RESUMENES

OR001
ANÁLISE DA EXPRESSÃO DOS GENES DE RESISTÊNCIA A MÚLTIPLAS- DROGAS MDR1 E MRP E A CORRELAÇÃO COM MUTAÇÕES NO GENE TP53 EM CARCINOMAS DA TIRÓIDE. 1Cerutti JM, 2Kimura E, 2Ebina KN e 1Maciel RMB. 1Laboratório de Endocrinologia Molecular, Disciplina de Endocrinologia, Universidade Federal de São Paulo-EPM, São Paulo, SP, Brasil. 2 Departamento de Histologia & Embriologia, ICB-USP, São Paulo, SP, Brasil.
      A agressividade e a refratariedade à terapêutica do carcinoma indiferenciado de tiróide (CIT) tem sido um dos grandes desafios no tratamento de cancer de tiróide. A análise de diferentes tipos de tumores revelou que a resistência ao tratamento tem sido associada a superexpressão da proteína codificada pelo gene de resistência à drogas-múltiplas (MDR1) que funciona como uma bomba de efluxo para vários agentes quimioterápicos. Outro mecanismo pelo qual as células podem adquirir resistência ao tratamento com drogas é através da superexpressão da proteína MRP. Estudos in vitro sugerem que o gene MDR1 é regulado negativamente pela proteína P53 normal e que a perda da função pode induzir à um aumento na transcricão do gene MDR1. Com o objetivo de melhor caracterizar o potencial biológico dos CIT, analisamos a expressão do gene de resistancia a drogas-múltiplas (MDR1) e correlacionamos estes achados com a presença de mutações no gene TP53 que é frequentemente mutado neste subtipo tumoral. Neste estudo foram analisados CIT (n=4), Carcinoma (Ca) papilífero (n=10), Ca folicular (n=4), tiróide Normal (n=3) e linhagens celulares derivadas de Ca Papilífero (NPA), Ca Folicular (WRO), CIT (ARO). A expressâo de MDR1 foi analisada por RT-PCR, e a pesquisa de mutação no gene TP53 por PCR-SSCP e sequenciamento de DNA. Ao contrário do esperado o gene MDR1 apresentou-se menos expresso nos CIT aonde foram identificadas mutações no gene TP53, quando comparado aos carcinoma diferenciados e controle normal. Também analisamos, utilizando imunohistoquímica (IHQ) a expressão da proteína MRP nas linhagens celulares ARO, NPA e WRO e verificamos uma correlação entre um aumento da expressão desta proteína na linhagem ARO quando comparada as linhagens NPA, WRO e controles normais. Estes achados mostram que a expressâo aumentada de MRP pode estar associadada à refratariedade terapêutica nos CIT, mas não se apresenta associada com o gene MDR1. E ainda, embora a literatura tenha sugerido que a integridade do gene TP53 seja responsável pela repressão do gene de MDR1,observamos que 2 dos CIT apresentavam mutaçao de p53 e expressâo diminuída de MDR1, não indicando uma associação TP53-MDR1

OR002
SEARCHING FOR IODINE DEFICIENCY DISORDERS (IDD) IN SCHOOLCHILDREN FROM BRAZIL: THE THYROMOBIL PROJECT
Rossi AC1, Pretell E2, Tomimori E1, Camargo RY1, Medeiros-Neto G1.
1Thyroid Unit, Division of Endocrinology, Univ. Sao Paulo Med School, Sao Paulo, Brazil, 2Universidad Peruana Cayetano Heredia, Lima, Peru
       The evaluation of IDD in Europe was recently conducted using an adapted vehicle "THYROMOBIL" equipped with an ultrasound apparatus, a computer unit, a freezer to store specimens, and an examining coach. The good results of this project in Europe made it possible to conduct similar study in Latin American and in Brazil. The target population was Brazilian schoolchildren (6-14 yrs. old) from randomly selected schools from villages and towns previously classified as "sentinels", at risk for IDD. The THYROMOBIL traveled 12,400 km in 9 states and visited 21 villages, examining 2,060 children. Results of ultrasonographic examination were available for 1,977 children. Thyroid pathological findings were present in 160 subjects (8%) including goiter (1.4%), nodules and cysts (0.75%), hypoechogeinity (6%) and 4 cases of hemiagenesis. Surprisingly the iodine concentration in domestic (household) samples of salt revealed that 47.4% of all specimens had more than 50 ppm Iodine and 10% of the salt had more than 100 ppm. Moreover the urinary excretion of iodine was considered to be very high in about 50% of all children. About 45% of all urinary specimens contained more than 500 µg I/liter urine and 3% of the specimens had more than 1,000 µg/L. It was concluded that the Brazil population is currently consuming an excessive amount of iodine per day and this may lead to thyroid dysfunction in many segments of the population, specially the elderly (hyperthyroidism) and post-menopausal women (hypothyroidism, autoimune disease).

