Effects of Triiodothyronine on Human Osteoblast-Like Cells: Novel Insights From a Global Transcriptome Analysis.

Background: Thyroid hormones play a significant role in bone development and maintenance, with triiodothyronine (T3) particularly being an important modulator of osteoblast differentiation, proliferation, and maintenance. However, details of the biological processes (BPs) and molecular pathways affected by T3 in osteoblasts remain unclear. Methods: To address this issue, primary cultures of human adipose-derived mesenchymal stem cells were subjected to our previously established osteoinduction protocol, and the resultant osteoblast-like cells were treated with 1 nm or 10 nm T3 for 72 h. RNA sequencing (RNA-Seq) was performed using the Illumina platform, and differentially expressed genes (DEGs) were identified from the raw data using Kallisto and DESeq2. Enrichment analysis of DEGs was performed against the Gene Ontology Consortium database for BP terms using the R package clusterProfiler and protein network analysis by STRING. Results: Approximately 16,300 genes were analyzed by RNA-Seq, with 343 DEGs regulated in the 1 nm T3 group and 467 upregulated in the 10 nm T3 group. Several independent BP terms related to bone metabolism were significantly enriched, with a number of genes shared among them (FGFR2, WNT5A, WNT3, ROR2, VEGFA, FBLN1, S1PR1, PRKCZ, TGFB3, and OSR1 for 1nM T3; and FZD1, SMAD6, NOG, NEO1, and ENG for 10 nm T3). An osteoblast-related search in the literature regarding this set of genes suggests that both T3 doses are unfavorable for osteoblast development, mainly hindering BMP and canonical and non-canonical WNT signaling. Conclusions: Therefore, this study provides new directions toward the elucidation of the mechanisms of T3 action on osteoblast metabolism, with potential future implications for the treatment of endocrine-related bone pathologies.

Historical evolution of spheroids and organoids, and possibilities of use in life sciences and medicine.

BACKGROUND: An impressive percentage of biomedical advances were achieved through animal research and cell culture investigations. For drug testing and disease researches, both animal models and preclinical trials with cell cultures are extremely important, but present some limitations, such as ethical concern and inability of representing complex tissues and organs. 3D cell cultures arise providing a more realistic in vitro representation of tissues and organs. Environment and cell type in 3D cultures can represent in vivo conditions and thus provide accurate data on cell-to-cell interactions, and cultivation techniques are based on a scaffold, usually hydrogel or another polymeric material, or without scaffold, such as suspended microplates, magnetic levitation, and microplates for spheroids with ultra-low fixation coating. PURPOSE AND SCOPE: This review aims at presenting an updated summary of the most common 3D cell culture models available, as well as a historical background of their establishment and possible applications. SUMMARY: Even though 3D culturing is incapable of replacing other current research types, they will continue to substitute some unnecessary animal experimentation, as well as complement monolayer cultures. CONCLUSION: In this aspect, 3D culture emerges as a valuable alternative to the investigation of functional, biochemical, and molecular aspects of human pathologies.

Pitfalls and challenges of the purinergic signaling cascade in obesity.

Obesity is a worldwide health problem which have reached pandemic proportions, now also including low and middle-income countries. Excessive or abnormal fat deposition in the abdomen especially in the visceral compartment is tightly associated with a high metabolic risk for arterial hypertension, type II diabetes, cardiovascular diseases, musculoskeletal disorders (especially articular degeneration) and some cancers. Contrariwise, accumulation of fat in the subcutaneous compartment has been associated with a neutral metabolic impact, favoring a lower risk of insulin resistance. Obesity results more often from an avoidable imbalance between food consumption and energy expenditure. There are several recommended strategies for dealing with obesity, including pharmacological therapies, but their success remains incomplete and may not compensate the associated adverse effects. Purinergic signaling operated by ATP and its metabolite, adenosine, has attracted increasing attention in obesity. The extracellular levels of purines often reflect the energy status of a given cell population. Adenine nucleotides and nucleosides fine tuning control adipogenesis and mature adipocytes function via the activation of P2 and P1 purinoceptors, respectively. These features make the purinergic signaling cascade a putative target for therapeutic intervention in obesity and related metabolic syndromes. There are, however, gaps in our knowledge regarding the role of purines in adipocyte precursors differentiation and mature adipocytes functions, as well as their impact among distinct adipose tissue deposits (e.g. white vs. brown, visceral vs. subcutaneous), which warrants further investigations before translation to clinical trials can be made.

