Leishmania infantum (syn. L. chagasi) parasites affect the release of soluble CD14 by infected macrophages.

Functionally, cluster of differentiation 14 (CD14) is a co-receptor of the complex formed by lipopolysaccharide (LPS) and LPS-binding protein expressed on the membrane of a variety of cells. However, CD14 can be shed from the cell membrane into the circulation as soluble CD14 (sCD14) upon cell activation. Previously, our group reported that elevated sCD14 serum levels were associated with the clinical and laboratory findings in the context of visceral leishmaniasis (VL), but not in the context of LPS stimulation or bacterial infection. In the present study, we investigated the secretion dynamics of sCD14 in the context of Leishmania infantum (syn. L. chagasi) in vitro infection. Macrophages from treated VL patients and delayed-type hypersensitivity positive (DTH+) subjects were infected with L. infantum (syn. L. chagasi) promastigotes, and the infection index was evaluated (number of amastigotes per 100 infected macrophages). Additionally, the levels of sCD14, Inteleukin (IL)10, IL-6 and tumour necrosis factor alpha (TNF-α) were measured in the culture supernatants using the Luminex assay. Interestingly, the release of sCD14 was inversely correlated with the L. infantum (syn. L. chagasi) infection index. Of note, the release of sCD14 was upregulated and downregulated in the context of infected macrophages from DTH+ subjects and treated VL patients, respectively. Additionally, we also observed that the levels of sCD14 in the culture supernatants were positively correlated with the levels of TNF-α, IL-6 and IL-10. Therefore, our data suggest that macrophages from treated VL patients and DTH+ subjects respond differently to L. infantum (syn. L. chagasi) infection in the context of the release of sCD14; therefore, the release of sCD14 may be associated with the outcome of VL.

Plasma Total Antioxidant Capacity and Cardiometabolic Risk in Non-Obese and Clinically Healthy Young Adults / Capacidade Antioxidante Total do Plasma e Risco Cardiometabólico em Adultos Jovens, Não Obesos e Clinicamente Saudáveis

Abstract Background: The oxidative biomarkers play an important role in the genesis of cardiometabolic risk-related processes. Objective: To investigate the total antioxidant capacity of plasma and its association with cardiometabolic risk in non-obese and clinically healthy young adults. Methods: University students of the state of Sergipe, Brazil, aged between 18 and 25 years, were recruited for this study from May of 2013 and October of 2014. Anthropometric, clinical and biochemical parameters were measured and analyzed using protocols which were previously standardized and described in the literature. The measurement of plasma total antioxidant capacity was based on the ability that all the antioxidants present in the sample (plasma) have to inhibit the oxidation of the oxidizable substrate ABTS (2,2`- Azino-di-[3-ethylbenzthiazoline sulphonate]) to ABTS•+ by metmyoglobin. Results: Approximately 25% of the sample presented more than one component of cardiometabolic risk. Low HDL-cholesterol was the most prevalent component. Compared to absence of components, the subjects with at least one component presented greater body weight and waist circumference, higher levels of diastolic blood pressure and fasting glucose, greater total cholesterol/HDL-c ratio, and lower levels of HDL-c (p < 0.05). Fasting glycemia was the only parameter which was associated with total antioxidant capacity (R2 = 0.10; β = 0.17; p = 0.001). Conclusions: The plasma total antioxidant capacity was not able to predict the cardiometabolic risk components due possibly to the establishment of compensatory mechanisms that become activated in physiological conditions.