OR003
ROLE OF THYROID HORMONE RECEPTOR HOMODIMER FORMATION IN THYROID HORMONE RESPONSES.
Velasco LFR, Caires FD, Neves FAR, Ribeiro RCJ. Molecular Pharmacology Laboratory, University of Brasília, DF, Brazil.
       Thyroid hormone (T3) receptors (TRs) bind to T3 response elements (TREs) as monomers and dimers. We previously found that residue L422 in the TRb1 is critical for homodimerization. This mutant is selectively impaired in dimer formation since it binds with normal affinity T3 and co-activators. The syndrome of T3 resistance (RTH) is commonly due to mutations in one allele of the TRb1 gene, whose protein product acts as dominant negative by interfering with the function of the normal TRb1, presumably through homodimerization. Here we tested the function of L422R on TREs to assess if disruption of dimerization impairs function. We also tested if G345R, TR mutant found in RTH that inhibits TRb1 function, can also inhibit L422R function on TREs. We employed gel shift assays to assess TR-DNA binding and transfection assays in U937 cells to test TR function on AGGTCA direct repeats (DR-4), the palindrome TREpal and the inverted palindrome F2. Our results showed that normal TRb1 and G345R bind well to all TREs, but L422R cannot homodimerize with itself, TRb1 or G345R on any TREs. In transfection assays, L422R has reduced activity on DR-4 and F2 but activates TREpal as well as TRb1. Interestingly, G345R, which repress TRb;1 function, could also repress L422R in transfection assays. Our data show that homodimer formation plays a role in TR function on DR-4 and F2, but not on TREpal suggesting TR homodimers do not function in all TREs. They also suggest that dominant negative activity of RTH mutants such as G345R may not require homodimer formation.

OR004
INVOLVEMENT OF THE cGMP PATHWAY IN THE INHIBITION OF THE IODIDE UPTAKE AND THE EXPRESSION OF THYROID SPECIFIC GENES IN FRTL-5
Bazzara LG, Costamagna ME, Cabanillas AM, Velez ML, Fozzatti L, Masini-Repiso AM.
Depto. de Bioq. Clínica, Fac. de Cs. Químicas, Universidad Nacional de Córdoba, Córdoba, Argentina.
       Previous reports indicated that nitric oxide (NO) inhibits the iodide uptake in several species and the thyroperoxidase (TPO) mRNA expression in FRTL-5 cells. This work was aimed to analyse the role of the guanylyl cyclase/cyclic GMP (cGMP) pathway in the inhibitory effect of NO on iodide uptake and in the expression of TPO and thyroglobulin (TG) mRNA in TSH (500 mU/ml)-stimulated FRTL-5. Sodium Nitroprusside (SNP), a NO donor, increased cGMP levels (RIA) in a concentration and time-dependent manner (Table 1). The cAMP level (RIA) decreased after 48 h of incubation with 500 mM SNP and was not modified at shorter times and lower SNP concentrations. An increase of cGMP was induced by TSH (Table 1). The inhibition exerted by SNP or 8-Br-cGMP for 24 h on the iodide uptake (131I) was reversed by KT-5823 (1-10 mM), an inhibitor of the cGMP-dependent protein kinase (PKG) (Table 2). The TPO and TG mRNA expression (Northern-blot) was significantly reduced by 8-Br-cGMP. Data (24 h) were (Absorbance ratio mRNA/18 S rRNA, mean±SEM, arbitrary units): Control (C): TPO= 2.00, TG= 2.00; 8-Br-cGMP 100 mM: TPO= 1.97±0.24, TG= 1.96±0.03; 300 mM: TPO= 1.56±0.13, TG= 1.23±0.16**; 1000 mM: TPO= 1.22±0.29*, TG= 1.01±0.26** (*P<0.05, **P<0.01 vs C; n=3, ANOVA, Student-Newman-Keuls test). In conclusion, an activation of PKG seems to be involved in the NO/cGMP-induced inhibition of iodide uptake in FRTL-5. A reduction in the cAMP production could be relevant in the action of high NO/cGMP levels. Evidence is provided for a role of the cGMP pathway in the regulation of thyroid specific genes.

