Lipid Fraction from Agaricus brasiliensis as a Potential Therapeutic Agent for Lethal Sepsis in Mice.

Sepsis is a potentially fatal clinical condition that results from an immune imbalance in the host during an infection. It presents systemic alterations due to excessive activation of pro-inflammatory mediators that contribute to inflammation, formation of reactive species, and tissue damage. Anti-inflammatory mediators are then extensively activated to regulate this process, leading to immune exhaustion and, consequently, immunosuppression of the host. Considering the biological activities of the nutraceutical Agaricus brasiliensis (A. brasiliensis), such as immunomodulatory, antioxidant, and antitumor activities, the present study investigated the therapeutic potential of the lipid fraction of A. brasiliensis (LF) in a model of lethal sepsis in mice (Mus musculus), induced by cecal ligation and perforation (CLP). The results showed that treatment of septic animals with LF or LF associated with ertapenem (LF-Erta) reduced systemic inflammation, promoting improvement in clinical parameters and increased survival. The data show a reduction in pro-inflammatory and oxidative stress markers, regulation of the anti-inflammatory response and oxidizing agents, and increased bacterial clearance in the peritoneal cavity and liver. Thus, it can be concluded that LF as a treatment, and in conjunction with antibiotic therapy, has shown promising effects as a hepatoprotective, antioxidant, antimicrobial, and immunomodulatory agent.

Anti-Leishmania effect of icetexanes from Salvia procurrens.

BACKGROUND AND PURPOSE: Leishmaniasis is a globally prevalent vector-borne disease caused by parasites of the genus Leishmania. The available chemotherapeutic drugs present problems related to efficacy, emergence of parasite resistance, toxicity and high cost, justifying the search for new drugs. Several classes of compounds have demonstrated activity against Leishmania, including icetexane-type diterpenes, previously isolated from Salvia and other Lamiaceae genera. Thus, in this study, compounds of Salvia procurrens were investigated for their leishmanicidal and immunomodulatory activities. METHODS: The exudate of S. procurrens was obtained by rapidly dipping the aerial parts in dichloromethane. The compounds were isolated by column and centrifugal planar chromatography over silica gel. The effects on L. amazonensis growth, survival, membrane integrity, reactive oxygen species (ROS) generation, mitochondrial membrane potential and cytotoxicity of the compounds towards human erythrocytes, peripheral blood mononuclear cells and macrophages were evaluated. The effects on intracellular amastigote forms, nitric oxide (NO) and TNF-α production were also investigated. RESULTS: The exudate from the leaves afforded the novel icetexane 7-hydroxyfruticulin A (1) as well as the known demethylisofruticulin A (2), fruticulin A (3) and demethylfruticulin A (4). The compounds (1-4) were tested against promastigotes of L. amazonensis and showed an effective inhibition of the parasite survival (IC50 = 4.08-16.26 µM). In addition, they also induced mitochondrial ROS production, plasma membrane permeability and mitochondrial dysfunction in treated parasites, and presented low cytotoxicity against macrophages. Furthermore, all diterpenes tested reduced the number of parasites inside macrophages, by mechanisms involving TNF-α, NO and ROS. CONCLUSION: The results suggest the potential of 7-hydroxyfruticulin A (1) as well as the known demethylisofruticulin A (2),fruticulin A (3) and demethylfruticulin A (4) as candidates for use in further studies on the design of anti-leishmanial drugs.

Polarization of HIV-1- and CMV-Specific IL-17-Producing T Cells among People with HIV under Antiretroviral Therapy with Cannabis and/or Cocaine Usage.

