Essential oil of oregano (Origanum vulgare L.) reduces infection and proliferation of Toxoplasma gondii in BeWo cells with induction of autophagy and death of tachyzoites through a mechanism similar to necrosis.

Toxoplasmosis poses a global health threat, ranging from asymptomatic cases to severe, potentially fatal manifestations, especially in immunocompromised individuals and congenital transmission. Prior research suggests that oregano essential oil (OEO) exhibits diverse biological effects, including antiparasitic activity against Toxoplasma gondii. Given concerns about current treatments, exploring new compounds is important. This study was to assess the toxicity of OEO on BeWo cells and T. gondii tachyzoites, as well as to evaluate its effectiveness in in vitro infection models and determine its direct action on free tachyzoites. OEO toxicity on BeWo cells and T. gondii tachyzoites was assessed by MTT and trypan blue methods, determining cytotoxic concentration (CC50), inhibitory concentration (IC50), and selectivity index (SI). Infection and proliferation indices were analyzed. Direct assessments of the parasite included reactive oxygen species (ROS) levels, mitochondrial membrane potential, necrosis, and apoptosis, as well as electron microscopy. Oregano oil exhibited low cytotoxicity on BeWo cells (CC50: 114.8 µg/mL ± 0.01) and reduced parasite viability (IC50 12.5 ± 0.06 µg/mL), demonstrating 9.18 times greater selectivity for parasites than BeWo cells. OEO treatment significantly decreased intracellular proliferation in infected cells by 84% after 24 h with 50 µg/mL. Mechanistic investigations revealed increased ROS levels, mitochondrial depolarization, and lipid droplet formation, linked to autophagy induction and plasma membrane permeabilization. These alterations, observed through electron microscopy, suggested a necrotic process confirmed by propidium iodide labeling. OEO treatment demonstrated anti-T. gondii action through cellular and metabolic change while maintaining low toxicity to trophoblastic cells.

Trilobolide-6-O-isobutyrate from Sphagneticola trilobata acts by inducing oxidative stress, metabolic changes and apoptosis-like processes by caspase 3/7 activation of human lung cancer cell lines.

BACKGROUND: Lung cancer, a chronic and heterogeneous disease, is the leading cause of cancer-related death on a global scale. Presently, despite a variety of available treatments, their effectiveness is limited, often resulting in considerable toxicity and adverse effects. Additionally, the development of chemoresistance in cancer cells poses a challenge. Trilobolide-6-O-isobutyrate (TBB), a natural sesquiterpene lactone extracted from Sphagneticola trilobata, has exhibited antitumor effects. Its pharmacological properties in NSCLC lung cancer, however, have not been explored. PURPOSE: This study evaluated the impact of TBB on the A549 and NCI-H460 tumor cell lines in vitro, examining its antiproliferative properties and initial mechanisms of cell death. METHODS: TBB, obtained at 98 % purity from S. trilobata leaves, was characterized using chromatographic techniques. Subsequently, its impact on inhibiting tumor cell proliferation in vitro, TBB-induced cytotoxicity in LLC-MK2, THP-1, AMJ2-C11 cells, as well as its effects on sheep erythrocytes, and the underlying mechanisms of cell death, were assessed. RESULTS: In silico predictions have shown promising drug-likeness potential for TBB, indicating high oral bioavailability and intestinal absorption. Treatment of A549 and NCI-H460 human tumor cells with TBB demonstrated a direct impact, inducing significant morphological and structural alterations. TBB also reduced migratory capacity without causing toxicity at lower concentrations to LLC-MK2, THP-1 and AMJ2-C11 cell lines. This antiproliferative effect correlated with elevated oxidative stress, characterized by increased levels of ROS, superoxide anion radicals and NO, accompanied by a decrease in antioxidant markers: SOD and GSH. TBB-stress-induced led to changes in cell metabolism, fostering the accumulation of lipid droplets and autophagic vacuoles. Stress also resulted in compromised mitochondrial integrity, a crucial aspect of cellular function. Additionally, TBB prompted apoptosis-like cell death through activation of caspase 3/7 stressors. CONCLUSION: These findings underscore the potential of TBB as a promising candidate for future studies and suggest its viability as an additional component in the development of novel anticancer drugs prototypes.

