Expansion of CD25-Negative Forkhead Box P3-Positive T Cells during HIV and Mycobacterium tuberculosis Infection.

Tuberculosis (TB) and HIV alter the immune system, and coinfected (HIV-TB) individuals usually present deregulations of T-lymphocytic immune response. We previously observed an increased frequency of "unconventional" CD4+CD25-FoxP3+ Treg (uTreg) population during HIV-TB disease. Therefore, we aimed to explore the phenotype and function of uTreg and conventional CD4+CD25+FoxP3+ Treg subsets (cTreg) in this context. We evaluated the expression of CD39, programmed cell death protein 1 (PD1), glucocorticoid-induced tumor necrosis factor receptor (GITR), and the effector/memory distribution by flow cytometry in cTreg and uTreg. Also, IL-10, TGF-ß, IFN-γ production, and the suppressor capacity of uTregs were analyzed in cocultures with effector lymphocytes and compared with the effect of regulatory T cells (Tregs). We found diminished expression of CD39 and higher levels of PD1 on uTreg compared to cTreg in both HIV-TB and healthy donors (HD). In addition, uTreg and cTreg showed differences in maturation status in both HIV-TB and HD groups, due to the expansion of effector memory uTregs. Interestingly, both HIV-TB and HD showed a pronounced production of IFN-γ in uTreg population, though no significant differences were observed for IL-10 and TGF-ß production between uTreg and cTreg. Moreover, IFN-γ+ cells were restricted to the CD39- uTreg population. Finally, when the suppressor capacity was evaluated, both uTreg and cTreg inhibited polyclonal T cell-proliferation and IFN-γ production in a similar extent. These findings suggest that uTregs, which are expanded during HIV-TB coinfection, exert regulatory functions in a similar way to cTregs despite an altered surface expression of Treg characteristic markers and differences in cytokine production.

Effect of simvastatin on endothelial cell apoptosis mediated by Fas and TNF-alpha.

Although there is evidence suggesting that statins may exert an endothelial protecting effect, recent in vitro data have shown that these compounds may induce endothelial cells (EC) apoptosis. We previously reported that the Fas-death receptor may induce apoptosis of the liver sinusoid endothelial cells (LSEC), and that TNF-alpha increases the susceptibility of these cells to suffer Fas-mediated apoptosis. Based on this evidence, in this study, we investigated the effect of simvastatin on Fas-mediated LSEC apoptosis. Simvastatin induced a significant reduction in LSEC viability, in a dose dependent manner, under serum-containing or serum-free conditions. This effect was prevented by mevalonate and GGPP, indicating the role of hydroxy-3-methylglutaryl-CoA reductase. The simvastatin effect on LSEC death was not associated with increased activation of caspase-3. We found that simvastatin increased the susceptibility of LSEC death mediated by Fas. Further, simvastatin increased LSEC-apoptosis induced by Fas and TNF-alpha. Mevalonate and GGPP partially prevented simvastatin-induced sensitization to LSEC death mediated by Jo2 and TNF-alpha, but not Jo2 alone. Simvastatin did not induce up-regulation of the Fas on the LSEC. Our results provide evidence of simvastatin in modulating Fas-mediated apoptosis in endothelial cells. These results may have clinical implications in those clinical conditions associated with high levels of FasL and TNF-alpha.

Leishmania chagasi y Trypanosoma cruzi: Conducta trófica en cultivos axénicos puros y mixtos / Leishmania chagasi and Trypanosoma cruzi: Trophic behavior in pure and mixed axenic cultures

El mecanismo adaptativo entre Leishmania y Trypanosoma ante la competencia por los recursos nutricionales en ambientes compartidos ha sido pobremente investigado. En particular, el estudio de la conducta trófica entre Leishmania chagasi y Trypanosoma cruzi podría contribuir al entendimiento de situaciones de gran relevancia médica en humanos, como son las infecciones mixtas por ambos tripanosomatídeos. En este trabajo se establecieron cultivos axénicos puros de Leishmania chagasi y Trypanosoma cruzi, así como cultivos axénicos mixtos (L. chagasi + T. cruzi) en médio BHI. Se registraron los cambios de dinámica poblacional (organismos por mililitro), la evolución de la estructura de las poblaciones, las variaciones de las concentraciones de colesterol, glucosa y proteínas totales, así como los cambios de pH en el medio de cultivo. El manejo cuantitativo de los conjuntos numéricos generados experimentalmente se abordó con técnicas univariadas (Análisis de la Varianza) y multivariadas (Análisis Discriminante Múltiple). Los resultados demuestran diferencias estadísticas significativas entre las medias de los parámetros considerados y prueban que el comportamiento “in vitro” investigado en el cultivo mixto L. chagasi – T. cruzi difiere taxativamente del estudiado en los cultivos puros de L. chagasi y T. Cruzi

