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O ácido butírico : possível regulador da expressão dos genes do relógio circadiano intestinal?
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Abstract(s)
O presente estudo foi elaborado em células Caco-2, para verificar se a glucose e o ácido butírico regulam a expressão dos genes do relógio circadiano intestinal. O seguimento da expressão de SRF sobre 18h, por imuno-detecção por epifluorescência, confirma que se trata de um input do relógio. A imunodetecção da expressão oscilatória de PERIOD1 sobre 18h e 48h (com intervalos de 6h) comprova que Period1 é um gene da resposta imediata, fundamental ao funcionamento do relógio. Verificamos ainda que a expressão de PERIOD1 é sincronizada com maior eficácia, utilizando um meio com 1mM de ácido butírico, relativamente a outros meios de cultura (DMEM 10% FCS, 25mM D-glucose, sem glucose nem ácido butírico). A redução significativa da concentração em ácido butírico (0,5, 1 e 2mM) nos sobrenadantes de monocamadas de células Caco-2, comparativamente a sobrenada-ntes incubados em placas sem células (ANOVA em triplicado; significância: 5%), sugere que o composto será consumido pelas células. Integrando essa observação com o resultado da experiência anterior, deduzimos que o ácido butírico utilizado pelas células servirá de regulador à expressão do gene Period1. O uso desse composto poderá assim diminuir a dessincronização dos ritmos de células do relógio, que estará na origem de determinadas patologias.
ABSTRACT: This study was realized in Caco-2 cells to determine whether glucose and butyric acid regulate the expression of circadian clock genes in the intestine. The follow-up of the expression of SRF on 18h, by immunodetection by epifluorescence, confirms that it is an input of the clock. Immunodetection of oscillatory PERIOD1 expression on 18h and 48h (with a 6 hours step) shows that Period1 is an immediate response gene, critical for the correct functioning of the clock. We also note that the expression of PERIOD1 is synchronized more effectively, using a medium with 1 mM butyric acid in relation to other culture media (DMEM 10% FCS, 25mM D-glucose, without glucose nor butyric acid). A significant reduction in the concentration of butyric acid (0.5, 1 and 2mM) in supernatants of monolayers of Caco-2 cells, compared with supernatants incubated on cell-free plastic wells (ANOVA in triplicate; significance: 5%), suggests that the compound can be consumed by the cells. Integrating this observation with the result of previous experience, we deduce that the butyric acid used by cells can serve as regulator of Period1 gene expression. The use of this compound could help reduce the desynchronization of clock cells rhythms, that can lead to certain diseases.
ABSTRACT: This study was realized in Caco-2 cells to determine whether glucose and butyric acid regulate the expression of circadian clock genes in the intestine. The follow-up of the expression of SRF on 18h, by immunodetection by epifluorescence, confirms that it is an input of the clock. Immunodetection of oscillatory PERIOD1 expression on 18h and 48h (with a 6 hours step) shows that Period1 is an immediate response gene, critical for the correct functioning of the clock. We also note that the expression of PERIOD1 is synchronized more effectively, using a medium with 1 mM butyric acid in relation to other culture media (DMEM 10% FCS, 25mM D-glucose, without glucose nor butyric acid). A significant reduction in the concentration of butyric acid (0.5, 1 and 2mM) in supernatants of monolayers of Caco-2 cells, compared with supernatants incubated on cell-free plastic wells (ANOVA in triplicate; significance: 5%), suggests that the compound can be consumed by the cells. Integrating this observation with the result of previous experience, we deduce that the butyric acid used by cells can serve as regulator of Period1 gene expression. The use of this compound could help reduce the desynchronization of clock cells rhythms, that can lead to certain diseases.
Description
Dissertação de Mestrado em Engenharia Zootécnica.
Keywords
Ácido Butírico Caco-2 Relógio Circadiano Intestinal Butyric Acid Intestinal Circadian Clock