Leishmania amazonensis infection regulates oxidate stress in hyperglycemia and diabetes impairing macrophage's function and immune response.

Leishmaniasis is a group of infectious diseases caused by protozoa of the Leishmania genus and its immunopathogenesis results from an unbalanced immune response during the infection. Diabetes is a chronic disease resulting from dysfunction of the body's production of insulin or the ability to use it properly, leading to hyperglycemia causing tissue damage and impairing the immune system. AIMS: The objective of this work was to evaluate the effects of hyperglycemia and diabetes during Leishmania amazonensis infection and how these conditions alter the immune response to the parasite. METHODS: An in vitro hyperglycemic stimulus model using THP-1-derived macrophages and an in vivo experimental diabetes with streptozotocin (STZ) in C57BL/6 mice was employed to investigate the impact of diabetes and hyperglicemia in Leishmania amazonensis infection. RESULTS: We observed that hyperglycemia impair the leishmanicidal capacity of macrophages derived from THP-1 cells and reverse the resistance profile that C57BL/6 mice have against infection by L. amazonensis, inducing more exacerbated lesions compared to non-diabetic animals. In addition, the hyperglycemic stimulus favored the increase of markers related to the phenotype of M2 macrophages. The induction of experimental diabetes in C57BL/6 mice resulted in a failure in the production of nitric oxide (NO) in the face of infection and macrophages from diabetic animals failed to process and present Leishmania antigens, being unable to activate and induce proliferation of antigen-specific lymphocytes. CONCLUSION: Together, these data demonstrate that diabetes and hyperglycemia can impair the cellular immune response, mainly of macrophages, against infection by parasites of the genus Leishmania.

Antioxidant Compounds and Health Benefits of Citrus Fruits.

Recent evidence emanating from epidemiological prospective studies shows that increased intakes of antioxidant-rich fruits, vegetables, and legumes are associated with a lower risk of developing chronic oxidative stress-related diseases like cardiovascular diseases and cancer, as well as with a lower risk of cardiovascular, cancer, and all-cause mortality rates [...].

Synergistic Antileishmanial Effect of Oregano Essential Oil and Silver Nanoparticles: Mechanisms of Action on Leishmania amazonensis.

American tegumentary leishmaniasis, a zoonotic disease caused by the Leishmania genus, poses significant challenges in treatment, including administration difficulty, low efficacy, and parasite resistance. Novel compounds or associations offer alternative therapies, and natural products such as oregano essential oil (OEO), extracted from Origanum vulgare, have been extensively researched due to biological effects, including antibacterial, antifungal, and antiparasitic properties. Silver nanoparticles (AgNp), a nanomaterial with compelling antimicrobial and antiparasitic activity, have been shown to exhibit potent leishmanicidal properties. We evaluated the in vitro effect of OEO and AgNp-Bio association on L. amazonensis and the death mechanisms of the parasite involved. Our results demonstrated a synergistic antileishmanial effect of OEO + AgNp on promastigote forms and L. amazonensis-infected macrophages, which induced morphological and ultrastructural changes in promastigotes. Subsequently, we investigated the mechanisms underlying parasite death and showed an increase in NO, ROS, mitochondrial depolarization, accumulation of lipid-storage bodies, autophagic vacuoles, phosphatidylserine exposure, and damage to the plasma membrane. Moreover, the association resulted in a reduction in the percentage of infected cells and the number of amastigotes per macrophage. In conclusion, our findings establish that OEO + AgNp elicits a late apoptosis-like mechanism to combat promastigote forms and promotes ROS and NO production in infected macrophages to target intracellular amastigote forms.

Biogenic silver nanoparticle exhibits schistosomicidal activity in vitro and reduces the parasitic burden in experimental schistosomiasis mansoni.