Irisin modulates genes associated with severe coronavirus disease (COVID-19) outcome in human subcutaneous adipocytes cell culture.

Obesity patients are more susceptible to develop COVID-19 severe outcome due to the role of angiotensin-converting enzyme 2 (ACE2) in the viral infection. ACE2 is regulated in the human cells by different genes associated with increased (TLR3, HAT1, HDAC2, KDM5B, SIRT1, RAB1A, FURIN and ADAM10) or decreased (TRIB3) virus replication. RNA-seq data revealed 14857 genes expressed in human subcutaneous adipocytes, including genes mentioned above. Irisin treatment increased by 3-fold the levels of TRIB3 transcript and decreased the levels of other genes. The decrease in FURIN and ADAM10 expression enriched diverse biological processes, including extracellular structure organization. Our results, in human subcutaneous adipocytes cell culture, indicate a positive effect of irisin on the expression of multiple genes related to viral infection by SARS-CoV-2; furthermore, translatable for other tissues and organs targeted by the novel coronavirus and present, thus, promising approaches for the treatment of COVID-19 infection as therapeutic strategy to decrease ACE2 regulatory genes.

The roles of triiodothyronine and irisin in improving the lipid profile and directing the browning of human adipose subcutaneous cells.

Triiodothyronine (T3) and irisin (I) can modulate metabolic status, increase heat production, and promote differentiation of white adipose tissue (WAT) into brown adipose tissue (BAT). Herein, human subcutaneous white adipocytes were treated with 10 nM T3 or 20 nM I for 24 h to evaluate intracellular lipid accumulation, triglyceride, and glycerol levels, oxidative stress, DNA damage, and protein levels of uncoupling protein 1 (UCP1), adiponectin, leptin, peroxisome proliferator-activated receptor gamma (PPARγ), and fibronectin type III domain-containing protein 5 (FNDC5). T3 and irisin improved UCP1 production, lipid profile, oxidative stress, and DNA damage. T3 elevated adiponectin and leptin levels with a concomitant decrease in PPARy and FNDC5 levels. However, irisin did not alter adipokine, PPARy, and FNDC5 levels. The results indicate that T3 may be used to increase leptin and adiponectin levels to improve insulin sensitivity, and irisin may be used to prevent obesity or maintain weight due to its impact on the lipid profile without altering adipokine levels.

Triiodothyronine activated extranuclear pathways upregulate adiponectin and leptin in murine adipocytes.

Adiponectin and leptin, important for metabolic regulation, are synthesized and secreted by adipose tissue and are influenced by triiodothyronine (T3) that activates the MAPK/ERK and integrin αVß3 pathways, modulating gene expression. Adipocytes were treated with T3 (10 nM), for 1 h, in the absence or presence of PD98059 (PD) and tetraiodothyroacetic acid (Tetrac), which are pathways inhibitors. The cells were incubated with Adipo Red/Oil Red O reagents, and intracellular lipid accumulation [glycerol and triacylglycerol (TAG)], MTT, 8-hydroxideoxyguanosine (8-OH-dG), and mRNA and protein expression were assessed. T3 increased leptin mRNA and protein expression, and, in contrast, there was a decrease in the Tetrac + T3 group. Adiponectin mRNA expression was not altered by T3, though it had increased its protein expression, which was terminated by inhibitors PD + T3 and Tetrac + T3. However, T3 did not alter PPARγ protein expression, lipid accumulation, TAG, glycerol, and DNA damage, but PD + T3 and Tetrac + T3 reduced these parameters. T3 activated the MAPK/ERK pathway on adipocytes to modulate the adiponectin protein expression and integrin αvß3 to alter the leptin gene expression.

The importance of estrogen for bone protection in experimental hyperthyroidism in human osteoblasts.

