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Title: Chitosan-mediated shRNA knockdown of cytosolic alanine aminotransferase improves hepatic carbohydrate metabolism
Author: González, Juan D.
Silva-Marrero, Jonás I.
Metón Teijeiro, Isidoro
Caballero Solares, Albert
Viegas, Iván
Fernández González, Felipe Javier
Miñarro Carmona, Montserrat
Fábregas, A. (Anna)
Ticó Grau, Josep R.
Jones, John G.
Silva-Marrero, Jonás I. Villasante, Juliana Rashi
Keywords: Quitosan
Teràpia genètica
Gene therapy
Sparus aurata
Issue Date: Feb-2016
Publisher: Springer Verlag
Abstract: Alanine aminotransferase (ALT) catalyses a transamination reaction that links carbohydrate and amino acid metabolism. In this study, we examined the effect of silencing cytosolic ALT (cALT) expression on the hepatic metabolism in Sparus aurata. A number of siRNA and shRNA designed to down-regulate cALT expression were validated in HEK-293 cells transfected with plasmids expressing S. aurata cALT or mitochondrial ALT (mALT) isoforms: ALT silencing significantly decreased the expression levels of S. aurata mRNA cALT1 to 62 % (siRNA) and 48 % (shRNA) of the values observed in control cells. The effect of cALT silencing was analysed in the liver of S. aurata 72 h after intraperitoneal injection of chitosan-tripolyphosphate (TPP) nanoparticles complexed with a plasmid encoding a shRNA to down-regulate cALT expression (pCpG-si1sh1). In fish fed diets with different ratio of protein to carbohydrate and treated with chitosan-TPP-pCpG-si1sh1, cALT1 and cALT2 mRNA levels significantly decreased irrespective of the diet. Consistently, ALT activity decreased in liver of treated animals. In the liver of S. aurata treated with chitosan-TPP-pCpG-si1sh1 nanoparticles, down-regulation of cALT expression increased the activity of key enzymes in glycolysis (6-phosphofructo-1-kinase and pyruvate kinase) and protein metabolism (glutamate dehydrogenase). Besides showing for the first time that administration of chitosan-TPP-pCpG-si1sh1 nanoparticles silences hepatic cALT expression in vivo, our data support that down-regulation of cALT could improve the use of dietary carbohydrates to obtain energy and spare protein catabolism
Note: Versió postprint del document publicat a:
It is part of: Marine Biotechnology, 2016, vol. 18, num. 1, p. 85-97
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ISSN: 1436-2228
Appears in Collections:Articles publicats en revistes (Farmàcia, Tecnologia Farmacèutica i Fisicoquímica)
Articles publicats en revistes (Bioquímica i Fisiologia)

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