Schistosomiasis is a neglected tropical parasitic disease that affects millions of people, being the second most prevalent parasitic disease worldwide. The current treatment has limited effectiveness, drug-resistant strains, and is not effective in different stages of the disease. This study investigated the antischistosomal activity of biogenic silver nanoparticles (Bio-AgNp) against Schistosoma mansoni. Bio-AgNp presented direct schistosomicidal activity on newly transformed schistosomula causing plasma membrane permeabilization. In S. mansoni adult worms, reduced the viability and affected the motility, increasing oxidative stress parameters, and inducing plasma membrane permeabilization, loss of mitochondrial membrane potential, lipid bodies accumulation, and autophagic vacuoles formation. During the experimental schistosomiasis mansoni model, Bio AgNp restored body weight, reduced hepatosplenomegaly, and decrease the number of eggs and worms in feces and liver tissue. The treatment also ameliorates liver damage and reduces macrophage and neutrophil infiltrates. A reduction in count and size was evaluated in the granulomas, as well as a change to an exudative-proliferative phase, with a local increase of IFN-γ. Together our results showed that Bio-AgNp is a promising therapeutic candidate for studies of new therapeutic strategies against schistosomiasis.

Naringenin and Hesperidin as Promising Alternatives for Prevention and Co-Adjuvant Therapy for Breast Cancer.

Citrus (genus Citrus L.) fruits are essential sources of bioactive compounds with antioxidant properties, such as flavonoids. These polyphenolic compounds are divided into subclasses, in which flavanones are the most prominent. Among them, naringenin and hesperidin are emerging compounds with anticancer potential, especially for breast cancer (BC). Several mechanisms have been proposed, including the modulation of epigenetics, estrogen signaling, induction of cell death via regulation of apoptotic signaling pathways, and inhibition of tumor invasion and metastasis. However, this information is sparse in the literature and needs to be brought together to provide an overview of how naringenin and hesperidin can serve as therapeutic tools for drug development and as a successful co-adjuvant strategy against BC. This review detailed such mechanisms in this context and highlighted how naringenin and hesperidin could interfere in BC carcinogenesis and be helpful as potential alternative therapeutic sources for breast cancer treatment.

Melatonin modulates the Warburg effect and alters the morphology of hepatocellular carcinoma cell line resulting in reduced viability and migratory potential.

AIMS: Hepatocellular Carcinoma (HCC) is a primary neoplasm derived from hepatocytes with low responsiveness and recurrent chemoresistance. Melatonin is an alternative agent that may be helpful in treating HCC. We aimed to study in HuH 7.5 cells whether melatonin treatment exerts antitumor effects and, if so, what cellular responses are induced and involved. MAIN METHODS: We evaluated the effects of melatonin on cell cytotoxicity and proliferation, colony formation, morphological and immunohistochemical aspects, and on glucose consumption and lactate release. KEY FINDINGS: Melatonin reduced cell motility and caused lamellar breakdown, membrane damage, and reduction in microvillus. Immunofluorescence analysis revealed that melatonin reduced TGF and N-cadherin expression, which was further associated with inhibition of epithelial-mesenchymal transition process. In relation to the Warburg-type metabolism, melatonin reduced glucose uptake and lactate production by modulating intracellular lactate dehydrogenase activity. SIGNIFICANCE: Our results indicate that melatonin can act upon pyruvate/lactate metabolism, preventing the Warburg effect, which may reflect in the cell architecture. We demonstrated the direct cytotoxic and antiproliferative effect of melatonin on the HuH 7.5 cell line, and suggest that melatonin is a promising candidate to be further tested as an adjuvant to antitumor drugs for HCC treatment.

Biogenic silver nanoparticles (AgNp-Bio) restore testosterone levels and increase TNF-α and IL-6 in Leydig cells infected with Toxoplasma gondii.