OR005
THYROID HORMONE DEFICIENCY SHOWS A SELECTIVE IMPAIRMENT ON MYELIN MARKERS DURING DEVELOPMENT.
Barradas, P.C., Vieira, R.S., De Freitas,M.S.
Instituto de Biologia, UERJ, 20551-030,Brasil.
       We previously described that mature myelinated fibers, from hypothyroid rats, showed a continuity of staining as revealed by 2'3'cyclic nucleotide 3'phosphodiesterase (CNPase) immunohistochemistry during development that contrasted with the pattern of the normal mature myelinated fibers and suggests a possible role for thyroid hormones on myelin compaction. The most abundant proteins of CNS myelin, myelin basic protein (MBP) and proteolipidic protein (PLP), are expressed when the myelin sheath formation is in progress. Another myelin constituent designated myelin-associated/oligodendrocytic basic protein (MOBP) was identified and related to myelin compaction. We assessed the developmental sequence of appearance of CNPase, MBP, MOPB and PLP proteins in cerebellum (Cb) and corpus callosum (cc) from experimental hypothyroidism model. Post-natal animals from 10 to 60 days (P10 to P60), normal (C) and hypothyroid (H) were used. To induce the hypothyroid status animals received methimazol since 19th gestational until the killing. We performed an immunoblotting analysis using an anti-MOBP antiserum (gift from Dr.Holz),monoclonal anti-CNPase or anti-MBP antibodies or polyclonal anti-PLP. The appearance of both MOBP isoforms occurred at P25 and P30 in cc and Cb, respectively, followed by an increase with age in C group. However, all the MOBP isoforms were weakly detectable in both regions at P30 from H group and one isoform remains decreased in cc, even at P90. The developmental pattern of CNPase, MBP and PLP proteins expression was also delayed in H group. CNPase and MBP expression was recovered in adulthood, while PLP isoforms remained below from control levels at P90 in cc. Our data show that hypothyroidism affects the developmental pattern of the oligodendrocytic/myelin markers. Furthermore, the permanent impairment on MOBP and PLP expression reinforces the suggestion of a role for thyroid hormone on myelin compaction. Supported by: TWAS, FAPERJ, CNPq, SR2/UERJ

OR006
LIPOPOLYSACCHARIDE (LPS) INHIBITS THE THYROGLOBULIN (Tg) AND THYROID PEROXIDASE (TPO) mRNA EXPRESSION AND MODULATES THE IODIDE UPTAKE IN FRTL-5
Vélez ML, Costamagna ME, Bazzara LG, Fozzatti L and Masini-Repiso AM. Depto. de Bioq. Clín., Fac. de Cs. Químicas, Universidad Nacional de Córdoba, Córdoba, Argentina
       Bacterial endotoxin LPS induces the expression of multiple genes including nitric oxide (NO) synthase. LPS is present in blood in certain infectious processes. This study analyzed the LPS action on the expression of thyroid specific genes and the iodide uptake in TSH (500mIU/ml)-stimulated FRTL-5. The Tg and TPO mRNA expression (Northern-blot) was decreased by LPS (0.01-1mg/ml): (absorbance ratio mRNA/18S rRNA; arbitrary units): Tg: Basal(B): 0.45±0.09; Control(C): 1.00±0.03; LPS (mg/ml) 0.01: 0.59±0.04**, 0.05: 0.55±0.05**, 0.1: 0.66±0.09*, 1: 0.79±0.09. TPO: B: 0.51±0.07; C: 1.00±0.06; LPS (mg/ml) 0.01: 0.64±0.03**, 0.05: 0.61±0.03**, 0.1: 0.61±0.08*, 1: 0.69±0.14 (n=3; **p<0.01; *p<0.05 vs C). A non significant reduction in the sodium-iodide transporter (NIS) mRNA expression was induced by LPS. The iodide uptake (131I) was increased by LPS (0.01-10mg/ml) in a concentration-dependent manner (maximum at 0.1mg/ml): (cpm/mg DNA; 48h): B: 91.2±11.4; C: 593.6±26.6; LPS (mg/ml) 0.01: 1161.9±110.6***; 0.05: 1103.2±96.1***; 0.1: 1532.6±148.6***; 1: 1071.5±212.0**; 5: 972.4±156.2**; 10: 899.2±123.9** (n=14; ***p<0.0001; **p=0.01 vs C). This effect was time-dependent (highest value at 48h) (Table). In contrast, LPS inhibited iodide uptake at higher concentrations: (% of control; 48h): C: 100±4%; LPS (mg/ml) 100: 144±36%, 500: 44±10%***, 1000: 21± 3%*** (n=9; ***p<0.0001 vs C). LPS (0.01-1000mg/ml; 48h) did not induce NO generation (nitrite accumulation). Results (mean±SEM) were analyzed by ANOVA and Student t test. In conclusion, these findings revealed that LPS differentially influence thyroid specific gene expression and iodide uptake. A mediation of NO in the LPS action is unlikely. The changes induced by LPS on parameters of thyroid hormone biosynthesis support a potencial ability of the endotoxin to alter the thyroid function.

