Polarisation of human macrophages towards an M1 subtype triggered by an atypical Brazilian strain of Toxoplasma gondii results in a reduction in parasite burden.

Toxoplasma gondii Nicolle et Manceaux, 1909, the etiologic agent of toxoplasmosis, was considered a clonal population with three distinct genetic lineages (I, II and III); however, sequence analysis of different strains has revealed distinct atypical genotypes. Macrophages are essential for immunity against toxoplasmosis and differential cell regulation may affect the course of the disease. In this context, our study aims to investigate the infection by TgChBrUD2, a highly virulent atypical Brazilian strain of T. gondii, on the activation and polarisation of human macrophages. Human macrophage-like cells obtained from THP-1 cells were infected with TgChBrUD2, RH or ME49 strains of T. gondii to evaluate the impact of parasite infection on macrophage polarisation. Our results indicate that the TgChBrUD2 and ME49 strains of T. gondii induced a classic activation of human macrophages, which was confirmed by the high rate of spindle-shaped macrophages, low amount of urea and increase in the levels of nitrite, as well as the down-regulation of M2-markers. In contrast, RH strain promoted an alternative activation of macrophages. The polarisation of human macrophages towards an M1 subtype mediated by TgChBrUD2 and ME49 strains resulted in a low parasite burden, with high levels of IL-6 and MIF. Finally, the M2 subtype triggered by the RH strain culminated in a lower intracellular proliferation index. We concluded that the atypical (TgChBrUD2) and clonal (ME49) strains are able to elicit an M1 subtype, which results in parasitism control, partially explained by the high levels of IL-6 and MIF produced during the infection by these genotypes. In contrast, the clonal (RH) strain promoted a macrophage polarisation towards an M2 subtype, marked by a high parasite burden, with a weak modulation of pro-inflammatory cytokines. Thus, atypical strains can present different mechanisms of pathogenicity and transmissibility compared to clonal strains, as well as they can use distinct strategies to evade the host's immune response and ensure their survival.

BEWO trophoblast cells and Toxoplasma gondii infection modulate cell death mechanisms in THP-1 monocyte cells by interference in the expression of death receptor and intracellular proteins.

Crosstalk between trophoblast and monocytes is essential for gestational success, and it can be compromised in congenital toxoplasmosis. Cell death is one of the mechanisms involved in the maintenance of pregnancy, and this study aimed to evaluate the role of trophoblast in the modulation of monocyte cell death in the presence or absence of Toxoplasma gondii infection. THP-1 cells were stimulated with supernatants of BeWo cells and then infected or not with T. gondii. The supernatants were collected and analyzed for the secretion of human Fas ligand, and cells were used to determine cell death and apoptosis, cell death receptor, and intracellular proteins expression. Cell death and apoptosis index were higher in uninfected THP-1 cells stimulated with supernatants of BeWo cells; however, apoptosis index was reduced by T. gondii infection. Macrophage migration inhibitory factor (MIF) and transforming growth factor (TGF)-ß1, secreted by BeWo cells, altered the cell death and apoptosis rates in THP-1 cells. In infected THP-1 cells, the expression of Fas/CD95 and secretion of FasL was significantly higher; however, caspase 3 and phosphorylated extracellular-signal-regulated kinase (ERK1/2) were downregulated. Results suggest that soluble factors secreted by BeWo cells induce cell death and apoptosis in THP-1 cells, and Fas/CD95 can be involved in this process. On the other hand, T. gondii interferes in the mechanism of cell death and inhibits THP-1 cell apoptosis, which can be associated with active caspase 3 and phosphorylated ERK1/2. In conclusion, our results showed that human BeWo trophoblast cells and T. gondii infection modulate cell death in human THP-1 monocyte cells.

Biogenic Silver Nanoparticles Can Control Toxoplasma gondii Infection in Both Human Trophoblast Cells and Villous Explants.

The combination of sulfadiazine and pyrimethamine plus folinic acid is the conventional treatment for congenital toxoplasmosis. However, this classical treatment presents teratogenic effects and bone marrow suppression. In this sense, new therapeutic strategies are necessary to reduce these effects and improve the control of infection. In this context, biogenic silver nanoparticles (AgNp-Bio) appear as a promising alternative since they have antimicrobial, antiviral, and antiparasitic activity. The purpose of this study to investigate the action of AgNp-Bio in BeWo cells, HTR-8/SVneo cells and villous explants and its effects against Toxoplasma gondii infection. Both cells and villous explants were treated with different concentrations of AgNp-Bio or combination of sulfadiazine + pyrimethamine (SDZ + PYZ) in order to verify the viability. After, cells and villi were infected and treated with AgNp-Bio or SDZ + PYZ in different concentrations to ascertain the parasite proliferation and cytokine production profile. AgNp-Bio treatment did not reduce the cell viability and villous explants. Significant reduction was observed in parasite replication in both cells and villous explants treated with silver nanoparticles and classical treatment. The AgNp-Bio treatment increased of IL-4 and IL-10 by BeWo cells, while HTR8/SVneo cells produced macrophage migration inhibitory factor (MIF) and IL-4. In the presence of T. gondii, the treatment induced high levels of MIF production by BeWo cells and IL-6 by HTR8SV/neo. In villous explants, the AgNp-Bio treatment downregulated production of IL-4, IL-6, and IL-8 after infection. In conclusion, AgNp-Bio can decrease T. gondii infection in trophoblast cells and villous explants. Therefore, this treatment demonstrated the ability to reduce the T. gondii proliferation with induction of inflammatory mediators in the cells and independent of mediators in chorionic villus which we consider the use of AgNp-Bio promising in the treatment of toxoplasmosis in BeWo and HTR8/SVneo cell models and in chorionic villi.

