Gene expression on Rat1 fibroblast cells after transformation by EVI1. Rev. Ang. de Ciênc. da Saúde. 2021 Jan – Jul 1 (1): 21-23
A comparison between Rat1neo and Rat15.6 according to the enzyme Luciferase activity as CAIII gene candrive the expression of firefly luciferase from its promotor and expression of renilla luciferase by thecytomegalovirus (CMV) also from its promotor, renilla luciferase was used to normalise the results as it is
permanently present, without vary, in transfected cells with plasmids carrying both exotic genes but theexpression of firefly may alter and this variation can be measured according to the intensity of luciferase activityin the gene promotor region and for this work it was higher in Rat1neo and lower for Rat15.6.Luciferase activity was higher for Rat1neo and it may indicate that CAIII is more expressed in Rat1neo cellsthan in Rat15.6 where EVI1 gene is expressed.Caspase 3 activity in Rat1 cells was measured after hydrogen peroxide (H 2 O 2 ) treatment to evaluateapoptosis. Therefore, siRNA was used to target the expression of CAIII gene (siRNA CAIII) in Rat1neo cells bytransfection in distinction to siRNA control and untreated cells. After, the role of CAIII gene in cell protectionagainst oxygen-stress induced apoptosis was observed by caspase 3 activity among untreated, siRNA controland siRNA CAIII.The Caspase 3 activity was higher in siRNA CAIII as the protection factor against H 2 O 2 was knocked downby the dicer silencing targeting CAIII RNA and apoptosis raised up.
CONCLUSION So, caspase 3 activity was higher in Rat1neo cells treated with siRNA CAIII and H 2 O 2 compared to theuntreated which prove the repression of CAIII by EVI1. Moreover, the luciferase activity demonstrated that EVI1overexpression, at the molecular level, represses CAIII expression by acting on its promotor region.Consequently, cells, where CAIII is knocked out, are no longer protected against H 2 O 2 and elevated caspase 3activity increases the level of apoptosis. Following these principles, it is suggested that in leukaemia if drugscontaining related H 2 O 2 agents are used, they may selectively target transformed cells with EVI1 geneoverexpression and these could be candidates for the cancer therapy..
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