Toxoplasma gondii, a protozoan parasite, is responsible for toxoplasmosis. The available therapy for patients with toxoplasmosis involves a combination of pyrimethamine and sulfadiazine, which have several adverse effects, including bone marrow suppression, megaloblastic anemia, leukopenia, and granulocytopenia. The development of therapeutic alternatives is essential for the management of toxoplasmosis, emphasizing the recent advances in nanomedicine. This study aimed to evaluate the in vitro effects of biogenic silver nanoparticles (AgNp-Bio) on tachyzoite forms and Leydig cells infected with T. gondii. We observed that the AgNp-Bio reduced the viability of the tachyzoites and did not exhibit cytotoxicity against Leydig cells at low concentrations. Additionally, treatment with AgNp-Bio reduced the rate of infection and proliferation of the parasite, and lowered the testosterone levels in the infected cells. It increased the levels of IL-6 and TNF-α and reduced the levels of IL- 10. Among the morphological and ultrastructural changes, AgNp-Bio induced a reduction in the number of intracellular tachyzoites and caused changes in the tachyzoites with accumulation of autophagic vacuoles and a decrease in the number of tachyzoites inside the parasitophorous vacuoles. Collectively, our data demonstrate that the AgNp-Bio affect T. gondii tachyzoites by activating microbicidal and inflammatory mechanisms and could be a potential alternative treatment for toxoplasmosis.

Vitamin D3 and Salinomycin synergy in MCF-7 cells cause cell death via endoplasmic reticulum stress in monolayer and 3D cell culture.

1α, 25, dihydroxyvitamin D3 (1,25D), the active form of vitamin D3, has antitumor properties in several cancer cell lines in vitro. Salinomycin (Sal) has anticancer activity against cancer cell lines. This study aims to examine the cytotoxic and antiproliferative effect of Sal associated with 1,25D on MCF-7 breast carcinoma cell line cultured in monolayer (2D) and three-dimensional models (mammospheres). We also aim to evaluate the molecular mechanism of Sal and 1,25D-mediated effects. We report that Sal and 1,25D act synergistically in MCF-7 mammospheres and monolayer causing G1 cell cycle arrest, reduction of mitochondrial membrane potential (MMP), and reactive oxygen species (ROS) overproduction with a long-lasting cytotoxic response represented by clonogenic and mammosphere assay. We observed the induction of cell death by apoptosis with upregulation in mRNA levels of apoptosis-related genes (CASP7, CASP9, and BBC3). Extensive cytoplasmic vacuolization, a morphological characteristic found in paraptosis, was also seen and could be triggered by endoplasmic reticulum stress (ER) as we found transcriptional upregulation of genes related to ER stress (ATF6, GADD153, GADD45G, EIF2AK3, and HSPA5). Overall, Sal and 1,25D act synergistically, inhibiting cell proliferation by activating simultaneously multiple death pathways and may be a novel and promising luminal A breast cancer therapy strategy.

Grandiflorenic acid from Wedelia trilobata plant induces apoptosis and autophagy cell death in breast adenocarcinoma (MCF7), lung carcinoma (A549), and hepatocellular carcinoma (HuH7.5) cells lines.

INTRODUCTION: Wedelia trilobata (Sphagneticola trilobata) is a plant used in this popular medicine for treating infectious, sores and swellings in some rural communities, and their extract has antioxidant, anti-inflammatory, antitumor and hepatoprotective effect. Cancer is a molecularly heterogeneous disease caused by environmental and, genetic factors, among others. Since the complexity of the disease leads to low response rates to the different treatments used, it is necessary to find alternative drugs aimed at its control. The objective of our study was to assess whether grandiflorenic acid (GFA) has antitumor activity on breast (MCF7), liver (HuH7.5), and lung (A549) tumor cell lines. METHODS: We used cell integrity assessment methods to assess whether (GFA) would be cytotoxic for tumor cell lines at doses ranging from and the pattern of death involved in this effect. RESULTS: Treatment using GFA significantly inhibited cell proliferation in the three studied cells, followed by a decrease in cell size. The assessment of the death mechanisms showed the treatments increased the production of reactive oxygen species, caused exposure of phosphatidylserine, depolarization of the mitochondrial membrane, and, decrease plasma membrane integrity, indicating mechanisms related to apoptosis. Besides, we found the formation of autophagy vacuoles in our tests. CONCLUSION: Finally, our study found the effect of GFA on breast (MCF7), lung (A549), and liver (HuH7.5) tumor cell lines induce cytotoxicity and patterns of death associated with apoptosis and autophagy, and oxidative stress generation plays a role in these two pathways of cell death. Thus, this study revealed GFA exhibits anti-cancer activity in vitro and could help future studies to improve strategies for cancer treatment with involving natural compounds.