Experimental models of maternal-fetal interface and their potential use for nanotechnology applications.

During pregnancy, the placenta regulates the transfer of oxygen, nutrients, and residual products between the maternal and fetal bloodstreams and is a key determinant of fetal exposure to xenobiotics from the mother. To study the disposition of substances through the placenta, various experimental models are used, especially the perfused placenta, placental villi explants, and cell lineage models. In this context, nanotechnology, an area of study that is on the rise, enables the creation of particles on nanometric scales capable of releasing drugs aimed at specific tissues. An important reason for furthering the studies on transplacental transfer is to explore the potential of nanoparticles (NPs), in new delivery strategies for drugs that are specifically aimed at the mother, the placenta, or the fetus and that involve less toxicity. Due to the fact that the placental barrier is essential for the interaction between the maternal and fetal organisms as well as the possibility of NPs being used in the treatment of various pathologies, the aim of this review is to present the main experimental models used in studying the maternal-fetal interaction and the action of NPs in the placental environment.

Increased Toxoplasma gondii Intracellular Proliferation in Human Extravillous Trophoblast Cells (HTR8/SVneo Line) Is Sequentially Triggered by MIF, ERK1/2, and COX-2.

Macrophage migration inhibitory factor (MIF) is a potent pro-inflammatory cytokine, which mediates the regulation of diverse cellular functions. It is produced by extravillous trophoblastic cells and has been found to be involved in the pathogenesis of diseases caused by some protozoa, including Toxoplasma gondii. Previous studies demonstrated the ability of T. gondii to take advantage of MIF action in human trophoblast cells. However, MIF action in T. gondii-infected extravillous trophoblastic cells (HTR8/SVneo cell line) has not been fully investigated. The present study aimed to investigate the role of MIF in T. gondii-infected HTR8/SVneo cells and verify the intracellular signaling pathways triggered by this cytokine. We found that T. gondii increased MIF production by HTR8/SVneo cells, and by contrast, MIF inhibition, by ISO-1, led to a significant decrease in T. gondii proliferation and CD74 expression in HTR8/SVneo cells. Moreover, in infected HTR8/SVneo cells, the addition of recombinant MIF (rMIF) increased CD44 co-receptor expression, ERK1/2 phosphorylation, COX-2 expression, and IL-8 production, which favored T. gondii proliferation. Our findings indicate that T. gondii can use MIF to modulate important factors in HTR8/SVneo cells, being a possible explanation for the higher susceptibility of extravillous trophoblast cells than other trophoblast cell populations.

Cyclooxygenase (COX)-2 Inhibitors Reduce Toxoplasma gondii Infection and Upregulate the Pro-inflammatory Immune Response in Calomys callosus Rodents and Human Monocyte Cell Line.

Toxoplasma gondii is able to infect a wide range of vertebrates, including humans. Studies show that cyclooxygenase-2 (COX-2) is a modulator of immune response in multiple types of infection, such as Trypanosoma cruzi. However, the role of COX-2 during T. gondii infection is still unclear. The aim of this study was to investigate the role of COX-2 during infection by moderately or highly virulent strains of T. gondii in Calomys callosus rodents and human THP-1 cells. C. callosus were infected with 50 cysts of T. gondii (ME49), treated with COX-2 inhibitors (meloxicam or celecoxib) and evaluated to check body weight and morbidity. After 40 days, brain and serum were collected for detection of T. gondii by real-time PCR and immunohistochemistry or cytokines by CBA. Furthermore, peritoneal macrophages or THP-1 cells, infected with RH strain or uninfected, were treated with meloxicam or celecoxib to evaluate the parasite proliferation by colorimetric assay and cytokine production by ELISA. Finally, in order to verify the role of prostaglandin E2 in COX-2 mechanism, THP-1 cells were infected, treated with meloxicam or celecoxib plus PGE2, and analyzed to parasite proliferation and cytokine production. The data showed that body weight and morbidity of the animals changed after infection by T. gondii, under both treatments. Immunohistochemistry and real-time PCR showed a reduction of T. gondii in brains of animals treated with both COX-2 inhibitors. Additionally, it was observed that both COX-2 inhibitors controlled the T. gondii proliferation in peritoneal macrophages and THP-1 cells, and the treatment with PGE2 restored the parasite growth in THP-1 cells blocked to COX-2. In the serum of Calomys, upregulation of pro-inflammatory cytokines was detected, while the supernatants of peritoneal macrophages and THP-1 cells demonstrated significant production of TNF and nitrite, or TNF, nitrite and MIF, respectively, under both COX-2 inhibitors. Finally, PGE2 treatment in THP-1 cells triggered downmodulation of pro-inflammatory mediators and upregulation of IL-8 and IL-10. Thus, COX-2 is an immune mediator involved in the susceptibility to T. gondii regardless of strain or cell types, since inhibition of this enzyme induced control of infection by upregulating important pro-inflammatory mediators against Toxoplasma.