Dataset Information

Extracellular Vesicles from the Protozoa Acanthamoeba castellanii: Their Role in Pathogenesis, Environmental Adaptation and Potential Applications.

ABSTRACT: Extracellular vesicles (EVs) are membranous compartments of distinct cellular origin and biogenesis, displaying different sizes and include exosomes, microvesicles, and apoptotic bodies. The EVs have been described in almost every living organism, from simple unicellular to higher evolutionary scale multicellular organisms, such as mammals. Several functions have been attributed to these structures, including roles in energy acquisition, cell-to-cell communication, gene expression modulation and pathogenesis. In this review, we described several aspects of the recently characterized EVs of the protozoa Acanthamoeba castellanii, a free-living amoeba (FLA) of emerging epidemiological importance, and compare their features to other parasites' EVs. These A. castellanii EVs are comprised of small microvesicles and exosomes and carry a wide range of molecules involved in many biological processes like cell signaling, carbohydrate metabolism and proteolytic activity, such as kinases, glucanases, and proteases, respectively. Several biomedical applications of these EVs have been proposed lately, including their use in vaccination, biofuel production, and the pharmaceutical industry, such as platforms for drug delivery.

SUBMITTER: Goncalves DS

PROVIDER: S-EPMC6466093 | biostudies-literature | 2019 Feb

REPOSITORIES: biostudies-literature

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Extracellular Vesicles from the Protozoa <i>Acanthamoeba castellanii</i>: Their Role in Pathogenesis, Environmental Adaptation and Potential Applications.

Gonçalves Diego de Souza DS Ferreira Marina da Silva MDS Guimarães Allan J AJ

Bioengineering (Basel, Switzerland) 20190201 1

Extracellular vesicles (EVs) are membranous compartments of distinct cellular origin and biogenesis, displaying different sizes and include exosomes, microvesicles, and apoptotic bodies. The EVs have been described in almost every living organism, from simple unicellular to higher evolutionary scale multicellular organisms, such as mammals. Several functions have been attributed to these structures, including roles in energy acquisition, cell-to-cell communication, gene expression modulation and ...[more]

PMID: 30717103

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Project description:We conducted a comprehensive comparative analysis of extracellular vesicles (EVs) from two Acanthamoeba castellanii strains, Neff (environmental) and T4 (clinical). Morphological analysis via transmission electron microscopy revealed slightly larger Neff EVs (average = 194.5 nm) compared to more polydisperse T4 EVs (average = 168.4 nm). Nanoparticle tracking analysis (NTA) and dynamic light scattering validated these differences. Proteomic analysis of the EVs identified 1,352 proteins, with 1,107 common, 161 exclusive in Neff, and 84 exclusively in T4 EVs. Gene ontology and Kyoto Encyclopedia of Genes and Genomes (KEGG) mapping revealed distinct molecular functions and biological processes and notably, the T4 EVs enrichment in serine proteases, aligned with its pathogenicity. Lipidomic analysis revealed a prevalence of unsaturated lipid species in Neff EVs, particularly triacylglycerols, phosphatidylethanolamines (PEs), and phosphatidylserine, while T4 EVs were enriched in diacylglycerols and diacylglyceryl trimethylhomoserine, phosphatidylcholine and less unsaturated PEs, suggesting differences in lipid metabolism and membrane permeability. Metabolomic analysis indicated Neff EVs enrichment in glycerolipid metabolism, glycolysis, and nucleotide synthesis, while T4 EVs, methionine metabolism. Furthermore, RNA-seq of EVs revealed differential transcript between the strains, with Neff EVs enriched in transcripts related to gluconeogenesis and translation, suggesting gene regulation and metabolic shift, while in the T4 EVs transcripts were associated with signal transduction and protein kinase activity, indicating rapid responses to environmental changes. In this novel study, data integration highlighted the differences in enzyme profiles, metabolic processes, and potential origins of EVs in the two strains shedding light on the diversity and complexity of A. castellanii EVs and having implications for understanding host-pathogen interactions and developing targeted interventions for Acanthamoeba-related diseases.IMPORTANCEA comprehensive and fully comparative analysis of extracellular vesicles (EVs) from two Acanthamoeba castellanii strains of distinct virulence, a Neff (environmental) and T4 (clinical), revealed striking differences in their morphology and protein, lipid, metabolites, and transcripts levels. Data integration highlighted the differences in enzyme profiles, metabolic processes, and potential distinct origin of EVs from both strains, shedding light on the diversity and complexity of A. castellanii EVs, with direct implications for understanding host-pathogen interactions, disease mechanisms, and developing new therapies for the clinical intervention of Acanthamoeba-related diseases.

Project description:BackgroundAcanthamoeba species are the causative agents of fatal granulomatous encephalitis in humans. Haematogenous spread is thought to be a primary step, followed by blood-brain barrier penetration, in the transmission of Acanthmaoeba into the central nervous system, but the associated molecular mechanisms remain unclear. Here, we evaluated the role of Src, a non-receptor protein tyrosine kinase in the biology and pathogenesis of Acanthamoeba.MethodsAmoebistatic and amoebicidal assays were performed by incubating amoeba in the presence of Src kinase-selective inhibitor, PP2 (4-amino-5-(4-chlorophenyl)-7-(t-butyl)pyrazolo[3,4-d]pyrimidine) and its inactive analog, PP3 (4-amino-7-phenylpyrazolo[3,4-d]pyrimidine). Using this inhibitor, the role of Src kinase in A. castellanii interactions with Escherichia coli was determined. Zymographic assays were performed to study effects of Src kinase on extracellular proteolytic activities of A. castellanii. The human brain microvascular endothelial cells were used to determine the effects of Src kinase on A. castellanii adhesion to and cytotoxicity of host cells.ResultsInhibition of Src kinase using a specific inhibitor, PP2 (4-amino-5-(4 chlorophenyl)-7-(t-butyl)pyrazolo [3,4-d] pyrimidine) but not its inactive analog, PP3 (4-amino-7-phenylpyrazolo[3,4-d] pyrimidine), had detrimental effects on the growth of A. castellanii (keratitis isolate, belonging to the T4 genotype). Interestingly, inhibition of Src kinase hampered the phagocytic ability of A. castellanii, as measured by the uptake of non-invasive bacteria, but, on the contrary, invasion by pathogenic bacteria was enhanced. Zymographic assays revealed that inhibition of Src kinases reduced extracellular protease activities of A. castellanii. Src kinase inhibition had no significant effect on A. castellanii binding to and cytotoxicity of primary human brain microvascular endothelial cells, which constitute the blood-brain barrier.ConclusionsFor the first time, these findings demonstrated that Src kinase is involved in A. castellanii proliferation, protease secretions and phagocytic properties. Conversely, invasion of Acanthamoeba by pathogenic bacteria was stimulated by Src kinase inhibition.

Dataset Information

Extracellular Vesicles from the Protozoa Acanthamoeba castellanii: Their Role in Pathogenesis, Environmental Adaptation and Potential Applications.

Publications

Extracellular Vesicles from the Protozoa <i>Acanthamoeba castellanii</i>: Their Role in Pathogenesis, Environmental Adaptation and Potential Applications.

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