Susceptibility to first choice antimicrobial treatment for urinary tract infections to Escherichia coli isolates from women urine samples in community South Brazil.

E. coli is the main pathogen of UTI. It is important to be aware the local epidemiological data for an appropriate initial treatment. Resistance to antimicrobial agents has increased, especially to first-choice antibiotics in the treatment of cystitis. There are few studies on the sensivity profile of community uropathogen in our region. OBJECTIVE: To characterize antimicrobials the sensitivity profile to E. coli isolated from urocultures of women treated at Basic Health Units and Emergency Care Units of Londrina- Paraná- Brazil during a period of 12 months (June 1, 2016 to June 1, 2017). METHODOLOGY: A cross-sectional study was carried out from June 2016 to June 2017. All urine samples collected in the Basic Health Units and Emergency Departments in the city of Londrina (Paraná State, Brazil) were sent to a Central Laboratory where the identification and antimicrobial susceptibility testing were performed. Clinical Laboratory Standards Institute (CLSI) breakpoints were used for the interpretation of susceptibility testing results. RESULTS: 56,555 urine cultures were performed in the period, of which 8,832 were positive, of which 5,377 were women. Of these samples, 4.7% were enterobacteria producing extended-spectrum beta-lactamases (ESBL) and 15.5% resistant to quinolones. TMP- SMX was resistant in more than 30% of the samples in all age groups. Among quinolone-resistant isolates, resistance to cephalothin, ampicillin and sulfamethoxazole-trimethoprim was greater than 60%. Nitrofurantoin was the only antimicrobial that showed 90% of sensitivity. CONCLUSION: The antimicrobials sensitivity profile was similar to that reported in the literature, with TMP- SMX resistance greater than 30% in the studied samples. Nitrofurantoin maintains high sensitivity rates greater than 90%. Resistance to quinolones increases proportionally with age, as well ESBL.

Biogenic silver nanoparticles reduce Toxoplasma gondii infection and proliferation in RAW 264.7 macrophages by inducing tumor necrosis factor-alpha and reactive oxygen species production in the cells.

Owing to the serious adverse effects caused by pyrimethamine and sulfadiazine, the drugs commonly used to treat toxoplasmosis, there is a need for treatment alternatives for this disease. Nanotechnology has enabled significant advances toward this goal. This study was conducted to evaluate the activity of biogenic silver nanoparticles (AgNp-Bio) in RAW 264.7 murine macrophages infected with the RH strain of Toxoplasma gondii. The macrophages were infected with T. gondii tachyzoites and then treated with various concentrations of AgNp-Bio. The cells were evaluated by microscopy, and culture supernatants were collected for ELISA determination of their cytokine concentration. Treatment with 6 µM AgNp-Bio reduced the infection and parasite load in infected RAW 264.7 macrophages without being toxic to the cells. The treatment also induced the synthesis of reactive oxygen species and tumor necrosis factor-alpha (both pro-inflammatory mediators), which resulted in ultrastructural changes in the tachyzoites and their intramacrophagic destruction. Our findings suggest that AgNp-Bio affect T. gondii tachyzoites by activating microbicidal and pro-inflammatory mechanisms and may be a potential alternative treatment for toxoplasmosis.

Investigation of the antileishmanial activity and mechanisms of action of acetyl-thiohydantoins.