OR007
IDENTIFICAÇÃO DE PROVÁVEIS MARCADORES TUMORAIS ASSOCIADOS À NEOPLASIAS DA TIRÓIDE HUMANA.
Arnaldi LAT, Maciel RMB, Cerutti JM.
Laboratório de Endocrinologia Molecular, Disciplina de Endocrinologia, Universidade Federal de São Paulo - EPM, São Paulo, Brasil.
       As neoplasias derivadas da célula folicular da tiróide representam um excelente modelo para análise de tumores que se desenvolvem como resultado de mutações seqüenciais nos genes que controlam o crescimento celular normal. Entre os eventos genéticos, foram identificados alterações nos oncogenes RET, RAS, TRK, GSP, CMYC, MET, nos genes supressores de tumor P53, RB e p16 além de um aumento de expressão da proteína de adesão Galectina 3. Embora alterações nestes genes tenham sido identificadas e estas associadas às características morfológicas, comportamento clínico e biológico dos tumores, a compreensão do mecanismo molecular que determina os diferentes fenótipos necessita de maiores informações. Este trabalho tem como objetivo a identificação de genes que poderão ser utilizados como novos marcadores tumorais correlacionados com a determinação e progressão dos diferentes subtipos de tumores da tiróide. Para isso, 220 genes que fazem parte do banco de seqüências do Projeto Genoma Humano do Câncer FAPESP/ILCR foram escolhidos e classificados de acordo com a função: fatores de crescimento, receptores de membrana, proteínas citoplasmáticas, proteínas nucleares e moléculas de adesão celular. Para screening inicial foi utilizada a técnica de Northern reverse onde os genes foram fixados em membranas e estas analisadas paralelamente utilizando sondas de cDNA sintetizadas a partir das linhagens celulares de carcinoma indiferenciado (ARO), carcinoma papilífero (NPA), carcinoma folicular (WRO) e controle derivado de tecido normal. Dos 220 genes analisados, 45 mostraram alteração no padrão de expressão quando comparados entre si. Destes, 3 genes foram inicialmente selecionados para análise nos tecidos derivados de pacientes com carcinomas da tiróide, os quais confirmaram variabilidade nos níveis de expressão. Estes resultados sugerem que estes genes tem potencialidade para serem novos marcadores tumorais e, portanto, facilitarão a compreensão dos mecanismos moleculares envolvidos na tumorigênese da tiróide.

OR008
DIFFERENTIAL IMMUNOHISTOCHEMISTRY REACTIVITY OF GALECTIN-3 IN HÜRTHLE CELL ADENOMAS AND CARCINOMAS
Nascimento MC1, Bisi H2, Alves V2, Longatto-Filho A3, Kanamura CT3 and Medeiros-Neto G1.
1Thyroid Unit (LIM-25), Univ Sao Paulo Med School, Sao Paulo, Brazil, 2Pathology, Univ Sao Paulo Med School, Sao Paulo, Brazil, 3Inst Adolfo Lutz, State Health Div, Sao Paulo, Brazil
      Hürthle cell carcinomas represent about 3.5% of all types of thyroid carcinomas, are a more aggressive variety of follicular carcinoma, with frequent recurrences and higher morbidity. Galectin-1 and galectin-3 have been implicated in the regulation of cellular growth, differentiation and malignant transformation in thyroid neoplasm. More specifically galectin-3, a beta-galactoside binding protein, is highly expressed in thyroid malignancies of epithelial origin (papillary and follicular carcinomas). The current study was undertaken to investigate reactivity of galectin-3 by immunohistochemistry in thyroid tissues with Hürthle cells adenomas and carcinomas (n=31), in other differentiated thyroid cancers (n=25), colloid goiter (n=30), Hashimoto thyroiditis (n=11) and normal thyroid tissues (n=18). Galectin-3 expression was examined using a mouse monoclonal Ab against galectin-3 in Hürthle cell neoplasia and other thyroid pathologies. As expected immunohistochemistry for galectin-3 was positive (60-100%) in all follicular and papillary cancers being higher when pathological examination revealed the presence of Hürthle cells. There was no tissue positivity in all specimens from Hashimoto thyroiditis, colloid goiters and normal thyroid tissues. The frequency of galectin-3 expression in Hürthle cell carcinoma was significantly higher (59%) as compared with the expression in Hürthle cell adenomas (7.1%, p<0.05). These results suggest that galectin-3 may potentially serve as a marker for distinguishing benign thyroid Hürthle cell adenomas from Hürthle cell carcinomas.