Resumo Fundamentos: Os biomarcadores oxidativos exercem um importante papel na gênese dos processos relacionados ao risco cardiometabólico. Objetivo: Investigar a capacidade antioxidante total do plasma e sua associação com risco cardiometabólico em adultos jovens, não obesos e clinicamente saudáveis. Métodos: Estudantes universitários do estado de Sergipe, Brasil, com idade entre 18 e 25 anos, foram recrutados entre maio de 2013 e outubro de 2014. Parâmetros antropométricos, clínicos e bioquímicos foram medidos e analisados usando protocolos previamente padronizados e descritos na literatura. A medida da capacidade antioxidante total do plasma baseou-se na capacidade de todos os antioxidantes presentes na amostra (plasma) em inibir a oxidação do substrato oxidável ABTS (2,2-Azino-bis-(3-etilbenzotiazolina-6-sulfonato) a ABTS•+ pela metamioglobina. Resultados: Aproximadamente 25% da amostra apresentaram mais de um componente do risco cardiometabólico. Valores baixos de HDL foram o componente mais prevalente. Em comparação à ausência de componentes, os indivíduos com pelo menos um componente apresentou valores mais altos de peso corporal, circunferência da cintura, pressão sanguínea diastólica, glicemia de jejum e razão colesterol total/HDL-c, e valores mais baixos de HDL-c (p < 0,05). A glicemia de jejum foi o único parâmetro que se associou com a capacidade antioxidante total (R2 = 0,10; β = 0,17; p = 0,001). Conclusões: A capacidade antioxidante total não foi capaz de predizer os componentes do risco cardiometabólico possivelmente devido ao estabelecimento de mecanismos compensatórios que se tornam ativados em condições fisiológicas.

Plasma Total Antioxidant Capacity and Cardiometabolic Risk in Non-Obese and Clinically Healthy Young Adults.

BACKGROUND:: The oxidative biomarkers play an important role in the genesis of cardiometabolic risk-related processes. OBJECTIVE:: To investigate the total antioxidant capacity of plasma and its association with cardiometabolic risk in non-obese and clinically healthy young adults. METHODS:: University students of the state of Sergipe, Brazil, aged between 18 and 25 years, were recruited for this study from May of 2013 and October of 2014. Anthropometric, clinical and biochemical parameters were measured and analyzed using protocols which were previously standardized and described in the literature. The measurement of plasma total antioxidant capacity was based on the ability that all the antioxidants present in the sample (plasma) have to inhibit the oxidation of the oxidizable substrate ABTS (2,2`- Azino-di-[3-ethylbenzthiazoline sulphonate]) to ABTS•+ by metmyoglobin. RESULTS:: Approximately 25% of the sample presented more than one component of cardiometabolic risk. Low HDL-cholesterol was the most prevalent component. Compared to absence of components, the subjects with at least one component presented greater body weight and waist circumference, higher levels of diastolic blood pressure and fasting glucose, greater total cholesterol/HDL-c ratio, and lower levels of HDL-c (p < 0.05). Fasting glycemia was the only parameter which was associated with total antioxidant capacity (R2 = 0.10; ß = 0.17; p = 0.001). CONCLUSIONS:: The plasma total antioxidant capacity was not able to predict the cardiometabolic risk components due possibly to the establishment of compensatory mechanisms that become activated in physiological conditions. FUNDAMENTOS:: Os biomarcadores oxidativos exercem um importante papel na gênese dos processos relacionados ao risco cardiometabólico. OBJETIVO:: Investigar a capacidade antioxidante total do plasma e sua associação com risco cardiometabólico em adultos jovens, não obesos e clinicamente saudáveis. MÉTODOS:: Estudantes universitários do estado de Sergipe, Brasil, com idade entre 18 e 25 anos, foram recrutados entre maio de 2013 e outubro de 2014. Parâmetros antropométricos, clínicos e bioquímicos foram medidos e analisados usando protocolos previamente padronizados e descritos na literatura. A medida da capacidade antioxidante total do plasma baseou-se na capacidade de todos os antioxidantes presentes na amostra (plasma) em inibir a oxidação do substrato oxidável ABTS (2,2-Azino-bis-(3-etilbenzotiazolina-6-sulfonato) a ABTS•+ pela metamioglobina. RESULTADOS:: Aproximadamente 25% da amostra apresentaram mais de um componente do risco cardiometabólico. Valores baixos de HDL foram o componente mais prevalente. Em comparação à ausência de componentes, os indivíduos com pelo menos um componente apresentou valores mais altos de peso corporal, circunferência da cintura, pressão sanguínea diastólica, glicemia de jejum e razão colesterol total/HDL-c, e valores mais baixos de HDL-c (p < 0,05). A glicemia de jejum foi o único parâmetro que se associou com a capacidade antioxidante total (R2 = 0,10; ß = 0,17; p = 0,001). CONCLUSÕES:: A capacidade antioxidante total não foi capaz de predizer os componentes do risco cardiometabólico possivelmente devido ao estabelecimento de mecanismos compensatórios que se tornam ativados em condições fisiológicas.