The currently available treatment options for leishmaniasis are associated with high costs, severe side effects, and high toxicity. In previous studies, thiohydantoins demonstrated some pharmacological activities and were shown to be potential hit compounds with antileishmanial properties. The present study further explored the antileishmanial effect of acetyl-thiohydantoins against Leishmania amazonensis and determined the main processes involved in parasite death. We observed that compared to thiohydantoin nuclei, acetyl-thiohydantoin treatment inhibited the proliferation of promastigotes. This treatment caused alterations in cell cycle progression and parasite size and caused morphological and ultrastructural changes. We then investigated the mechanisms involved in the death of the protozoan; there was an increase in ROS production, phosphatidylserine exposure, and plasma membrane permeabilization and a loss of mitochondrial membrane potential, resulting in an accumulation of lipid bodies and the formation of autophagic vacuoles on these parasites and confirming an apoptosis-like process. In intracellular amastigotes, selected acetyl-thiohydantoins reduced the percentage of infected macrophages and the number of amastigotes/macrophages by increasing ROS production and reducing TNF-α levels. Moreover, thiohydantoins did not induce cytotoxicity in murine macrophages (J774A.1), human monocytes (THP-1), or sheep erythrocytes. In silico and in vitro analyses showed that acetyl-thiohydantoins exerted in vitro antileishmanial effects on L. amazonensis promastigotes in apoptosis-like and amastigote forms by inducing ROS production and reducing TNF-α levels, indicating that they are good candidates for drug discovery studies in leishmaniasis treatment. Additionally, we carried out molecular docking analyses of acetyl-thiohydantoins on two important targets of Leishmania amazonensis: arginase and TNF-alpha converting enzyme. The results suggested that the acetyl groups in the N1-position of the thiohydantoin ring and the ring itself could be pharmacophoric groups due to their affinity for binding amino acid residues at the active site of both enzymes via hydrogen bond interactions. These results demonstrate that thiohydantoins are promising hit compounds that could be used as antileishmanial agents.

Botryosphaeran, [(1 → 3)(1 → 6)-ß-D-glucan], induces apoptosis-like death in promastigotes of Leishmania amazonensis, and exerts a leishmanicidal effect on infected macrophages by activating NF-kB and producing pro-inflammatory molecules.

Leishmaniasis is an infectious-parasitic disease caused by the protozoan Leishmania spp. The available treatments are based upon expensive drugs bearing adverse side-effects. The search for new therapeutic alternatives that present a more effective action without causing adverse effects to the patient is therefore important. The objective of this study was to evaluate the in vitro effect of botryosphaeran, a (1 â†’ 3)(1 â†’ 6)-ß-D-glucan, on the promastigote and intracellular amastigote forms of Leishmania amazonensis. The direct activity of botryosphaeran on promastigote forms was evaluated in vitro and inhibited proliferation, the IC50 7 µg/mL in 48 h was calculated. After 48 h treatment, botryosphaeran induced nitric oxide production (NO), caused mitochondrial membrane hyperpolarization, increased reactive oxygen species (ROS), and accumulation of lipid vesicles in promastigotes, resulting in apoptosis, necrosis and autophagy, and was accompanied by morphological and ultrastructural changes. The range of concentrations used did not alter the viability of peritoneal macrophages from BALB/c mice and erythrocytes of sheep. Botryosphaeran was able to reduce the number of infected macrophages and the number of amastigotes per macrophage at 12.5 µg/mL (50.75% ± 6.48), 25 µg/mL (55.66% ± 3.93) and 50 µg/mL (72.9% ± 6.98), and IC50 9.3 µg/mL (±0.66) for intracellular amastigotes forms. The leishmanicidal effect was due to activation of NF-κB and promoted an increase in pro-inflammatory cytokines (TNF-α and IL-6), iNOS and microbial-derived ROS and NO, in addition to decreasing the levels of SOD. Based upon the data obtained, we infer that botryosphaeran exerted an active leishmanicidal and immunomodulatory effect, acting on promastigotes through autophagic, apoptotic and necrosis processes, and in the intracellular amastigote form, through the action of ROS and NO.

Grandiflorenic acid isolated from Sphagneticola trilobata against Trypanosoma cruzi: Toxicity, mechanisms of action and immunomodulation.