Triiodothyronine (T3) and estrogen (E2) play important roles in the bone remodeling process and signaling of receptor activator of the nuclear factor-kappa ß (RANKL) and osteoprotegerin (OPG) expressed by osteoblasts. However, little is known of the molecular action of these hormones in conditions of hyperthyroidism and associated E2 in human cells. AIMS: This study evaluated the effects of the physiological concentration of E2 (10 nM), alone or in association with physiological (1 nM) and supraphysiological (10 nM) concentrations of T3, on RANKL and OPG gene expression in human osteoblasts. MAIN METHODS: Alkaline phosphatase and osteocalcin assays were performed to verify the presence of mature osteoblasts. After mimicking the experimental hyperthyroidism in osteoblasts untreated or treated with E2, RANKL and OPG gene expression was analyzed by real-time PCR and protein expression by western Blot and ELISA. Alizarin Red staining analyzed the amount of bone matrix after hormonal treatments. KEY FINDINGS: E2 enhanced the gene expression of OPG when associated with 1 nM and 10 nM T3. E2 was able to restore the bone matrix after an initial decrease using 1 nM and 10 nM T3. The protective effect of E2 on the RANKL and OPG signaling pathway was demonstrated. E2 restored the bone matrix induced by experimental hyperthyroidism. SIGNIFICANCE: The data highlight the importance of E2 to maintain OPG expression and osteoblast activity against possible loss of bone mass, especially in conditions where T3 is in excess.

Disruptive Effect of Organotin on Thyroid Gland Function Might Contribute to Hypothyroidism.

A considerable increase in endocrine abnormalities has been reported over the last few decades worldwide. A growing exposure to endocrine-disrupting chemicals (EDCs) can be one of the causes of endocrine disorders in populations, and these disorders are not only restricted to the metabolic hormone system but can also cause abnormal functions. Thyroid hormone (TH) disruption is defined as an abnormal change in TH production, transport, function, or metabolism, which results in some degree of impairment in body homeostasis. Many EDCs, including organotin compounds (OTCs), are environmental contaminants that are commonly found in antifouling paints used on ships and in several other industrial procedures. OTCs are obesogenic and can disrupt TH metabolism; however, abnormalities in thyroid function resulting from OTC exposure are less well understood. OTCs, one of the most prevalent EDCs that are encountered on a daily basis, modulate the thyroid axis. In most toxicology studies, it has been reported that OTCs might contribute to hypothyroidism.

Adiponectin and Serine/Threonine Kinase Akt Modulation by Triiodothyronine and/or LY294002 in 3T3-L1 Adipocytes.

Adipose tissue (AT), an endocrine organ that modulates several physiological functions by synthesizing and releasing adipokines such as adiponectin, is a metabolic target of triiodothyronine (T3). T3 and adiponectin play important roles in controlling normal metabolic functions such as stimulation of fatty acid oxidation and increase in thermogenesis. The phosphatidylinositol 3-kinase (PI3K) pathway is important for the differentiation of preadipocytes into adipocytes and can be activated by T3 for the transcription of specific genes, such as adiponectin. We examined the role of PI3K in adiponectin modulation by T3 action in murine adipocytes (3T3-L1). The 3T3-L1 adipocytes were treated with 1000 nM T3 for 1 h in the presence or absence of 50 µM LY294002 (LY), a PI3K inhibitor. Then, we assessed the expression of adiponectin and the phosphorylated serine/threonine kinase Akt (pAkt), a PI3K signaling protein, in the adipocytes. Adiponectin and pAKT levels were higher in the T3-adipocyte cells, whereas in the LY group adiponectin was elevated and pAKT was decreased compared to the control (C). PI3K pathway inhibition for 1 h and posterior treatment with T3, in LY + T3, reduced the adiponectin level and increased pAKT levels compared to those in LY. T3 stimulated adiponectin levels by PI3K pathway activation and T3 can compensate alteration in the PI3K pathway, because with inhibition of the pathway it is able to maintain the basal levels of adiponectin and pAKT.

