI3K (LY294002). A ausência de qualquer tratamento foi considerada o grupo controle (C). RT-qPCR foi utilizado para analisar a expressão do mRNA. Para as análises dos dados, utilizou-se ANOVA complementada com o teste de Tukey a 5% de significância. Resultados O T3 aumentou a expressão de mRNA de TRα em P (1,91±0,13, p<0,001) e SI (2,14±0,44, p<0,001) em comparação com o grupo C (1±0,08). Esse aumento foi completamente abolido por LY294002 em P (0,53±0,03, p<0,001) e SI (0,31±0,03, p<0,001). Para examinar se a expressão de TRα foi diretamente induzida pelo T3, utilizou-se o inibidor de tradução, ciclohexamida (CHX). A presença de CHX reduziu os níveis de mRNA de TRα em P (1,15±0,05, p>0,001) e SI (0,99±0,15, p>0,001), induzidos pelo T3. Conclusão Esses resultados demonstram que a ativação da via de sinalização de PI3K tem um papel importante na expressão do gene TRα mediada indiretamente pelo T3, em adipócitos 3T3-L1. .

Modulation of thyroid hormone receptors, TRα and TRβ, by using different doses of triiodothyronine (T3) at different times / Modulação dos receptores de hormônio tireoidiano, TRα e TRβ, utilizando diferentes doses de triiodotironina (T3) em diferentes tempos

OBJECTIVE: To examine the effect of different doses of triiodothyronine (T3) on mRNA levels of thyroid hormone receptors, TRα and TRβ, at different times. MATERIALS AND METHODS: 3T3-L1 adipocytes were incubated with T3 (physiological dose: F; supraphysiological doses: SI or SII), or without T3 (control, C) for 0.5, 1, 6, or 24h. TRα and TRβ mRNA was detected using real-time polymerase chain reaction. RESULTS: F increased TRβ mRNA levels at 0.5h. After 1h, TRα levels increased with F and SI and TRβ levels decreased with SII compared with C, F, and SI. After 6h, both genes were suppressed at all concentrations. In 24h, TRα and TRβ levels were similar to those of C group. CONCLUSIONS: T3 action with F began at 1h for TRα and at 0.5h for TRβ. These results suggest the importance of knowing the times and doses that activate T3 receptors in adipocytes.

OBJETIVO: Examinar o efeito de diferentes doses de triiodotironina (T3) sobre a expressão gênica dos receptores TRα e TRβ em diferentes tempos. MATERIAIS E MÉTODOS: Adipócitos, 3T3-L1, foram incubados com T3 nas doses fisiológica (F, 10nM) e suprafisiológicas (SI, 100nM ou SII, 1000nM) ou veículo (controle, C) durante 0,5, 1, 6 ou 24h. mRNA dos TRs foram detectados utilizando PCR em tempo real. RESULTADOS: Níveis de TRβ aumentaram em F em 0,5h. Após 1h, níveis de TRα aumentaram em F e SI comparado ao C, enquanto TRβ diminuiu no SII comparado com C, F, e SI. Após 6h, ambos os genes foram suprimidos em todas concentrações. Em 24h, níveis de TRα e TRβ retornaram aos do C. CONCLUSÕES: Ação do T3 em F iniciou-se em 1h para TRα e 0,5h para TRβ. Esses resultados são importantes para determinar tempo inicial e dose de T3 em que os receptores de HT são ativados em adipócitos.

Perfil dos hormônios da tireoide em pacientes com câncer de mama em estado de menopausa / Thyroid hormone profile in breast cancer patients in postmenopause

OBJETIVO: Verificar o perfil dos hormônios tireóideos (HTs) em pacientes pós-menopausa portadoras de carcinoma de mama (CaM). SUJEITOS E MÉTODOS: Participaram 12 pacientes com CaM em estádio I ou II sem intervenções que pudessem interferir na progressão tumoral e um grupo controle com 18 pacientes em pós-menopausa sem CaM. Foram dosados os níveis séricos de anticorpo antitiroperoxidase (TPOAB), hormônio estimulante da tireoide (TSH), tiroxina livre (T4L), estradiol (E2), hormônio folículo estimulante (FSH) e hormônio luteinizante (LH) antes e após a cirurgia, e realizada a imunoistoquímica dos receptores de estrógeno (ER) e progesterona (PR). RESULTADOS: Quatro pacientes com CaM apresentaram alterações do perfil hormonal tireoidiano: dois hipertireoidismo, um hipotireoidismo e positividade TPO-AB, todas com ER e PR positivos. Os níveis de TSH dessas pacientes não foram diferentes dos níveis encontrados no grupo controle (1,89 ± 1,56 vs. 2,86 ± 3,12 mUI/mL), porém os níveis de T4L nas pacientes com CaM foram estatisticamente maiores que o controle (1,83 ± 0,57 vs. 1,10 ± 0,20 ng/dL). CONCLUSÃO: Esses resultados reforçam a necessidade de avaliação do status tireoidiano em pacientes com CaM, uma vez que, na ausência de E2, mudanças clínicas nos HTs podem atuar em vias controladas pelo E2.

OBJECTIVE: The aim of this study was to determine thyroid hormone (TH) profile in postmenopausal patients with breast cancer (BC). SUBJECTS AND METHODS: 12 CaM patients stages I or II, without interventions that could interfere with tumor progression were selected, as well as and a control group with 18 postmenopausal women without CaM. We measured serum anti-thyroperoxidase antibody (TPOAB), thyroid-stimulating hormone (TSH), free thyroxine (T4L), estradiol (E2), follicle-stimulating hormone (FSH), and luteinizing hormone (LH), before and after surgery, besides immunohistochemistry for estrogen (ER) and progesterone (PR) receptors. RESULTS: Four patients with CaM showed changes in thyroid hormone profile: two had hyperthyroidism, one hypothyroidism, and one was positive for TPO-AB. All of them positive for ER and PR. TSH levels in breast cancer patients were not different from levels found in the control group (1.89 ± 1.56 vs. 2.86 ± 3.12 mIU/mL), but the levels of T4L in patients with CaM were statistically higher than those of the control group (1.83 ± 0.57 vs. 1.10 ± 0.20 ng/dL). CONCLUSION: These results reinforce the need for assessment of thyroid status in CaM patients, since in the absence of E2, changes in clinical HTs can act in E2-controlled processes.