OBJECTIVE: This study evaluated the influence of cannabis and/or cocaine use in human immunodeficiency virus (HIV)- and cytomegalovirus (CMV)-specific T-cell responses of people with HIV (PWH). RESULTS: There was a higher percentage of IL-17-producing HIV-Gag-specific CD8+ T-cells in all drug users than that in PWH non-drug users. Stratifying the drug-user groups, increased percentages of IL-17-producing HIV-Gag-specific CD4+ and CD8+ T-cells were found in PWH cannabis plus cocaine users compared to PWH non-drug users. In response to CMV, there were higher percentage of IL-17-producing CMV-specific CD8+ T-cell in PWH cocaine users than that in PWH non-drug users. Considering all drug users together, there was a higher percentage of SEB-stimulated IL-17-producing CD4+ T-cells than that in PWH non-drug users, whereas cannabis users had higher percentages of IL-17-producing CD4+ T-cells compared to non-drug users. METHODS: Cryopreserved peripheral blood mononuclear cells from 37 PWH undergoing antiretroviral therapy (ART) using cannabis (10), cocaine (7), or cannabis plus cocaine (10) and non-drug users (10) were stimulated with HIV-1 Gag or CMV-pp65 peptide pools, or staphylococcal enterotoxin B (SEB) and evaluated for IFN-γ- and/or IL-17A-producing CD4+ and CD8+ T-cells using flow cytometry. CONCLUSIONS: Cannabis plus cocaine use increased HIV-specific IL-17 producing T-cells and cocaine use increased IL-17 CMV-specific CD8+ T-cell responses which could favor the inflammatory conditions associated with IL-17 overproduction.

Antiprotozoal potential of Vismia species (Hypericaceae), medicinal plants used to fight cutaneous leishmaniasis.

ETHNOPHARMACOLOGICAL RELEVANCE: Species of Vismia (Hypericaceae), known in Brazil as "lacre", are commonly used in traditional Amazonian medicine for the treatment of skin lesions, including those caused by Leishmania infection. AIM OF THE STUDY: Hexane extracts from the leaves of Vismia cayennensis, V. gracilis, V. sandwithii and V. guianensis, as well as from the fruits of the latter, in addition to the anthraquinones vismiaquinone, physcion and chrysophanol isolated from these species were explored for their anti-promastigote and anti-amastigote activity on Leishmania amazonensis. MATERIALS AND METHODS: Extracts were prepared by static maceration with n-hexane. The compounds, isolated by chromatographic techniques, were identified by spectroscopic methods (1H and 13C NMR). Promastigotes of L.amazonensis were incubated with hexane extracts (1-50 µg/mL) or anthraquinones (1-50 µM) and the parasite survival analyzed. The action of compounds on reactive oxygen species (ROS) production, mitochondrial membrane potential, and membrane integrity of promastigotes were evaluated by flow cytometer, and the cytotoxicity on mammalian cells using MTT assay. Furthermore, the activity of compounds against amastigotes and nitric oxide production were also investigated. RESULTS: Vismiaquinone and physcion were obtained from the leaves of V. guianensis. Physcion, as well as chrysophanol, were isolated from V. sandwithii. Vismia cayennensis and V. gracilis also showed vismiaquinone, compound detected in lower quantity in the fruits of V. guianensis. All extracts were active against the parasite, corroborating the popular use. The greatest activity against promastigotes was achieved with V. guianensis extract (IC50 4.3 µg/mL), precisely the most used Vismia species for treating cutaneous leishmaniasis. Vismiaquinone and physcion exhibited relevant activity with IC50 12.6 and 2.6 µM, respectively. Moreover, all extracts and anthraquinones tested induced ROS production, mitochondrial dysfunction, membrane disruption and were able to kill intracellular amastigote forms, being worthy of further in vivo studies as potential antileishmanial drugs. CONCLUSIONS: The overall data achieved in the current investigation scientifically validate the traditional use of Vismia species, mainly V. guianensis, as an anti-Leishmania agent. Furthermore, the promising results presented here indicate species of Vismia as potentially useful resources of Brazilian flora for the discovery of therapeutic solutions for neglected diseases.

Distinct anti-NP, anti-RBD and anti-Spike antibody profiles discriminate death from survival in COVID-19.