Infection of Human Macrophages by Leishmania infantum Is Influenced by Ecto-Nucleotidases.

Ecto-nucleotidase activity is involved in the infection process of Leishmania and various other parasites that enables modulation of host immune responses to promote disease progression. One of the enzymes responsible for this activity is the ecto-nucleoside triphosphate diphosphohydrolase (E-NTPDase). The enzyme hydrolyzes nucleotides tri- and/or di-phosphate into monophosphate products, which are subsequently hydrolyzed into adenosine. These nucleotides can serve as purinergic signaling molecules involved in diverse cellular processes that govern immune responses. Given the importance of the extracellular metabolism of these nucleotides during intracellular pathogen infections, this study evaluates the role of ecto-nucleotidase activity during Leishmania infantum (L. infantum) infection in human macrophages. E-NTPDase protein expression and activity was evaluated in L. infantum during purine starvation, adenosine-enriched medium, or in the presence of an inhibitor of ecto-nucleotidases. Results show that E-NTPDase is expressed in L. infantum parasites, including on the cell membrane. Furthermore, functional activity of the enzyme was modulated according to the availability of adenosine in the medium. Purine starvation increased the hydrolytic capacity of nucleotides leading to higher infectivity, while growth in adenosine-enriched medium led to lower infectivity. Moreover, inhibiting E-NTPDase function decreased L. infantum infection in macrophages, suggesting the enzyme may serve as a ligand. Taken together, the ability of L. infantum to hydrolyze nucleotides is directly associated with increased infectivity in macrophages.

Trypanosoma cruzi nucleoside triphosphate diphosphohydrolase 1 (TcNTPDase-1) biochemical characterization, immunolocalization and possible role in host cell adhesion.

Previous work has suggested that Trypanosoma cruzi diphosphohydrolase 1 (TcNTPDase-1) may be involved in the infection of mammalian cells and serve as a potential target for rational drug design. In this work, we produced recombinant TcNTPDase-1 and evaluated its nucleotidase activity, cellular localization and role in parasite adhesion to mammalian host cells. TcNTPDase-1 was able to utilize a broad range of triphosphate and diphosphate nucleosides. The enzyme's Km for ATP (0.096 mM) suggested a capability to influence the host's ATP-dependent purinergic signaling. The use of specific polyclonal antibodies allowed us to confirm the presence of TcNTPDase-1 at the surface of parasites by confocal and electron microscopy. In addition, electron microscopy revealed that TcNTPDase-1 was also found in the flagellum, flagellum insertion region, kinetoplast, nucleus and intracellular vesicles. The presence of this enzyme in the flagellum insertion region and vesicles suggests that it may have a role in nutrient acquisition, and the widespread distribution of TcNTPDase-1 within the parasite suggests that it may be involved in other biological process. Adhesion assays using anti-TcNTPDase-1 polyclonal antibodies as a blocker or purified recombinant TcNTPDase-1 as a competitor revealed that the enzyme has a role in parasite-host cell adhesion. These data open new frontiers to future studies on this specific parasite-host interaction and other unknown functions of TcNTPDase-1 related to its ubiquitous localization.