Grandiflorenic acid (GFA) is one of the main kaurane diterpenes found in different parts of Sphagneticola trilobata. It has several biological activities, especially antiprotozoal action. In turn, Chagas disease is a complex systemic disease caused by the protozoan Trypanosoma cruzi, and the drugs available to treat it involve significant side effects and impose an urgent need to search for therapeutic alternatives. In this context, our goal was to determine the effect of GFA on trypomastigote and intracellular amastigote forms. Our results showed that GFA treatment led to significantly less viability of trypomastigote forms, with morphological and ultrastructural changes in the parasites treated with IC50 of GFA (24.60 nM), and larger levels of reactive oxygen species (ROS), mitochondrial depolarization, lipid droplets accumulation, presence of autophagic vacuoles, phosphatidylserine exposure, and plasma membrane damage. In addition, the GFA treatment was able to reduce the percentage of infected cells and the number of amastigotes per macrophage (J774A.1) without showing cytotoxicity in mammalian cell lines (J774A.1, LLCMK2, THP-1, AMJ2-C11), in addition to increasing TNF-α and reducing IL-6 levels in infected macrophages. In conclusion, the GFA treatment exerted influence on trypomastigote forms through an apoptosis-like mechanism and by eliminating intracellular parasites via TNF-α/ROS pathway, without generating cellular cytotoxicity.

Patterns of Cell Death Induced by Thiohydantoins in Human MCF-7 Breast Cancer Cells.

BACKGROUND: Conventional therapies for breast cancer are still a challenge due to cytotoxic drugs not being highly effective with significant adverse effects. Thiohydantoins are biologically active heterocyclic compounds reported for several biological activities, including anticarcinogenic properties, etc. This work aims to assess the use of thiohydantoin as a potential antitumor agent against MCF-7 breast cancer cells. METHODS: MTT and neutral red assays were used to assess the possible cytotoxic activity of compounds against MCF-7 cells. Cell volume measurement and analysis were performed by flow cytometry. Fluorescence analysis was carried out to determine patterns of cell death induced by thiohydantoins. RESULTS: The treatment with micromolar doses of thiohydantoins promoted a decrease in the viability of MCF-7 breast tumor cells. An increase in the ROS and NO production, reduction in cell volume, loss of membrane integrity, mitochondrial depolarization, and increased fluorescence for annexin-V and caspase-3 were also observed. These findings indicate cell death by apoptosis and increased autophagic vacuoles, stopping the cell cycle in the G1/ G0 phase. CONCLUSION: Our results indicate that thiohydantoins are cytotoxic to breast tumor cells, and this effect is linked to the increase in ROS production. This phenomenon changes tumorigenic pathways, which halt the cell cycle in G1/G0. This is an essential checkpoint for DNA errors, which may have altered how cells produce energy, causing a decrease in mitochondrial viability and thus leading to the apoptotic process. Furthermore, the results indicate increased autophagy, a vital process linked to a decrease in lysosomal viability and thus considered a cell death and tumor suppression mechanism.

Exploring the antileishmanial activity of N1,N2-disubstituted-benzoylguanidines: synthesis and molecular modeling studies.

In this report, we describe the synthesis and evaluation of nine N1,N2-disubstituted-benzoylguanidines against promastigotes and amastigotes forms of Leishmania amazonensis. The derivatives 2g and 2i showed low IC50 values against promastigote form (90.8 ± 0.05 µM and 68.4 ± 0.03 µM, respectively), low cytotoxicity profile (CC50 396 ± 0.02 µM and 857.9 ± 0.06 µM) for peritoneal macrophages cells and SI of 5.5 and 12.5, respectively. Investigations about the mechanism of action of 2g and 2i showed that both compounds cause mitochondrial depolarization, increase in ROS levels, and generation of autophagic vacuoles on free promastigotes forms. These compounds were also capable of reducing the number of infected macrophages with amastigotes forms (59.5% ± 0.08% and 98.1% ± 0.46%) and the number of amastigotes/macrophages (79.80% ± 0.05% and 96.0% ± 0.16%), through increasing induction of microbicide molecule NO. Additionally, ADMET-Tox in silico predictions showed drug-like features and free of toxicological risks. The molecular docking studies with arginase and gp63 showed that relevant intermolecular interactions could explain the experimental results. Therefore, these results reinforce that benzoylguanidines could be a starting scaffold for the search for new antileishmanial drugs.Communicated by Ramaswamy H. Sarma.

