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Development of an in vitro media perfusion model of Leishmania major macrophage infection

O'Keeffe, A; Hyndman, L; McGinty, S; Riezk, A; Murdan, S; Croft, SL; (2019) Development of an in vitro media perfusion model of Leishmania major macrophage infection. PLoS One , 14 (7) , Article e0219985. 10.1371/journal.pone.0219985. Green open access

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Abstract

BACKGROUND: In vitro assays are widely used in studies on pathogen infectivity, immune responses, drug and vaccine discovery. However, most in vitro assays display significant differences to the in vivo situation and limited predictive properties. We applied medium perfusion methods to mimic interstitial fluid flow to establish a novel infection model of Leishmania parasites. METHODS: Leishmania major infection of mouse peritoneal macrophages was studied within the Quasi Vivo QV900 macro-perfusion system. Under a constant flow of culture media at a rate of 360μl/min, L. major infected macrophages were cultured either at the base of a perfusion chamber or raised on 9mm high inserts. Mathematical and computational modelling was conducted to estimate medium flow speed, shear stress and oxygen concentration. The effects of medium flow on infection rate, intracellular amastigote division, macrophage phagocytosis and macropinocytosis were measured. RESULTS: Mean fluid speeds at the macrophage cell surface were estimated to be 1.45 x 10^{-9} m/s and 1.23 x 10^{-7} m/s for cells at the base of the chamber and cells on an insert, respectively. L. major macrophage infection was significantly reduced under both media perfusion conditions compared to cells maintained under static conditions; a 85±3% infection rate of macrophages at 72 hours in static cultures compared to 62±5% for cultures under slow medium flow and 55±3% under fast medium flow. Media perfusion also decreased amastigote replication and both macrophage phagocytosis (by 44±4% under slow flow and 57±5% under fast flow compared with the static condition) and macropinocytosis (by 40±4% under slow flow and 62±5% under fast flow compared with the static condition) as measured by uptake of latex beads and pHrodo Red dextran. CONCLUSIONS: Perfusion of culture medium in an in vitro L. major macrophage infection model (simulating in vivo lymphatic flow) reduced the infection rate of macrophages, the replication of the intracellular parasite, macrophage phagocytosis and macropinocytosis with greater reductions achieved under faster flow speeds.

Type: Article
Title: Development of an in vitro media perfusion model of Leishmania major macrophage infection
Location: United States
Open access status: An open access version is available from UCL Discovery
DOI: 10.1371/journal.pone.0219985
Publisher version: https://doi.org/10.1371/journal.pone.0219985
Language: English
Additional information: © 2019 O’Keeffe et al. This is an open access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/4.0/).
Keywords: Macrophages, Parasitic diseases, Fluid flow, Oxygen, Phagocytosis, Flow rate, Shear stresses, Amastigotes
UCL classification: UCL
UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences
UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Life Sciences
UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Life Sciences > UCL School of Pharmacy
UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Life Sciences > UCL School of Pharmacy > Pharmaceutics
URI: https://discovery.ucl.ac.uk/id/eprint/10078941
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