The importance of estrogen for bone protection in experimental hyperthyroidism in human osteoblasts

Triiodothyronine (T3) and estrogen (E2) play important roles in the bone remodeling process and signaling of receptor activator of the nuclear factor-kappa ß (RANKL) and osteoprotegerin (OPG) expressed by osteoblasts. However, little is known of the molecular action of these hormones in conditions of hyperthyroidism and associated E2 in human cells. AIMS: This study evaluated the effects of the physiological concentration of E2 (10?nM), alone or in association with physiological (1nM) and supraphysiological (10nM) concentrations of T3, on RANKL and OPG gene expression in human osteoblasts. MAIN METHODS: Alkaline phosphatase and osteocalcin assays were performed to verify the presence of mature osteoblasts. After mimicking the experimental hyperthyroidism in osteoblasts untreated or treated with E2, RANKL and OPG gene expression was analyzed by real-time PCR and protein expression by western Blot and ELISA. Alizarin Red staining analyzed the amount of bone matrix after hormonal treatments. KEY FINDINGS: E2 enhanced the gene expression of OPG when associated with 1nM and 10nMT3. E2 was able to restore the bone matrix after an initial decrease using 1nM and 10nMT3. The protective effect of E2 on the RANKL and OPG signaling pathway was demonstrated. E2 restored the bone matrix induced by experimental hyperthyroidism. SIGNIFICANCE: The data highlight the importance of E2 to maintain OPG expression and osteoblast activity against possible loss of bone mass, especially in conditions where T3 is in excess.

The importance of estrogen for bone protection in experimental hyperthyroidism in human osteoblasts

Triiodothyronine (T3) and estrogen (E2) play important roles in the bone remodeling process and signaling of receptor activator of the nuclear factor-kappa ß (RANKL) and osteoprotegerin (OPG) expressed by osteoblasts. However, little is known of the molecular action of these hormones in conditions of hyperthyroidism and associated E2 in human cells. AIMS: This study evaluated the effects of the physiological concentration of E2 (10?nM), alone or in association with physiological (1nM) and supraphysiological (10nM) concentrations of T3, on RANKL and OPG gene expression in human osteoblasts. MAIN METHODS: Alkaline phosphatase and osteocalcin assays were performed to verify the presence of mature osteoblasts. After mimicking the experimental hyperthyroidism in osteoblasts untreated or treated with E2, RANKL and OPG gene expression was analyzed by real-time PCR and protein expression by western Blot and ELISA. Alizarin Red staining analyzed the amount of bone matrix after hormonal treatments. KEY FINDINGS: E2 enhanced the gene expression of OPG when associated with 1nM and 10nMT3. E2 was able to restore the bone matrix after an initial decrease using 1nM and 10nMT3. The protective effect of E2 on the RANKL and OPG signaling pathway was demonstrated. E2 restored the bone matrix induced by experimental hyperthyroidism. SIGNIFICANCE: The data highlight the importance of E2 to maintain OPG expression and osteoblast activity against possible loss of bone mass, especially in conditions where T3 is in excess.

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

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.

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.

Triiodothyronine (T3) induces HIF1A and TGFA expression in MCF7 cells by activating PI3K.

High expression levels of hypoxia inducing factor 1 alpha are related to mammary carcinogenesis. In previous studies, we demonstrated that expression of transforming growth factor alpha increases upon treatment with triiodothyronine, but this expression does not occur in cellular models that do not express the estrogen receptor, or when cells are co-treated with the anti-estrogen, tamoxifen. The aim of this study was to determine the effect of the hormone triiodothyronine on the expression of the genes HIF1A and TGFA in the breast cancer cell line MCF7. The cell line was subjected to treatment with triiodothyronine at the supraphysiological dose of 10(-8)M for 10min, 30min, 1h, and 4h in the presence or absence of actinomycin D, the gene expression inhibitor, cycloheximide, the protein synthesis inhibitor, and LY294002, the phosphoinositide 3 kinase inhibitor. HIF1A and TGFA mRNA expression was analyzed by reverse transcription polymerase chain reaction. For data analysis, we used analysis of variance complemented by Tukey test and an adopted minimum of 5% significance. We found that HIF1A and TGFA expression increased in the presence of triiodothyronine at all times studied. HIF1A expression decreased in triiodothyronine-treated cells when gene transcription was also inhibited; however, TGFA expression decreased after 10 and 30min of treatment even when transcription was not inhibited. We found that activation of PI3K was necessary for triiodothyronine to modulate HIF1A and TGFA expression.

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.

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.

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.

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.

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.