Evento
Design of a lentiviral vector as a therapeutic strategy against alzheimer´s disease
Abrey Recalde, Maria Jimena
; Gonzalez Hermida, Paula; Baez, Veronica; Jerusalinsky, Diana Alicia
; Frecha, Cecilia Ariana
Tipo del evento:
Reunión
Nombre del evento:
LXI Reunión anual de la Sociedad Argentina de Investigación Clinica; LXVI, Reunión anual de la Sociedad Argentina de Inmunología; XLVIII Reunión Anual de la Sociedad Argentina de Farmacología Experimental; VII Reunión Anual de la Sociedad Argentina de Nanomedicina y V Congreso Nacional de la Asociación Argentina de Ciencia y Tecnología de Animales de Laboratorio
Fecha del evento:
15/11/2016
Institución Organizadora:
Sociedad Argentina de Investigación Clinica;
Sociedad Argentina de Inmunología;
Sociedad Argentina de Farmacología Experimental;
Sociedad Argentina de Nanomedicina;
Asociación Argentina de ciencia y tecnología de animales de Laboratorio;
Título de la revista:
Medicina
Editorial:
Fundacion Revista Medicina
Idioma:
Inglés
Clasificación temática:
Resumen
Alzheimer’s disease (AD) is a chronic neurodegenerative disorder characterized by a progressive loss of cognitive functions. One of the hallmarks is the formation of amyloid plaques, composed mainly by Aß peptide oligomers (AβOs). Neprilysin (NEP) is the most important endopeptidase and crucial for the degradation of Aß in the brain, avoiding amyloid plaques formation. Because NEP is decreased in brains of patients with AD, we aim to develop a lentiviral vector (LV) capable of specifically expressing in hippocampal neurons, to study its role in AD and therefore its possible therapeutic function.First, a construct containing the complete cDNA of NEP downstream of the hippocampal-specific promoter human synapsin (SYN-1) was performed. NEP cDNA was obtained from pBOB-NEP plasmid and was cloned under the SYN-1 promoter to obtain SYN-NEP plasmid. SYN-NEP also contains the ubiquitous CMV promoter, located outside the sequence to be packaged in LV. The correct cloning was checked by BamHI/KpnI digestion followed by 1% agarose gel electrophoresis and was verified by sequencing. To test NEP expression under the CMV promoter, 293T cells were transfected with SYN-NEP. SYN-RFP plasmid expressing the reporter red fluorescent protein (RFP) and pBOB-NEP were used as transfection and positive controls, respectively. After 48 hours NEP expression was evaluated by western blot (WB) and RFP by fluorescence microscopy.Electrophoresis of SYN-NEP digestion resulted in two bands of 3392pb and 7644pb, which coincided with the molecular weights of the insert containing NEP and backbone, respectively. This in turn was validated by sequencing. Transfection efficiency calculated by expression of RFP was 80%. WB showed a 85kDa band only in those lanes corresponding to transfection with pBOB-NEP and SYN-NEP.In conclusion, NEP was successfully cloned downstream of SYN-1 promoter and is expressed correctly under a ubiquitous promoter. This construction is the first step for the production of LV.
Palabras clave:
Alzheimer's disease
,
NEPYLISIN
,
LENTIVIRAL VECTORS
,
SYNAPSIN PROMOTER
Archivos asociados
Licencia
Identificadores
Colecciones
Eventos (IMTIB)
Eventos de INSTITUTO DE MEDICINA TRASLACIONAL E INGENIERIA BIOMEDICA
Eventos de INSTITUTO DE MEDICINA TRASLACIONAL E INGENIERIA BIOMEDICA
Eventos(SEDE CENTRAL)
Eventos de SEDE CENTRAL
Eventos de SEDE CENTRAL
Citación
Design of a lentiviral vector as a therapeutic strategy against alzheimer´s disease; LXI Reunión anual de la Sociedad Argentina de Investigación Clinica; LXVI, Reunión anual de la Sociedad Argentina de Inmunología; XLVIII Reunión Anual de la Sociedad Argentina de Farmacología Experimental; VII Reunión Anual de la Sociedad Argentina de Nanomedicina y V Congreso Nacional de la Asociación Argentina de Ciencia y Tecnología de Animales de Laboratorio; Argentina; 2016; 110-111
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