Construction and effect of recombinant adenovirus vector containing siRNA targeting Raf-1 on cardiomyocyte hypertrophy in neonatal rats / 中国比较医学杂志
Chinese Journal of Comparative Medicine
;
(6): 18-23, 2016.
Artigo
em Chinês
| WPRIM
| ID: wpr-486232
ABSTRACT
Objective To construct an adenovirus vector expressing small interfering RNA ( siRNA) targeting to rat Raf-1 gene and identify its function in cardiomyocytes.Methods The siRNA containing DNA sequence targeting to Raf-1 and its negative control sequence were designed, synthesized, annealed and subcloned into adenoviral shuttle vector pAdTrack-CMV.The recombinant adenovirus vector pAd-siRaf-1 was obtained by homologous recombination with pAdTrack-siRaf-1 linearized by PmeI and pAdeasy-1 in bacteria BJ5183, then transfected into HEK293 cells to package the adenovirus.Cardiomyocytes were infected with the adenovirus pAd-siRaf-1, and the expressions of Raf-1 and NF-κB protein were detected by Western blotting.[ 3 H ]-leu incorporation was evaluated by scintillation.The surface area of cardiomyocytes was measured using a HJ2000 image analysis system.Results The adenovirus vectors were verified by enzyme digestion and DNA sequencing.Compared with the Ang II group, Raf-1 and NF-κB expression, the surface area and [ 3 H]-leu incorporation of cardiomyocytes were significantly decreased in cardiomyocytes infected with the adenovirus PAd-siRaf-1.Conclusions A recombinant adenovirus vector containing rat siRaf-1 gene is successfully constructed.It can effectively reduce Raf-1 and NF-κB expression and cardiomyocyte hypertrophy induced by Ang II.
Texto completo:
DisponíveL
Índice:
WPRIM (Pacífico Ocidental)
Idioma:
Chinês
Revista:
Chinese Journal of Comparative Medicine
Ano de publicação:
2016
Tipo de documento:
Artigo
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