The ethanolic extract of the fungus Trichoderma stromaticum decreases the Toxoplasma gondii replication in vitro and ameliorates the experimental toxoplasmosis in vivo.

Trichoderma are fungi that are well-known to inhibit the growth of a variety of plant pathogens. Currently, there is an increasing search for new drugs to treat toxoplasmosis. The aims of this study were to investigate the effect of ExtTs in the control of Toxoplasma gondii proliferation in vitro and the course of toxoplasmosis in a mouse model. Firstly, the cytotoxicity of the ExtTs was evaluated by cultivating macrophages with different concentrations of the extract and cell viability was assessed by the MTT assay. Next, the infectivity of the T. gondii treated with extract was analyzed by infecting J774 macrophages. To evaluate the effect of the ExtTs in vivo, C57BL/6 mice were infected orally with T. gondii, ME-49, treated daily with ExtTs, and clinical, biochemical and histological changes were monitored. It was demonstrated that the extract did not affect the host cellular viability and, the treatment of parasites with ExtTs altered their morphology and decreased their ability to proliferate inside macrophages. Additionally, the treatment of mice with ExtTs decreased the parasitism and inflammation in the small intestine and liver of infected mice in parallel with increased IL-10/TNF ratio systemically and prevented alterations to serum VLDL and triglyceride levels. Thus, ExtTs could be considered an alternative/complementary therapy to control toxoplasmosis.

Effects of Notch signaling pathway inhibition by dibenzazepine in acute experimental toxoplasmosis.

Notch signaling pathway plays a crucial role in cellular fate across species, being important for the differentiation and development of several cell types. The aim of this study was to evaluate the effect of Notch inhibition pathway by dibenzazepine (DBZ) in histological and inflammatory alterations and, tissue parasitism in acute Toxoplasma gondii infection. For this, C57BL/6 mice were treated with DBZ before infection with T. gondii, and the small intestine, lungs and liver were analyzed. The genes related to Notch signaling pathway were assayed through qPCR in the organs, and cytokine measurement was performed in serum samples. In the small intestine, T. gondii infection impaired the Hes1 and Math1 mRNA expressions, increased the inflammation and decreased goblet and Paneth cell numbers. The DBZ-treatment was able to partially preserve these cells, however, the parasitism and inflammation were not altered. In parallel, the high IL-2, IL-6, TNF and, IFN-γ levels induced by infection were not changed with the DBZ treatment, with the IFN-γ levels even higher. In contrast, in the liver and lungs, the DBZ-treatment diminished parasitism and inflammation. Our results highlight that Notch pathway inhibition in T.gondii infection results in different parasitological and inflammatory outcomes depending on the organ analyzed.

Toxoplasma gondii Infection Decreases Intestinal 5-Lipoxygenase Expression, while Exogenous LTB4 Controls Parasite Growth.

5-Lipoxygenase (5-LO) is an enzyme required for the production of leukotrienes and lipoxins and interferes with parasitic infections. In vitro, Toxoplasma gondii inhibits leukotriene B4 (LTB4) production, and mice deficient in 5-LO are highly susceptible to infection. The aim of this study was to investigate the effects of the pharmacological inhibition of the 5-LO pathway and exogenous LTB4 supplementation during experimental toxoplasmosis. For this purpose, susceptible C57BL/6 mice were orally infected with T. gondii and treated with LTB4 or MK886 (a selective leukotriene inhibitor through inhibition of 5-LO-activating protein [FLAP]). The parasitism, histology, and immunological parameters were analyzed. The infection decreased 5-LO expression in the small intestine, and treatment with MK886 reinforced this reduction during infection; in addition, MK886-treated infected mice presented higher intestinal parasitism, which was associated with lower local interleukin-6 (IL-6), interferon gamma (IFN-γ), and tumor necrosis factor (TNF) production. In contrast, treatment with LTB4 controlled parasite replication in the small intestine, liver, and lung and decreased pulmonary pathology. Interestingly, treatment with LTB4 also preserved the number of Paneth cells and increased α-defensins expression and IgA levels in the small intestine of infected mice. Altogether, these data demonstrated that T. gondii infection is associated with a decrease in 5-LO expression, and on the other hand, treatment with the 5-LO pathway product LTB4 resulted in better control of parasite growth in the organs, adding to the knowledge about the pathogenesis of T. gondii infection.

