Development of a Novel Ex-vivo 3D Model to Screen Amoebicidal Activity on Infected Tissue

Nancy Elena Guzmán-Delgado,Javier Morán-Martínez,G M M Groothuis,Gildardo Rivera,Nadia Denys Betancourt-Martínez,Sara García-Davis,Pilar Carranza-Rosales,I A M De Graaf,Irma Edith Carranza-Torres

doi:10.1038/s41598-019-44899-5

Abstract

Amoebiasis is a parasitic disease that causes thousands of deaths every year, its adverse effects and resistance to conventional treatments have led to the search of new treatment options, as well as the development of novel screening methods. In this work, we implemented a 3D model of intestine and liver slices from hamsters that were infected ex vivo with virulent E. histolytica trophozoites. Results show preserved histology in both uninfected tissues as well as ulcerations, destruction of the epithelial cells, and inflammatory reaction in intestine slices and formation of micro abscesses, and the presence of amoebae in the sinusoidal spaces and in the interior of central veins in liver slices. The three chemically synthetized compounds T-001, T-011, and T-016, which act as amoebicides in vitro, were active in both infected tissues, as they decreased the number of trophozoites, and provoked death by disintegration of the amoeba, similar to metronidazole. However, compound T-011 induced signs of cytotoxicity to liver slices. Our results suggest that ex vivo cultures of precision-cut intestinal and liver slices represent a reliable 3D approach to evaluate novel amoebicidal compounds, and to simultaneously detect their toxicity, while reducing the number of experimental animals commonly required by other model systems.

Highlights

On the other hand, in recent years the interest in designing and synthetizing new antiparasitic molecules has been renewed, facilitated by the application of in silico docking; by using such techniques several research groups have investigated newly synthetized molecules as potential antiprotozoal compounds, reporting active molecules against E. histolytica and other protozoa[11,12]
T-001, T-011, and T-016 were effective against E. histolytica, with IC50 values < 2 μM
E. histolytica is an enteric protozoan that causes human amoebiasis, which is considered the fourth cause of parasitic diseases[23,24]

Summary

Introduction

In recent years the interest in designing and synthetizing new antiparasitic molecules has been renewed, facilitated by the application of in silico docking; by using such techniques several research groups have investigated newly synthetized molecules as potential antiprotozoal compounds, reporting active molecules against E. histolytica and other protozoa[11,12]. Our group reported that hamster liver slices infected with E. histolytica represent an ex vivo 3D model to study hepatic amoebiasis based on morphological and molecular characterization of the disease ex vivo[18,19] In these studies, we observed that morphologic characteristics in amoebae were conserved at the evaluated experimental times, as were eritophagocytic activity, immune response reaction, and virulence factors induction. Infected precision-cut hamster liver and intestinal slices were used to analyze anti-amoebic activity, and at the same time, the possible toxicity of the amoebicidal molecules T-001, T-011, and T-016 in the infected tissue, in comparison to metronidazole as a reference compound This experimental approach allowed us to investigate effectivity of the tested compounds against E. histolytica in tissue, and demonstrates the feasibility of the model for the research of new antiprotozoal drugs

Methods

Results

Conclusion