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Functional evaluation of gene silencing on macrophages derived from U937 cells using interference RNA (shRNA) in a model of macrophages infected with Leishmania (Viannia) braziliensis

Published online by Cambridge University Press:  07 October 2015

CLEMENCIA OVALLE-BRACHO ,
DIANA A. LONDOÑO-BARBOSA ,
CARLOS FRANCO-MUÑOZ  and
CARLOS CLAVIJO-RAMÍREZ
CLEMENCIA OVALLE-BRACHO*
Affiliation:
Centro Dermatológico Federico Lleras Acosta, Avenida 1 # 13ª-61, Bogotá D.C., Colombia Facultad de Medicina, Universidad Nacional de Colombia, Carrera 45 # 26-85, Bogotá D.C., Colombia
DIANA A. LONDOÑO-BARBOSA
Affiliation:
Centro Dermatológico Federico Lleras Acosta, Avenida 1 # 13ª-61, Bogotá D.C., Colombia
CARLOS FRANCO-MUÑOZ
Affiliation:
Centro Dermatológico Federico Lleras Acosta, Avenida 1 # 13ª-61, Bogotá D.C., Colombia
CARLOS CLAVIJO-RAMÍREZ
Affiliation:
Departamento de Biología, Universidad Nacional de Colombia, Carrera 45 # 26-85, Bogotá D.C., Colombia
*
*Corresponding Author: Clemencia Ovalle-Bracho, Centro Dermatológico Federico Lleras Acosta, Avenida 1 # 13ª-61, Bogotá D.C., Colombia. E-mail: clemovalle@gmail.com
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Summary

Leishmaniasis development is multifactorial; nonetheless, the establishment of the infection, which occurs by the survival and replication of the parasite inside its main host cell, the macrophage, is mandatory. Thus, the importance of studying the molecular mechanisms involved in the Leishmania–macrophage interaction is highlighted. The aim of this study was to characterize a cellular model of macrophages derived from U937 cells that would allow for the identification of infection phenotypes induced by genetic silencing with interference RNA in the context of macrophages infected with Leishmania (Viannia) braziliensis. The model was standardized by silencing an exogenous gene (gfp), an endogenous gene (lmna) and a differentially expressed gene between infected and non-infected macrophages (gro-β). The silencing process was successful for the three genes studied, obtaining reductions of 88·9% in the GFP levels, 87·5% in LMNA levels and 74·4% for Gro-β with respect to the corresponding control cell lines. The cell model revealed changes in the infection phenotype of the macrophages in terms of number of amastigotes per infected macrophage, number of amastigotes per sampled macrophage and percentage of infected macrophages as a result of gene silencing. Thus, this cell model constitutes a research platform for the study of parasite–host interactions and for the identification of potentially therapeutic targets.

Keywords

Leishmaniamonocyte-derived macrophagesgene silencingRNAsmall interferingU937 cells
Type
Research Article
Information
Parasitology , Volume 142 , Issue 14 , December 2015 , pp. 1682 - 1692
Copyright
Copyright © Cambridge University Press 2015 

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