The imbalance of TNF and IL-6 levels and FOXP3 expression at the maternal-fetal interface is involved in adverse pregnancy outcomes in a susceptible murine model of congenital toxoplasmosis.

Vertical transmission of Toxoplasma gondii leads to adverse pregnancy outcomes depending on the time at which the infection occurs and the immunological state of the mother. C57BL/6 and BALB/c mice have been described as susceptible and resistant mouse lineages to congenital T. gondii infection, respectively. This study aimed to elucidate the systemic and local cytokine profile of pregnant mice infected with T. gondii and whether the expression of the transcription factor FOXP3, related to T regulatory cells, is associated with the resistance/susceptibility of these lineages of mice in the context of experimental congenital toxoplasmosis. For this purpose, C57BL/6 and BALB/c females were orally infected with the T. gondii ME-49 strain on the day of vaginal plug detection or day 14 of gestation, examined 7 or 5 days later, respectively, as models of early and late pregnancy. Cytokine levels were measured systemically and in the uterus/placenta. Additionally, the uterus/placenta were evaluated macroscopically for resorption rates and histologically for parasite and FOXP3 immunostaining. The FOXP3 protein expression was also evaluated by western blotting assay. It was found that, during early pregnancy, the infection leads to high IFN-γ, TNF and IL-6 levels systemically, with the TNF levels being higher in C57BL/6 mice. At the maternal-fetal interface, the infection induced high levels of IFN-γ in both mouse lineages; however, higher levels were observed in BALB/c, while high TNF and IL-6 levels were found in C57BL/6, but not in BALB/c mice. In contrast, in late gestation, T. gondii interfered less strongly with the cytokine profile. In early pregnancy, a reduction of FOXP3 expression at the maternal-fetal interface of infected mice was also observed, and the reduction was larger in C57BL/6 compared with BALB/c mice. Additionally, the parasite was seldom found in the uterus/placenta. Thus, the worse pregnancy outcomes observed in C57BL/6 mice were associated with higher TNF systemically, and TNF and IL-6 at the maternal-fetal interface, with lower FOXP3 expression.

Anti-parasitic activity of Annona muricata L. leaf ethanolic extract and its fractions against Toxoplasma gondii in vitro and in vivo.

ETHNOPHARMACOLOGICAL RELEVANCE: Sulfadiazine and pyrimethamine are the two drugs used as part of the standard therapy for toxoplasmosis, however; they may cause adverse side effects and fail to prevent relapse in many patients, rendering infected individuals at risk of reactivation upon becoming immunocompromised. Extracts from various parts of Annona muricata have been widely used medicinally for the management, control and/or treatment of several human diseases, acting against parasites that cause diseases in humans. AIM OF THE STUDY: This study was performed to investigate the action of the ethanolic extract of A. muricata (EtOHAm) and its fractions in the control of the apicomplexan parasite Toxoplasma gondii in vitro and in vivo, and the effect of EtOHAm on the inflammatory response and lipid profile alteration induced by in vivo T. gondii infection. MATERIALS AND METHODS: The cytotoxicity of EtOHAm and its fractions ethyl acetate (EtOAcAm), n-butanol (BuOHAm), aqueous (H2OAm), hexane (HexAm) and dichloromethane (CH2Cl2Am) was evaluated in NIH/3T3 fibroblasts using the (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) assay. The cells were infected with T. gondii, treated with the extracts, and parasite proliferation was analyzed. For the in vivo experiments, C57BL/6 mice were orally infected with T. gondii and, treated with different concentrations of extract fractions that were effective in vitro (EtOHAm, EtOAcAm, HexAm and CH2Cl2Am). Tissue parasitism, histological alterations, systemic cytokine and lipid profile were investigated. RESULTS: EtOHAm, EtOAcAm, BuOHAm, H2OAm presented low cytotoxicity until doses of 200 µg/mL, while HexAm and CH2Cl2Am presented toxicity from doses of 100µg/mL. EtOHAm, HexAm and CH2Cl2Am decreased the parasitism in vitro, presenting a therapeutic index of 2.62, 2.44, and 2.96, respectively. In vivo, EtOHAm, HexAm and CH2Cl2Am improved the survival rate of infected animals, however, only EtOHAm was able to decrease the parasitism in the small intestine and lung. Additionally, EtOHAm decreased the systemic interferon (IFN)-γ and tumor necrosis factor (TNF) systemically in infected mice, and was able to maintain the triglycerides and very-low-density lipoprotein (VLDL) lipid fractions at similar levels to uninfected animals. Although treatment with EtOHAm could not control the inflammation induced by oral infection in the tissues analyzed, it was able to preserve the number of goblet cells in the small intestine. CONCLUSIONS: Ethanolic A. muricata leaf extract could be considered as a good candidate for the development of a complementary/alternative therapy against toxoplasmosis, and also as an anti-inflammatory alternative for decreasing TNF and IFN-γ concentrations and lipid fractions in specific diseases.