Introduction: Infection by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) induces rapid production of IgM, IgA, and IgG antibodies directed to multiple viral antigens that may have impact diverse clinical outcomes. Methods: We evaluated IgM, IgA, and IgG antibodies directed to the nucleocapsid (NP), IgA and IgG to the Spike protein and to the receptor-binding domain (RBD), and the presence of neutralizing antibodies (nAb), in a cohort of unvaccinated SARS-CoV-2 infected individuals, in the first 30 days of post-symptom onset (PSO) (T1). Results: This study included 193 coronavirus disease 2019 (COVID-19) participants classified as mild, moderate, severe, critical, and fatal and 27 uninfected controls. In T1, we identified differential antibody profiles associated with distinct clinical presentation. The mild group presented lower levels of anti-NP IgG, and IgA (vs moderate and severe), anti-NP IgM (vs severe, critical and fatal), anti-Spike IgA (vs severe and fatal), and anti-RBD IgG (vs severe). The moderate group presented higher levels of anti-RBD IgA, comparing with severe group. The severe group presented higher levels of anti-NP IgA (vs mild and fatal) and anti-RBD IgG (vs mild and moderate). The fatal group presented higher levels of anti-NP IgM and anti-Spike IgA (vs mild), but lower levels of anti-NP IgA (vs severe). The levels of nAb was lower just in mild group compared to severe, critical, and fatal groups, moreover, no difference was observed among the more severe groups. In addition, we studied 82 convalescent individuals, between 31 days to 6 months (T2) or more than 6 months (T3), PSO, those: 12 mild, 26 moderate, and 46 severe plus critical. The longitudinal analyzes, for the severe plus critical group showed lower levels of anti-NP IgG, IgA and IgM, anti-Spike IgA in relation T3. The follow-up in the fatal group, reveals that the levels of anti-spike IgG increased, while anti-NP IgM levels was decreased along the time in severe/critical and fatal as well as anti-NP IgG and IgA in several/critical groups. Discussion: In summary, the anti-NP IgA and IgG lower levels and the higher levels of anti-RBD and anti-Spike IgA in fatal compared to survival group of individuals admitted to the intensive care unit (ICU). Collectively, our data discriminate death from survival, suggesting that anti-RBD IgA and anti-Spike IgA may play some deleterious effect, in contrast with the potentially protective effect of anti-NP IgA and IgG in the survival group.

Heterologous Booster with BNT162b2 Induced High Specific Antibody Levels in CoronaVac Vaccinees.

Immune responses after COVID-19 vaccination should be evaluated in different populations around the world. This study compared antibody responses induced by ChAdOx1 nCoV-19, CoronaVac, and BNT162b2 vaccines. Blood samples from vaccinees were collected pre- and post-vaccinations with the second and third doses. The study enrolled 78 vaccinees, of whom 62.8% were women, with the following median ages: 26 years-ChAdOx1 nCoV-19; 40 years-CoronaVac; 30 years-BNT162b2. Serum samples were quantified for anti-RBD IgG and anti-RBD IgA and anti-spike IgG by ELISA. After two vaccine doses, BNT162b2 vaccinees produced higher levels of anti-RBD IgA and IgG, and anti-spike IgG compared to ChAdOx1 nCoV-19 and CoronaVac vaccinees. The third dose booster with BNT162b2 induced higher levels of anti-RBD IgA and IgG, and anti-spike IgG in CoronaVac vaccinees. Individuals who reported a SARS-CoV-2 infection before or during the study had higher anti-RBD IgA and IgG production. In conclusion, two doses of the studied vaccines induced detectable levels of anti-RBD IgA and IgG and anti-spike IgG in vaccinees. The heterologous booster with BNT162b2 increased anti-RBD IgA and IgG and anti-spike IgG levels in CoronaVac vaccinees and anti-RBD IgA levels in ChAdOx1 nCoV-19 vaccinees. Furthermore, SARS-CoV-2 infection induced higher anti-RBD IgA and IgG levels in CoronaVac vaccinees.

Integrated Metabolic and Inflammatory Signatures Associated with Severity of, Fatality of, and Recovery from COVID-19.

