Extracellular vesicles isolated from Toxoplasma gondii induce host immune response

Abstract

This study established a protocol to purify Toxoplasma gondii tachyzoite microvesicles and exosomes, called as extracellular vesicles (EVs). In addition, the investigations were conducted to determine the kinetic of EV release by tachyzoites and whether EV proteins are able to modulate the host immune response. The particle size and concentration released by tachyzoites in culture medium at different incubation-period were characterized by nanoparticle tracking analysis. Tachyzoites (1 × 10 ) released around 4.37 ± 0.81 × 10 EVs/mL/h, with size varying between 138.2 and 171.9 nm. EVs released into the medium were purified by gel-exclusion chromatography and screened by ELISA, using a pool of human positive sera for toxoplasmosis. EV-fractions contained high concentration of proteins, and EVs were analyzed by scanning and transmission electron microscopies. Tachyzoites released EVs into the culture medium throughout all membrane surface, and these vesicles contain small RNAs/miRNA. Pooled sera from chronically infected human or mice (infected with 2 different T. gondii strains) recognized distinct EV electrophoretic patterns in immunoblotting. T. gondii EVs significantly induced IL-10, TNF-α and iNOS in murine macrophages. In conclusion, this study shows that T. gondii secrete/excrete EVs (microvesicles and exosomes) contain miRNA and they were immunologically recognized by host immune response.

Human extracellular vesicles and correlation with two clinical forms of toxoplasmosis

“…At the same time, T. gondii EVs contain specific proteins as surface antigens, which are release by tachyzoites and compose the excreted-secreted antigens. These proteins participate of tachyzoite invasion, replication within host cells and are recognized by host immune system [29,31,43,44].…”

“…Human EVs were previously fixed in 2% paraformaldehyde/PBS (v:v) for 1 hour. One drop of suspension was put on EM grid and performed by negative staining technique with 2% potassium phosphotungstate at pH 6,8, as previously described [31]. Grids were observed under a JEOL Transmission Electron Microscope (model JEM1011) (JEOL/Massachusetts/USA) operating at 80 kV.…”

“…Regarding on T. gondii, proteomic profile of exosomes containing a wide range of proteins [29]. T. gondii derived-EVs (exosomes and microvesicles) contain miRNA and they were immunologically recognized by host immune response inducing humoral and cellular responses as IL-10, TNF-α, and iNOS [30,31]. Exosomes excreted by infected host cells constituted an effective non-cellular vaccine with antigen and adjuvant properties for the immune response as well as, immunization [32,33].…”

Role of Extracellular Vesicles in Cellular Cross Talk in Malaria

“…EVs from T. gondii also carry nucleic acids including mRNAs and small RNAs, such as miRNA. The miRNAs inside EVs act as potential immune regulatory mediators involved in the host cell manipulation (132). Treatment of macrophages with T. gondii EVs leads to the upregulation on one hand of the anti-inflammatory IL-10 and, on the other hand, the pro-inflammaotry TNF-α, IFNγ, and iNOS.…”

Perils and Promises of Pathogenic Protozoan Extracellular Vesicles

“…Also, some PPEV-encapsulated biomolecules are classical EV markers (Mantel et al, 2013), immune modulators, mediators of intracellular signaling, host-parasite interactions, membrane fusion, transporters, and oxidation-reduction processes (Geiger et al, 2010;Moreira et al, 2019). However, the functions of PPEVs rely on preparation, time of reaction, temperature, pH and most importantly, species and strain of origin (Twu et al, 2013;Montaner et al, 2014;Nogueira et al, 2015;Silva et al, 2018;Moreira et al, 2019) (Table 1). PPEV bioactive molecules immensely contribute to parasite development, and it is likely that protozoan parasites secrete biochemically different EVs at every developmental stage so as to adapt to a changing environment as exemplified by L. infatum which has significant enrichment of ribosomal and RNA transport proteins during the parasite growth at the log phase but an abundance of cellular processes and oxidative phosphorylation proteins at the stationary phase (Santarém et al, 2013).…”