Annona muricata Linn. leaf as a source of antioxidant compounds with in vitro antidiabetic and inhibitory potential against α-amylase, α-glucosidase, lipase, non-enzymatic glycation and lipid peroxidation.

Annona muricata leaves are used in traditional medicine to manage diabetes mellitus and its complications. The aim of this study was to evaluate the potential in vitro antidiabetic properties of Annona muricata leaf by identifying its main phytochemical constituents and characterizing the phenolic-enriched fractions for their in vitro antioxidant capacity and inhibitory activities against glycoside and lipid hydrolases, advanced glycation end-product formation and lipid peroxidation. Ethanol extract of A. muricata leaf was subjected to a liquid-liquid partitioning and its fractions were used in enzymatic assays to evaluate their inhibitory potential against α-amylase, α-glucosidase and lipase, as well as their antioxidant (DPPH, ORAC, FRAP and Fe2+-ascorbate-induced lipid peroxidation assays) and anti-glycation (BSA-fructose, BSA-methylglyoxal and arginine-methylglyoxal models) capacities. In addition, identification of the main bioactive compounds of A. muricata leaf by HPLC-ESI-MS/MS analysis was carried out. Ethyl acetate (EtOAc) and n-butanol (BuOH) fractions showed, respectively, antioxidant properties (ORAC 3964 ±â€¯53 and 2707 ±â€¯519 µmol trolox eq g-1, FRAP 705 ±â€¯35 and 289 ±â€¯18 µmol trolox eq g-1, and DPPH IC50 4.3 ±â€¯0.7 and 9.3 ±â€¯0.8 µg mL-1) and capacity to reduce liver lipid peroxidation (p < .01). Also, EtOAc and BuOH, respectively, inhibited glycation in BSA-fructose (IC50 45.7 ±â€¯13.5 and 61.9 ±â€¯18.2 µg mL-1), BSA-methylglyoxal (IC50 166.1 ±â€¯21.6 and 413.2 ±â€¯49.5 µg mL-1) and arginine-methylglyoxal (IC50 437.9 ±â€¯89.0 and 1191.0 ±â€¯199.0 µg mL-1) assays, α-amylase (IC50 9.2 ±â€¯2.3 and 6.1 ±â€¯1.6 µg mL-1), α-glucosidase (IC50 413.1 ±â€¯121.1 and 817.4 ±â€¯87.9 µg mL-1) and lipase (IC50 74.2 ±â€¯30.1 and 120.3 ±â€¯50.5 µg.mL-1), and presented lower cytotoxicity, when compared to the other fractions and crude extract. Various biomolecules known as potent antioxidants were identified in these fractions, such as chlorogenic and caffeic acids, procyanidins B2 and C1, (epi)catechin, quercetin, quercetin-hexosides and kaempferol. This study presents new biological activities not yet described for A. muricata, which contributes to the understanding of the potential effectiveness in the use of the A. muricata leaf, especially its polyphenols-enriched fractions, for the management of diabetes mellitus and its complications.