Severe manifestations of coronavirus disease 2019 (COVID-19) and mortality have been associated with physiological alterations that provide insights into the pathogenesis of the disease. Moreover, factors that drive recovery from COVID-19 can be explored to identify correlates of protection. The cellular metabolism represents a potential target to improve survival upon severe disease, but the associations between the metabolism and the inflammatory response during COVID-19 are not well defined. We analyzed blood laboratorial parameters, cytokines, and metabolomes of 150 individuals with mild to severe disease, of which 33 progressed to a fatal outcome. A subset of 20 individuals was followed up after hospital discharge and recovery from acute disease. We used hierarchical community networks to integrate metabolomics profiles with cytokines and markers of inflammation, coagulation, and tissue damage. Infection by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) promotes significant alterations in the plasma metabolome, whose activity varies according to disease severity and correlates with oxygen saturation. Differential metabolism underlying death was marked by amino acids and related metabolites, such as glutamate, glutamyl-glutamate, and oxoproline, and lipids, including progesterone, phosphocholine, and lysophosphatidylcholines (lysoPCs). Individuals who recovered from severe disease displayed persistent alterations enriched for metabolism of purines and phosphatidylinositol phosphate and glycolysis. Recovery of mild disease was associated with vitamin E metabolism. Data integration shows that the metabolic response is a hub connecting other biological features during disease and recovery. Infection by SARS-CoV-2 induces concerted activity of metabolic and inflammatory responses that depend on disease severity and collectively predict clinical outcomes of COVID-19. IMPORTANCE COVID-19 is characterized by diverse clinical outcomes that include asymptomatic to mild manifestations or severe disease and death. Infection by SARS-CoV-2 activates inflammatory and metabolic responses that drive protection or pathology. How inflammation and metabolism communicate during COVID-19 is not well defined. We used high-resolution mass spectrometry to investigate small biochemical compounds (<1,500 Da) in plasma of individuals with COVID-19 and controls. Age, sex, and comorbidities have a profound effect on the plasma metabolites of individuals with COVID-19, but we identified significant activity of pathways and metabolites related to amino acids, lipids, nucleotides, and vitamins determined by disease severity, survival outcome, and recovery. Furthermore, we identified metabolites associated with acute-phase proteins and coagulation factors, which collectively identify individuals with severe disease or individuals who died of severe COVID-19. Our study suggests that manipulating specific metabolic pathways can be explored to prevent hyperinflammation, organ dysfunction, and death.

Exposure to fine particulate matter partially counteract adaptations on glucose metabolism, oxidative stress, and inflammation of endurance exercise in rats.

BACKGROUND: Long-term exposure to air pollution triggers metabolic alterations along with oxidative stress and inflammation, while exercise interventions are widely used to improve those parameters. OBJECTIVE: Our study aimed to determine the effects of subchronic exposure to particulate matter 2.5 (PM2.5) and endurance exercise training on glucose metabolism, oxidative stress, and inflammation of the heart and gastrocnemius muscle of rats. MATERIAL AND METHODS: Thirty-two male Wistar rats were assigned to 4 experimental groups: Untrained; Endurance training (ET); Untrained + PM2.5; Endurance training + PM2.5. Rats exposed to air pollution received 50 µg of PM2.5 via intranasal instillation daily for 12 weeks. Exercised groups underwent endurance training, consisting in running on an electronic treadmill (70% of maximal capacity, 5 days/week, 5 times/week) for 12 weeks. Glucose metabolism markers, redox state, and inflammatory variables were evaluated in the heart and gastrocnemius muscle. RESULTS: ET and ET + PM2.5 group had lower body mass gain and higher exercise capacity, and higher glycogen concentration in the heart and gastrocnemius muscle. In the heart, ET and ET + PM2.5 groups had higher levels of GSH, and lower TBARS and TNF-α concentrations. In the gastrocnemius muscle, the ET group showed higher leptin and lower TBARS and IL-1ß concentrations, ET and ET + PM2.5 showed higher superoxide dismutase activity and ROS content. CONCLUSION: PM2.5 exposure partially blunts metabolic and inflammatory adaptations in heart and gastrocnemius muscle tissues induced by exercise training.

In vivo Acute Anti-Inflammatory Activity of Essential Oils: A Review.

In recent years, there has been a significant increase in the search for new therapeutic strategies for the treatment of inflammatory diseases. In this sense, natural products emerge as a potential source for the discovery of new drugs, with the research of the pharmacological properties of these products being very important. In addition to its function in plants (insect attraction and repellency), essential oils present pharmacological effects, such as antibacterial, antifungal, antimutagenic, antiviral, antiprotozoal, antioxidant, antidiabetic and anti-inflammatory properties. In this review, we describe the mostly used in vivo acute inflammatory experimental models and the studies showing the in vivo anti-inflammatory activity of essential oils. Essential oil from species from the Apiaceae, Asteraceae, Burseraceae, Boraginaceae, Cupressaceae, Euphorbiaceae, Fabaceae, Lamiaceae, Lauraceae, Myrtaceae, Piperaceae, Poaceae, Rutaceae, Verbenaceae and Zingiberaceae families were described as being anti-inflammatory in vivo. Five models of acute inflammation are commonly used to investigate the anti-inflammatory activity in vivo: ear and paw edema, pleurisy, peritonitis and the subcutaneous air pouch model. In addition to in vivo analysis, ex vivo and in vitro experiments are carried out to study the anti-inflammatory action of essential oils. The most commonly used model was paw edema, especially due to this model being easy to perform. In order to suggest or elucidate the mechanisms involved in the anti-inflammatory effect, many studies measured some inflammatory mediators, such as cytokines, COX-2 expression and the levels of PGE2, and NO, or evaluated the effect of essential oils or their major compounds on inflammation response directly induced by inflammatory mediators.

