Therapeutic Potential of a Combination of Two Gene-Specific Small Interfering RNAs against Clinical Strains of Acanthamoeba

Author:

Lorenzo-Morales Jacob12,Martín-Navarro Carmen M.1,López-Arencibia Atteneri1,Santana-Morales María A.1,Afonso-Lehmann Raquel N.1,Maciver Sutherland K.2,Valladares Basilio1,Martínez-Carretero Enrique1

Affiliation:

1. University Institute of Tropical Diseases and Public Health of the Canary Islands, University of La Laguna, Avda. Astrofísico Fco. Sánchez, S/N 38203 La Laguna, Tenerife, Canary Islands, Spain

2. Centre for Integrative Physiology, School of Biomedical Sciences, University of Edinburgh, Hugh Robson Building, George Square, Edinburgh EH8 9XD, United Kingdom

Abstract

ABSTRACT Pathogenic strains of the genus Acanthamoeba are causative agents of severe infections, such as fatal encephalitis and a sight-threatening amoebic keratitis. Antimicrobial therapy for these infections is generally empirical, and patient recovery is often problematic, due to the existence of a highly resistant cyst stage in these amoebae. In previous studies, small interfering RNAs (siRNAs) against the catalytic domains of extracellular serine proteases and glycogen phosphorylase from Acanthamoeba were designed and evaluated for future therapeutic use. The silencing of proteases resulted in Acanthamoeba failing to degrade human corneal cells, and silencing of glycogen phosphorylase caused amoebae to be unable to form mature cysts. After the siRNA design and concentration were optimized in order to avoid toxicity problems, cultures of Acanthamoeba were treated with a combination of both siRNAs, and cells were evaluated under an inverted microscope. This siRNA-based treatment dramatically affected the growth rate and cellular survival of the amoebae. These results were observed less than 48 h after the initiation of the treatment. In order to check possible toxic effects of the siRNA combination, three eukaryotic cell lines (HeLa, murine macrophages, and osteosarcoma cells) were treated with the same molecules, and cytotoxicity was examined by measuring lactate dehydrogenase release. The future use of the combination of these siRNAs is proposed as a potential therapeutic approach against pathogenic strains of Acanthamoeba .

Publisher

American Society for Microbiology

Subject

Infectious Diseases,Pharmacology (medical),Pharmacology

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