Susceptibility to first choice antimicrobial treatment for urinary tract infections to Escherichia coli isolates from women urine samples in community South Brazil

ABSTRACT E. coli is the main pathogen of UTI. It is important to be aware the local epidemiological data for an appropriate initial treatment. Resistance to antimicrobial agents has increased, especially to first-choice antibiotics in the treatment of cystitis. There are few studies on the sensivity profile of community uropathogen in our region. Objective: To characterize antimicrobials the sensitivity profile to E. coli isolated from urocultures of women treated at Basic Health Units and Emergency Care Units of Londrina-Paraná- Brazil during a period of 12 months (June 1, 2016 to June 1, 2017). Methodology: A cross-sectional study was carried out from June 2016 to June 2017. All urine samples collected in the Basic Health Units and Emergency Departments in the city of Londrina (Paraná State, Brazil) were sent to a Central Laboratory where the identification and antimicrobial susceptibility testing were performed. Clinical Laboratory Standards Institute (CLSI) breakpoints were used for the interpretation of susceptibility testing results. Results: 56,555 urine cultures were performed in the period, of which 8,832 were positive, of which 5,377 were women. Of these samples, 4.7% were enterobacteria producing extended-spectrum beta-lactamases (ESBL) and 15.5% resistant to quinolones. TMP- SMX was resistant in more than 30% of the samples in all age groups. Among quinolone-resistant isolates, resistance to cephalothin, ampicillin and sulfamethoxazole-trimethoprim was greater than 60%. Nitrofurantoin was the only antimicrobial that showed 90% of sensitivity. Conclusion: The antimicrobials sensitivity profile was similar to that reported in the literature, with TMP- SMX resistance greater than 30% in the studied samples. Nitrofurantoin maintains high sensitivity rates greater than 90%. Resistance to quinolones increases proportionally with age, as well ESBL.

Murine Susceptibility to Leishmania amazonensis Infection Is Influenced by Arginase-1 and Macrophages at the Lesion Site.

Cutaneous leishmaniasis is a zoonotic infectious disease broadly distributed worldwide, causing a range of diseases with clinical outcomes ranging from self-healing infections to chronic disfiguring disease. The effective immune response to this infection is yet to be more comprehensively understood and is fundamental for developing drugs and vaccines. Thus, we used experimental models of susceptibility (BALB/c) and partial resistance (C57BL/6) to Leishmania amazonensis infection to investigate the local profile of mediators involved in the development of cutaneous leishmaniasis. We found worse disease outcome in BALB/c mice than in C57BL/6 mice, with almost 15 times higher parasitic load, ulcerated lesion formation, and higher levels of IL-6 in infected paws. In contrast, C57BL/6 presented higher levels of IFN-γ and superoxide anion (•O2-) after 11 weeks of infection and no lesion ulcerations. A peak of local macrophages appeared after 24 h of infection in both of the studied mice strains, followed by another increase after 240 h, detected only in C57BL/6 mice. Regarding M1 and M2 macrophage phenotype markers [iNOS, MHC-II, CD206, and arginase-1 (Arg-1)], we found a pronounced increase in Arg-1 levels in BALB/c after 11 weeks of infection, whereas C57BL/6 showed an initial predomination of markers from both profiles, followed by an M2 predominance, coinciding with the second peak of macrophage infiltration, 240 h after the infection. Greater deposition of type III collagen and lesion resolution was also observed in C57BL/6 mice. The adoptive transfer of macrophages from C57BL/6 to infected BALB/c at the 11th week showed a reduction in both edema and the number of parasites at the lesion site, in addition to lower levels of Arg-1. Thus, C57BL/6 mice have a more effective response against L. amazonensis, based on a balance between inflammation and tissue repair, while BALB/c mice have an excessive Arg-1 production at late infection. The worst evolution seems to be influenced by recruitment of Arg-1 related macrophages, since the adoptive transfer of macrophages from C57BL/6 mice to BALB/c resulted in better outcomes, with lower levels of Arg-1.