Reversal of haloperidol-induced orofacial dyskinesia and neuroinflammation by isoflavones.

BACKGROUND: Schizophrenia is a mental illness and its pharmacological treatment consists in the administration of antipsychotics like haloperidol. However, haloperidol often causes extrapyramidal motor disorders such as tardive dyskinesia (TD). So far, there is no effective treatment against TD and alternatives for it have been sought. Isoflafones have been studied as neuroprotector and inhibitor of monoamine oxidase enzyme. Thus, the objective is to evaluate the possible protective effect of isoflavones against the induction of involuntary movements induced by haloperidol in an animal model. METHODS AND RESULTS: Male Wistar rats were treated with haloperidol (1 mg/kg/day) and/or isoflavones (80 mg/kg) for 28 days. Rats were submitted to behavioral evaluation to quantify vacuous chewing movements (VCM) and locomotor activity. In addition, the levels of pro-inflammatory cytokines were measured in the striatum. Haloperidol treatment reduced the locomotor activity and increased the number of VCM in rats. Co-treatment with isoflavones was able to reverse hypolocomotion and reduce the number of VCM. Besides, haloperidol caused significant increase in the proinflammatory cytokines (interleukin-1ß:IL-1ß, tumor necrosis factor-α: TNF-α and IL-6 and the co-treatment with isoflavones was able to reduce the levels of IL-1ß and TNF-α, but not IL-6. CONCLUSIONS: It is believed that the beneficial effect found with this alternative treatment is related to its anti-inflammatory potential and to the action on estrogen receptors (based on scientific literature findings). Finally, further studies are needed to elucidate the mechanisms of isoflavones in reducing motor disorders induced by antipsychotics.

O3 concentration and duration of exposure are factors influencing the environmental health risk of exercising in Rio Grande, Brazil.

Ozone (O3) represents a great threat to human health, contributing to respiratory diseases and premature mortality. This pollutant is often considered a critical pollutant in regions of southern Brazil. Exposure to this pollutant during vigorous physical activity should be the subject of thorough investigations due to the increased ventilation rate and altered breathing pattern present during vigorous physical activity that result in greater inhalation of O3. Thus, this study aimed to evaluate the health risk of exposure to low, mean, and high concentrations of O3 during different durations of exercise in the city of Rio Grande (southern Brazil). Healthy young men (n = 45) performed cardiopulmonary exercise testing, and ventilation rate data were collected to predict total ventilation and pollutant inhalation during a 5 km running session. The O3 concentration in the city of Rio Grande was obtained from data reported by the Copernicus Atmosphere Monitoring Service (CAMS). The environmental health risk was calculated based on the potential intake dose. The lowest, mean, and highest concentrations of O3 detected during the monitoring period were 32.5, 64.9, and 115.2 µg/m3, respectively. In all evaluated scenarios, there was a toxicological risk (RQ > 1), except when exercising when the O3 concentration was lowest for the shortest length of time (p < 0.001). As the concentration of O3 and the duration of the exposure increase, the health risk is increased. Therefore, O3 concentration and duration of exposure are factors influencing the health risk of exercising. These findings are extremely relevant in cities that have high levels of O3, such as the city of Rio Grande.

Air pollution inhalation during acute exercise is dependent of the body mass index and ventilation of young men.

