Marzieh Rostaminejad1,2,3, Abdolmohamad Rostami4, Abbas Behzad-Behbahani2, Gholam Reza Rafiei Dehbidi2, Tahereh Kalantari5,6. 1. Division of Medical Biotechnology, Department of Medical Laboratory Sciences, School of Paramedical Sciences, Shiraz University of Medical Sciences, Meshkin-Fam St., Shiraz, Iran. 2. Diagnostic Laboratory Sciences and Technology Research Center, School of Paramedical Sciences, Shiraz University of Medical Sciences, Shiraz, Iran. 3. Epilepsy Research Center, Shiraz University of Medical Sciences, Shiraz, Iran. 4. Department of Neurology, Thomas Jefferson University, Philadelphia, PA, 19107, USA. 5. Division of Medical Biotechnology, Department of Medical Laboratory Sciences, School of Paramedical Sciences, Shiraz University of Medical Sciences, Meshkin-Fam St., Shiraz, Iran. taherehk@yahoo.com. 6. Diagnostic Laboratory Sciences and Technology Research Center, School of Paramedical Sciences, Shiraz University of Medical Sciences, Shiraz, Iran. taherehk@yahoo.com.
Abstract
BACKGROUND: The kynurenine pathway (KP) can be involved in the pathogenesis of neurodegenerative diseases and excessive neurotoxic metabolite production. This study aimed to evaluate the effects of overexpression of murine 2-amino-3-carboxymuconate-6-semialdehyde decarboxylase (Acmsd) gene in inflammatory conditions in RAW 264.7 cell line to present more information about the effect of this gene on inflammatory conditions and the KP cycle. METHODS AND RESULTS: The coding sequence of the Acmsd gene was cloned into pCMV6-AC-IRES-GFP expression vector with a green fluorescent protein (GFP) marker. To simulate inflammatory conditions, RAW 264.7 macrophage cells were stimulated by Lipopolysaccharide (LPS) 24 h before transfection, and transfected by Polyethyleneimine (PEI) with constructed plasmids expressing the Acmsd gene. The effect of Acmsd gene expression level on murine Interferon-gamma (Ifn-γ) and murine Indoleamine 2,3-dioxygenase 1 (Ido1) gene expression level was investigated by Real-Time PCR. According to the results of this study, good transfection efficiency was observed 72 h after transfection, and Acmsd expression level increased 29-fold (P < 0.001) in transfected LPS-stimulated cells compared to the control group (LPS-stimulated cells that were not transfected). Additionally, increased Acmsd expression level significantly down-regulated Ifn-γ (P < 0.001) and Ido1 (P < 0.01) expression level in transfected LPS-stimulated cells compared to LPS-stimulated cells. CONCLUSIONS: Acmsd gene overexpression in inflammatory conditions can reduce the expression levels of the Ido1 gene, and its regulator, Ifn-γ. Consequently, it may be considered as a novel regulatory factor in the KP balance.
BACKGROUND: The kynurenine pathway (KP) can be involved in the pathogenesis of neurodegenerative diseases and excessive neurotoxic metabolite production. This study aimed to evaluate the effects of overexpression of murine 2-amino-3-carboxymuconate-6-semialdehyde decarboxylase (Acmsd) gene in inflammatory conditions in RAW 264.7 cell line to present more information about the effect of this gene on inflammatory conditions and the KP cycle. METHODS AND RESULTS: The coding sequence of the Acmsd gene was cloned into pCMV6-AC-IRES-GFP expression vector with a green fluorescent protein (GFP) marker. To simulate inflammatory conditions, RAW 264.7 macrophage cells were stimulated by Lipopolysaccharide (LPS) 24 h before transfection, and transfected by Polyethyleneimine (PEI) with constructed plasmids expressing the Acmsd gene. The effect of Acmsd gene expression level on murine Interferon-gamma (Ifn-γ) and murine Indoleamine 2,3-dioxygenase 1 (Ido1) gene expression level was investigated by Real-Time PCR. According to the results of this study, good transfection efficiency was observed 72 h after transfection, and Acmsd expression level increased 29-fold (P < 0.001) in transfected LPS-stimulated cells compared to the control group (LPS-stimulated cells that were not transfected). Additionally, increased Acmsd expression level significantly down-regulated Ifn-γ (P < 0.001) and Ido1 (P < 0.01) expression level in transfected LPS-stimulated cells compared to LPS-stimulated cells. CONCLUSIONS: Acmsd gene overexpression in inflammatory conditions can reduce the expression levels of the Ido1 gene, and its regulator, Ifn-γ. Consequently, it may be considered as a novel regulatory factor in the KP balance.
Authors: Michael D Lovelace; Bianca Varney; Gayathri Sundaram; Matthew J Lennon; Chai K Lim; Kelly Jacobs; Gilles J Guillemin; Bruce J Brew Journal: Neuropharmacology Date: 2016-03-16 Impact factor: 5.250
Authors: Michael D Lovelace; Bianca Varney; Gayathri Sundaram; Nunzio F Franco; Mei Li Ng; Saparna Pai; Chai K Lim; Gilles J Guillemin; Bruce J Brew Journal: Front Immunol Date: 2016-08-04 Impact factor: 7.561