OR009
AN UPDATE OF THYROPEROXIDASE (TPO) GENE MUTATIONS IN CONGENITAL GOITROUS HYPOTHYROIDISM WITH PARTIAL IODIDE ORGANIFICATION DEFECT.
Nascimento AC1, Guedes DR1, Santos CS1, Rubio I1 and Medeiros-Neto G1.
1Thyroid Unit (LIM-25), Univ Sao Paulo Med School, Sao Paulo, Brazil
      In most patients with total iodide organification defect (TIOD), the peroxidase and hormone producing activities are undetectable. A subgroup of patients, however, exhibit a partial discharge of iodide after perchlorate, have some TPO activity in the thyroid tissue, resulting in low but detectable hormone production. We have studied five unrelated families with nine affected children with congenital hypothyroidism. All patients had elevated radioiodine uptake that was partially (30-48%) discharged by perchlorate. DNA from peripheral blood lymphocites was isolated, the TPO gene was studied by DGGE of PCR products. Exons with aberrant DGGE patterns were sequenced. Several polymorphisms were found, respectively, in exon 2 (position 102), in exon 8 (position 1209), in intron 8 (insertion of a single nucleotide), in intron 8 (substitution of G by T), in exon 11 (position 2088) and intron 13 (substitution C by T). It is unclear if these polymorphisms will affect the translated protein. Previously described mutations were found in our patients such as two mutations in exon 8 in positions, respectively, 1207 (Ala 373 Ser) and 1283 (Ser 398 Thr). New mutations were found in exon 8 at position 1242 (Glu 384 Asp), in intron 9 (AG/AT) that putatively affect splicing, in exon 10 (position 1780, Leu 564 Ile) and exon 11 (position 2084, Arg 665 Glu). By contrast with TIOD none of these TPO gene mutations were followed by a premature stop codon and frameshift. It is possible that some of these point mutations will only partially affect the catalytic action of the TPO mutant protein allowing for some thyroid hormone formation in vivo.

OR010
EXPRESSÃO DIFERENCIADA DOS PROTO-ONCOGENES H-RAS E K-RAS NO BÓCIO NODULAR.
Golbert L, Posser M, Lobato R, Maia AL.
Serviço de Endocrinologia, Hospital de Clínicas de Porto Alegre, Faculdade de Medicina, UFRGS, Porto Alegre, Brasil.
       Os proto-oncogenes estão envolvidos no controle do crescimento celular e mutações/aumento da expressão desses genes estão relacionados ao crescimento celular excessivo em detrimento da diferenciação. Alterações no proto-oncogene ras têm sido descritas em tumores benignos e malignos da tireóide, sugerindo que possam ser um evento inicial na transformação neoplásica da célula tireoidiana. Neste estudo avaliamos a expressão gênica e a presença de mutações no ras em tecido de bócio multinodular. Foram incluídos 115 pacientes submetidos à tireoidectomia, entre março de 1999 a fevereiro de 2001, no HCPA, tendo sido coletado fragmentos de tireóide normal e neoplásica durante o ato cirúrgico e imediatamente congelados. O RNA total foi extraído pelo método de Trizol (Gibco BRL) e o cDNA sintetizado através do Reverse Transcriptidase. Os genes H-ras e K-ras foram amplificados por PCR com primers específicos e os amplicons submetidos a eletroforese em gel para análise e quantificação (AMBIS System). Na detecção de mutações utilizou-se a técnica de SSCP. A média de expressão do H-ras no tecido neoplásico foi maior em comparação com a do normal, porém não houve diferença significativa na expressão do K-ras.
Entretanto, a proporção de casos com aumento da expressão desses proto-oncogenes no tecido neoplásico foi semelhante para os dois genes analisados, sendo que 50% do total de casos apresentaram aumento da expressão do H e/ou K-ras. Na correlação com os dados clínicos, a presença de maior expressão do ras foi associada a um crescimento mais rápido do bócio. Os resultados encontrados sugerem papéis diferenciados para os genes da família ras na gênese das neoplasias de tireóide.