Obesity and physical inactivity threaten human health, and both could be solved with exercise. However, a higher amount of pollutants is inhaled during exercise. Exposure to air pollution increases the incidence and progression of diseases. Therefore, the aim of the study was to investigate the rate of pollution inhalation of lean, overweight, and obese individuals in a low and high-intensity hypothetical exercise session. Healthy sedentary men (n = 135) classified as lean, overweight, or obese were enrolled in our study. All participants performed a cardiopulmonary exercise testing (CPX) to collect ventilation rate (VE) data, which was used to predict total ventilation and pollutant inhalation of a 5-km running session. Air pollutant concentration of São Paulo City, Brazil was evaluated and the toxicological risk was estimated based on the potential intake dose. The concentrations of PM2.5 were 29.57 µg/m3 and 51.71 µg/m3, PM10 were 45.85 µg/m3 and 74 µg/m3, NO2 were 63.71 µg/m3 and 66.65 µg/m3, and O3 were 69 µg/m3 and 37 µg/m3, respectively in the summer and winter. In the hypothetical exercise session, total VE and time in both the first and second threshold were increased in the obese group (p < 0.001) (p < 0.001). The inhalation of PM2.5, PM10, NO2, and O3, during the hypothetical session, was increased in obese individuals (p < 0.001). Obese individuals should be considered a susceptible population, once they are more exposed to air pollution during exercise.

Agaricus brasiliensis Mushroom Protects Against Sepsis by Alleviating Oxidative and Inflammatory Response.

Sepsis is characterized by the host's dysregulated immune response to an infection followed by a potentially fatal organ dysfunction. Although there have been some advances in the treatment of sepsis, mainly focused on broad-spectrum antibiotics, mortality rates remain high, urging for the search of new therapies. Oxidative stress is one of the main features of septic patients, so antioxidants can be a good alternative treatment. Agaricus brasiliensis is a nutraceutical rich in bioactive compounds such as polyphenols and polysaccharides, exhibiting antioxidant, antitumor, and immunomodulatory activities. Here, we investigated the immunomodulatory and antioxidant effects of A. brasilensis aqueous extract in the cecal ligation and puncture (CLP) sepsis model. Our data showed that aqueous extract of A. brasiliensis reduced systemic inflammatory response and improved bacteria clearance and mice survival. In addition, A brasiliensis decreased the oxidative stress markers in serum, peritoneal cavity, heart and liver of septic animals, as well as ROS production (in vitro and in vivo) and tert-Butyl hydroperoxide-induced DNA damage in peripheral blood mononuclear cells from healthy donors in vitro. In conclusion, the aqueous extract of A. brasiliensis was able to increase the survival of septic animals by a mechanism involving immunomodulatory and antioxidant protective effects.

Clinical and Pathological Aspects of Canine Cutaneous Leishmaniasis: A Meta-analysis.

PURPOSE: In the present study, we described the most prevalent clinical symptoms, the most affected organs, and the macro and microscopic lesions associated with cutaneous leishmaniasis. METHODS: Two independent researchers performed an extensive systematic review of the literature in four stages (identification, screening, eligibility, and inclusion) to identify studies published between January 2002 and November 2018 from the following electronic databases: Web of Science, PubMed, SciELO, Science Direct, and Google Scholar. Meta-analysis was conducted in "Metaprop" package of R 3.4.2 software. RESULTS: The electronic search yielded 3896 results, out of which 155 were further analyzed based on the full-text. Data extracted from 16 articles were included in the meta-analysis, representing a total of 430 leishmaniasis cases. Only 43% of all animals were identified to exhibit the clinical and cutaneous changes characteristic of leishmaniasis based on the observation that skin lesions were the most prevalent clinical sign and were present in 86% of all cases. Other less prevalent symptoms included weight loss, cachexia, apathy and lymph node enlargement. Histopathological analysis showed that the skin was the most affected organ, affecting 64% of cases, followed by lymph nodes (12%), spleen (8%) and liver (7%). CONCLUSIONS: Therefore, our current findings suggest that cutaneous leishmaniasis could lead to visceral disease. Notably, our findings indicated no clinical manifestation patterns in cutaneous leishmaniasis, since the same host species may present different clinical conditions.

Leishmanicidal and antichemotactic activities of icetexanes from Salvia uliginosa Benth.

