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Previous issue date: 2016-02-26 / Coordena??o de Aperfei?oamento de Pessoal de N?vel Superior - CAPES / The 17?-estradiol (E2) regulate many cardiac genes via its estrogen receptor (ER). The propagation of electrical activity in the myocardium depends on the current transfer at gap junctions. Connexins 40 (Cx40) and 43 are the predominant junctional proteins. In mice, Cx40 is restricted to the atrium and conduction system. Alterations of Cx40 expression or activity are associated with atrial fibrillation. Here we evaluate the effect of the E2 onCx40 mRNA expression, in vitro, and in vivo the effect on atrium expression of Cx40 mRNA in correlation to ECG studies. We treated A7r5 cells (smooth muscle cells from rat thoracic aorta) with low and high doses ofestradiol benzoate, EB, (10-8 M and 10-6M) for 24h and rat neonatal cardiomyocytes with EB 10-6 M for 2, 4 and8 hours. A7r5 cells were trasiently transfected with 0.5 microgramas of the test plasmid (-1190/+121Cx40Luc pGL3) and treated with 10-6M of EB for 24 hours. Female mice were subjected to ovariectomy (OVX) and then, treated with a low (2?g) and a high dose (20?g) of estradiol benzoate (EB; OVX+EB2 and OVX+EB20) for 15 days. ECG recordings were obtained from mice and atrium Cx40 mRNA expression were evaluated by RT-PCR real time. First, we observed that high doses of EB down regulated Cx40 mRNA in vitro (A7r5 cells and rat atrial cardiomyocytes). The transcriptional activity of Cx40 promoter was inhibited by 10-6M of EB in A7r5 cells. We observed lower heart rate for OVX animals when to compared to control animals, FO. In this regard, no difference was observed between OVX+EB2 and OVX+BE20 heart rate. The OVX group showed decrease P wave duration when to compared to FO group, but no difference in the P wave duration was observed among the others groups. No difference was observed in another ECG intervals among the experimental groups. The atrial Cx40mRNA level was reduced in OVX+BE20group when to compared to FO group. Our data indicate, for the first time, high doses of estradiol benzoate reduce Cx40 mRNA in vitro and ovariectomized mice. We proposed that when E2 levels are high, the complex ER-E2 acts directly from the DNA, inhibiting the transcriptional activity of Cx40 promoter / O 17?-estradiol (E2), regula muitos genes card?acos atrav?s de seu receptor (receptor para estr?geno, RE). A propaga??o da atividade el?trica no mioc?rdio depende da transfer?ncia de corrente atrav?s das jun??es comunicantes. As conexinas 40 (Cx40) e 43 s?o as principais conexinas que formam as jun??es expressas no cora??o. Em camundongos, a Cx40 ? restrita ao ?trio e sistema de condu??o. Altera??es na express?o ou atividade da Cx40 est?o associadas a fibrila??o atrial. Aqui n?s avaliamos o efeito do E2 in vitro e in vivo sobre a express?o do RNAm da Cx40 nos ?trios e correlacionamos aos estudos eletrocardiogr?ficos (ECG). N?s tratamos a linhagem A7r5 (derivada de m?sculo liso a?rtico de rato embrion?rio) com alta e baixa concentra??es de benzoato de estradiol, BE, (10-6 M e 10-8 M) durante 24 horas e cultura prim?ria de cardiomi?citos atriais de ratos neonatos foram incubados com BE 10-6 M durante 2, 4 e 8 horas. Ensaios de transfec??o transiente foram realizados na linhagem A7r5 com um plasm?deo contendo um segmento da regi?o promotora da Cx40 (-1190/+121Cx40LucpGL3) e tratadas com 10-6 M de BE durante 24 horas. Camundongos f?meas foram ovariectomizadas (OVX) e ent?o tratadas com baixa (2 microgramas) e alta (20 microgramas) doses de benzoato de estradiol (OVX+BE2 e OVX+BE20) durante 15 dias, sendo que o grupo controle sofreu apenas estresse cir?rgico (FO). Foram realizados registros ECG e a express?o atrial do RNAm da Cx40 foi avaliada por RT-PCR em tempo real. N?s observamos que altas doses de BE diminui a express?o do RNAm da Cx40 in vitro (na linhagem A7r5 e na cultura de cardiomi?citos). A atividade transcricional do promotor foi inibida por BE10-6 M. N?s observamos diminui??o da frequ?ncia card?aca dos animais do grupo OVX em rela??o ao controle, grupo FO. Nenhuma diferen?a foi observada na frequ?ncia card?aca entre os grupos OVX+BE2 e OVX+BE20. Os animais do grupo OVX apresentaram diminui??o na dura??o da onda P em rela??o ao grupo FO, no entanto nenhuma diferen?a foi observada na dura??o da onda P entre os outros grupos. Nenhuma diferen?a foi observada nos outros intervalos eletrocardiogr?ficos entre os grupos experimentais. A express?o atrial do RNAm da Cx40 estava reduzida nos animais do grupo OVX+BE20 em rela??o ao grupo FO. Nossos dados demonstram que altas doses de benzoato de estradiol reduzem a express?o do RNAm da Cx40 in vitro e em camundongos ovariectomizados. N?s propomos que altas doses de E2 ativa o seu receptor, e o complexo RE-E2 age diretamente no DNA, inibindo a atividade transcricional do promotor da Cx40
| Identifer | oai:union.ndltd.org:IBICT/oai:localhost:jspui/1394 |
| Date | 26 February 2016 |
| Creators | Amarante, D?bora Barbosa |
| Contributors | Almeida, Norma Aparecida dos Santos, Fortes, Fabio da Silva de Azevedo, Oliveira, Elaine de |
| Publisher | Universidade Federal Rural do Rio de Janeiro, Programa de P?s-Gradua??o em Ci?ncias Fisiol?gicas, UFRRJ, Brasil, Instituto de Ci?ncias Biol?gicas |
| Source Sets | IBICT Brazilian ETDs |
| Language | Portuguese |
| Detected Language | English |
| Type | info:eu-repo/semantics/publishedVersion, info:eu-repo/semantics/masterThesis |
| Format | application/pdf |
| Source | reponame:Biblioteca Digital de Teses e Dissertações da UFRRJ, instname:Universidade Federal Rural do Rio de Janeiro, instacron:UFRRJ |
| Rights | info:eu-repo/semantics/openAccess |
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