OR011
UP-REGULATION OF MYOGLOBIN (MB) GENE EXPRESSION BY THYROID HORMONE (T3) IN RAT CARDIAC MUSCLE: POSSIBLE INVOLVEMENT OF E-BOX-ASSOCIATED PROTEINS.
Gisele Giannocco., Rosangela A. Santos, Maria Tereza Nunes.
       Department of Physiology and Biophysics, ICB1, University of São Paulo, São Paulo, Brazil. T3 markedly increases heart function. Considering that Mb gene is highly expressed in the heart, improving O2 diffusion and mitochondrial respiration, we investigated if T3 is involved in Mb gene expression and if DNA binding activity of nuclear proteins are altered by T3 using E-box consensus sequence as a probe. Time-course studies were performed in euthyroid and thyroidectomized (Tx) rats treated, or not, with 100 ug/100 g of T3, BW, iv, for 30, 60, 120 min, 6 and 24 h. Rats were killed by decapitation; nuclear and cytosolic protein from ventricular (V) muscle were isolated. The cytosolic protein was electrophoresed and the Mb abundance were determined by western blot, using a laser-scanning densitometer. Nuclear protein was used to perform the electrophoretic mobility shift assays (EMSA). It was shown that Mb expression is under T3 control in V muscle, as indicated by decreased Mb protein content in Tx rats. The time-course study showed a progressive increase in the Mb protein abundance, as early as 30 min, which exceeded the control levels at 24 h of the T3 treatment. EMSA showed an enhanced capacity of nuclear proteins to bind the E-box probe after T3 treatment while Tx rats displayed a diminished binding. The present results showed that (1) Mb protein and, as previously shown, mRNA content are influenced by T3 and (2) T3 enhanced the binding capacity of E-box- associated proteins in rat cardiac muscle.
Apoio financeiro: FAPESP

OR012
EXPRESSÃO DA MIOGLOBINA (Mb) INDUZIDA PELO HORMÔNIO TIREOIDEANO EM CULTURA PRIMÁRIA DE MÚSCULO ESQUELÉTICO DE RATOS.
Bielavsky, M e Nunes, M.T.
       Depto. Fisiologia e Biofísica ICB1 - USP, São Paulo, SP. Trabalhos anteriores de nosso laboratório demonstraram que o hormônio tireoideano (T3) está envolvido na regulação da expressão gênica da mioglobina (Mb), proteína importante no transporte e consumo de oxigênio no músculo cardíaco e esquelético. Neste trabalho, utilizamos culturas primárias de músculo esquelético com o objetivo de verificar se o T3 pode regular sua expressão gênica, bem como da proteína, na ausência de influências hormonais ou de inervação. As culturas foram obtidas da musculatura da coxa de ratos de 4 dias de vida (Horn e Brodwick, 1980) e mantidas em meio de cultura com soro fetal depletado de T3, após 6-7 dias em cultura foram incubadas com T3 nas concentrações de 10-9 a 10-6 M por 24, 48 e 72 h. O conteúdo de RNA foi analisado através de Northern Blot, hibridando-se a membrana com 32P-cDNA da Mb. Os valores da densitometria das bandas corrigidos com sonda para a 18S ribossomal. O tratamento das culturas com T3 (10-9 a 10-6 M) por 24 e 48 h não induziu diferença na expressão gênica da Mb, entretanto, o tratamento por 72 h com T3 resultou em um aumento do conteúdo de mRNA de Mb nas concentrações de 10-8 a 10-6M. A expressão da proteína mioglobina foi avaliada por imunoprecipitação seguida de Western Blot nas concentrações de 10-8 a 10-6M por 48, 72 e 96 h. e mostrou-se fracamente expressa em culturas tratadas e não tratadas. O hormônio tireoideano causa um aumento na expressão gênica da Mb apenas em 72 h de tratamento neste modelo livre de influências neurais. A expressão da proteína Mb não acompanha o aumento de seu RNAm neste tempo, tampouco após 96 h de tratamento. Estes dados sugerem que o efeito do T3 na expressão gênica da Mb depende de um programa de ativação gênica tempo-dependente que ocorre de maneira precisa na célula esquelética.
Apoio Financeiro FAPESP.

[VOLVER] [SIGUIENTE]