BACKGROUND: Several species of Salvia are used as medicinal plants around the world. Biological activities of isolated compounds have been described, being diterpenes frequently responsible for the effects. PURPOSE: Isolation of diterpenes from Salvia uliginosa Benth. and evaluation of the antichemotactic and leishmanicidal activities of the isolated compounds. STUDY DESIGN: To isolate diterpenes from S. uliginosa and evaluate their antichemotactic and leishmanicidal activities in vitro. METHODS: The exudate of S. uliginosa was obtained by rapidly dipping the aerial parts in dichloromethane. The compounds were isolated by repeated column chromatography over silica gel. The effects on L. amazonensis growth, survival, DNA degradation, ROS generation, as well as the antichemotactic activity and cytotoxicity of the compounds towards human erythrocytes and macrophages were evaluated. RESULTS: A novel icetexane diterpene, isoicetexone (IsoICT) along with the known diterpenes icetexone (ICT), and 7-acetoxy-6,7-dihydroicetexone were isolated from the dichloromethane surface exudate of S. uliginosa. The structures were elucidated using NMR and MS experiments, and by comparison with previously reported data. IsoICT and ICT at low concentrations caused completely inhibition of neutrophils migration in vitro. In addition, IsoICT and ICT showed high leishmanicidal activity against L. amazonensis, induced ROS production in parasites and presented low cytotoxicity against macrophages and human erythrocytes, and moderate to high selectivity index. CONCLUSION: These data indicated that IsoICT and ICT exhibit potent antichemotactic and leishmanicidal effects. Further studies are needed in order to evaluate the in vivo activities as well as the toxicity of the compounds.

Aedes-Chikungunya Virus Interaction: Key Role of Vector Midguts Microbiota and Its Saliva in the Host Infection.

Aedes mosquitoes are important vectors for emerging diseases caused by arboviruses, such as chikungunya (CHIKV). These viruses' main transmitting species are Aedes aegypti and Ae. albopictus, which are present in tropical and temperate climatic areas all over the globe. Knowledge of vector characteristics is fundamentally important to the understanding of virus transmission. Only female mosquitoes are able to transmit CHIKV to the vertebrate host since they are hematophagous. In addition, mosquito microbiota is fundamentally important to virus infection in the mosquito. Microorganisms are able to modulate viral transmission in the mosquito, such as bacteria of the Wolbachia genus, which are capable of preventing viral infection, or protozoans of the Ascogregarina species, which are capable of facilitating virus transmission between mosquitoes and larvae. The competence of the mosquito is also important in the transmission of the virus to the vertebrate host, since their saliva has several substances with biological effects, such as immunomodulators and anticoagulants, which are able to modulate the host's response to the virus, interfering in its pathogenicity and virulence. Understanding the Aedes vector-chikungunya interaction is fundamentally important since it can enable the search for new methods of combating the virus' transmission.

In vitro and in silico Activity of Iridoids Against Leishmania amazonensis.

BACKGROUND: Leishmaniasis reaches millions of people around the world. The control of the disease is difficult due to the restricted access to the diagnosis and medication, and low adherence to the treatment. Thus, more efficient drugs are needed and natural products are good alternatives. Iridoids, natural products with reported leishmanicidal activity, can be exploited for the development of anti- Leishmania drugs. The aim of this study was to isolate and to investigate the in vitro activity of iridoids against Leishmania amazonensis and to compare the activity in silico of these compounds with those reported as active against this parasite. METHODS: Iridoids were isolated by chromatographic methods. The in vitro activity of asperuloside (1) and geniposide (2) from Escalonia bifida, galiridoside (3) from Angelonia integerrima and theveridoside (4) and ipolamiide (5) from Amphilophium crucigerum was investigated against promastigote forms of Leishmania amazonensis. Molecular modeling studies of 1-5 and iridoids cited as active against Leishmania spp. were performed. RESULTS: Compounds 1-5 (5-100 µM) did not inhibit the parasite survival. Physicochemical parameters predicted for 1-5 did not show differences compared to those described in literature. The SAR and the pharmacophoric model confirmed the importance of maintaining the cyclopentane[C]pyran ring of the iridoid, of oxygen-linked substituents at the C1 and C6 positions and of bulky substituents attached to the iridoid ring to present leishmanicidal activity. CONCLUSION: The results obtained in this study indicate that iridoids are a promising group of secondary metabolites and should be further investigated in the search for new anti-Leishmania drugs.

Salivary Gland Extract from Aedes aegypti Improves Survival in Murine Polymicrobial Sepsis through Oxidative Mechanisms.

Sepsis is a systemic disease with life-threatening potential and is characterized by a dysregulated immune response from the host to an infection. The organic dysfunction in sepsis is associated with the production of inflammatory cascades and oxidative stress. Previous studies showed that Aedes aegypti saliva has anti-inflammatory, immunomodulatory, and antioxidant properties. Considering inflammation and the role of oxidative stress in sepsis, we investigated the effect of pretreatment with salivary gland extract (SGE) from Ae. aegypti in the induction of inflammatory and oxidative processes in a murine cecum ligation and puncture (CLP) model. Here, we evaluated animal survival for 16 days, as well as bacterial load, leukocyte migration, and oxidative parameters. We found that the SGE pretreatment improved the survival of septic mice, reduced bacterial load and neutrophil influx, and increased nitric oxide (NO) production in the peritoneal cavity. With regard to oxidative status, SGE increased antioxidant defenses as measured by Trolox equivalent antioxidant capacity (TEAC) and glutathione (GSH), while reducing levels of the oxidative stress marker malondialdehyde (MDA). Altogether, these data suggest that SGE plays a protective role in septic animals, contributing to oxidative and inflammatory balance during sepsis. Therefore, Ae. aegypti SGE is a potential source for new therapeutic molecule(s) in polymicrobial sepsis, and this effect seems to be mediated by the control of inflammation and oxidative damage.

Imidazolium salts as innovative agents against Leishmania amazonensis.

The available chemotherapeutic drugs for the treatment of leishmaniasis present problems relating to efficacy, emergence of parasite resistance, and adverse effects and cost. Azole antifungal drugs have been repurposed for this proposition but the clinical response has been variable. In this sense, this study assessed the leishmanicidal and immunomodulatory activities of azoles-derived imidazolium salts (IS), being the ionic imidazole-derived equivalents: 1-n-butyl-3-methylimidazolium chloride (C4MImCl), 1-n-decyl-3-methylimidazolium chloride (C10MImCl), 1-n-hexadecyl-3-methylimidazolium chloride (C16MImCl), 1-n-hexadecyl-3-methylimidazolium methanesulfonate (C16MImMeS), 1-n-hexadecyl-3-methylimidazolium bis(trifluoromethanesulfonyl)imide (C16MImNTf2) and 1-methyl-3-n-octadecylimidazolium chloride (C18MImCl). Promastigotes of Leishmania amazonensis were incubated with IS at concentrations ranging from 0.1 to 100 µM, and the parasite survival was monitored. The effects of IS on reactive oxygen species (ROS) production and mitochondrial membrane potential of promastigotes, as well as on cytotoxicity against peripheral blood mononuclear cells (PBMC) and human erythrocytes were determined. Besides, the activities of IS against amastigotes and nitric oxide production were also evaluated. The IS inhibited parasite growth and showed potent leishmanicidal activity against promastigotes of L. amazonensis. In addition, IS induced mitochondrial dysfunction and ROS production in parasites, and presented low cytotoxicity against PBMC and human erythrocytes. Furthermore, at very low concentration (0.5 µM), C18MImCl, C16MImMeS, C16MImCl, C10MImCl and C16MImNTf2 were able to kill intramacrophage parasites at levels of 91.3, 100, 94.4, 95.3 and 35.6%, respectively. These results indicate that IS are promising candidates for the development of drugs against L. amazonensis.

Molecular Method Confirms Canine Leishmania Infection Detected by Serological Methods in Non-Endemic Area of Brazil.

In Brazil, visceral leishmaniasis (VL) is expanding and becoming urbanized, especially in non-endemic areas such as the State of Rio Grande do Sul. Considering that infected dogs are the main reservoir for zoonotic VL, this study evaluated the prevalence of canine visceral leishmaniasis (CVL) in the metropolitan area of Porto Alegre, a new area of expansion of VL in Brazil. Serum and plasma from 405 asymptomatic dogs from the municipalities of Canoas (n=107), São Leopoldo (n=216), and Novo Hamburgo (n=82) were tested for CVL using immunochromatographic (DPP®) and ELISA EIE® assays (2 assays officially adopted by the Brazilian government for the diagnosis of CVL) and real-time PCR to confirm the results. There was no agreement among serological and real-time PCR results, indicating that the Leishmania infection in asymptomatic animals with low parasite load, confirmed by negative parasitological tests (smears and parasite culture), need to be evaluated by molecular methods. The prevalence of LVC in the metropolitan region of Porto Alegre, confirmed by real-time PCR was 4% (5.6% in Canoas and 4.6% in São Leopoldo). The use of molecular method is essential for accurate diagnosis of CVL, especially in asymptomatic dogs in non-endemic areas.