The adaptive mechanisms betweenLeishmania and Trypanosoma facing the competencefor the nutritional resources in shared environmentshave been poorly investigated. Particularly, thestudy of the trophic behavior between Leishmaniachagasi and Trypanosoma cruzi could contribute tothe understanding of relevant medical situations, asmixed human infections. In this work pure axeniccultures of Leishmania chagasi and Trypanosomacruzi, as well as mixed cultures (L. chagasi + T.cruzi) were established in BHI medium. Changes ofpopulation dynamics (organisms/mL), the evolutionof the population’s structure, the variations of theconcentration of cholesterol, glucose and total proteins,as well as the changes of the medium’s pH wereregistered. The generated numerical sets were tackledwith univariate (Analysis of Variance) and multivariate(Multiple Discriminant Analysis) techniques. Theresults demonstrate significant statistical differencesbetween the media of the considered parameters andprove that the “in vitro” behavior investigated in theL. chagasi – T. cruzi mixed culture precisely differsfrom the L. chagasi and T. cruzi pure cultures

Antioxidant capacity, polyphenol content and iron bioavailability from algae (Ulva sp., Sargassum sp. and Porphyra sp.) in human subjects.

Marine algae are easily produced and are good sources of Fe. If this Fe is bioavailable, algae consumption could help to combat Fe deficiency and anaemia worldwide. The objective of the present study was to evaluate Fe bioavailability, polyphenol content and antioxidant capacity from three species of marine algae distributed worldwide. A total of eighty-three subjects received maize- or wheat-based meals containing marine algae (Ulva sp., Sargassum sp. and Porphyra sp.) in different proportions (2.5, 5.0 and 7.5 g) added to the water to prepare the dough. All meals administered contained radioactive Fe. Absorption was evaluated calculating radioactive Fe incorporation in subjects' blood. The three species of marine algae were analysed for polyphenol content and reducing power. Algae significantly increased Fe absorption in maize- or wheat-based meals, especially Sargassum sp., due to its high Fe content. Increases in absorption were dose-dependent and higher in wheat- than in maize-based meals. Total polyphenol content was 10.84, 18.43 and 80.39 gallic acid equivalents/g for Ulva sp., Porphyra sp. and Sargassum sp., respectively. The antioxidant capacity was also significantly higher in Sargassum sp. compared with the other two species analysed. Ulva sp., Sargassum sp. and Porphyra sp. are good sources of bioavailable Fe. Sargassum sp. resulted in the highest Fe intake due to its high Fe content, and a bread containing 7.5 g Sargassum sp. covers daily Fe needs. The high polyphenol content found in Sargassum sp. could be partly responsible for the antioxidant power reported here, and apparently did not affect Fe absorption.

High iron content and bioavailability in humans from four species of marine algae.

Searching for economical, nonconventional sources of iron is important in underdeveloped countries to combat iron deficiency and anemia. Our objective was to study iron, vitamin C, and phytic acid composition and also iron bioavailability from 4 species of marine algae included in a rice-based meal. Marine algae (Ulva sp, Sargassum sp, Porphyra sp, and Gracilariopsis sp) were analyzed for monthly variations in iron and for ascorbic acid and phytic acid concentrations. A total of 96 subjects received rice-based meals containing the 4 species of marine algae in different proportions, raw or cooked. All meals contained radioactive iron. Absorption was evaluated by calculating the radioactive iron incorporation in subjects' blood. Iron concentrations in algae were high and varied widely, depending on the species and time of year. The highest iron concentrations were found in Sargassum (157 mg/100 g) and Gracilariopsis (196 mg/100 g). Phytates were not detected in the algae and ascorbic acid concentration fluctuated between 38 microg/g dry weight (Ulva) and 362 microg/g dry weight (Sargassum). Algae significantly increased iron absorption in rice-based meals. Cooking did not affect iron absorption compared with raw algae. Results indicate that Ulva sp, Sargassum sp, Porphyra sp, and Gracilariopsis sp are good sources of ascorbic acid and bioavailable iron. The percentage of iron absorption was similar among all algae tested, although Sargassum sp resulted in the highest iron intake. Based on these results, and on the high reproduction rates of algae during certain seasons, promoting algae consumption in some countries could help to improve iron nutrition.

Activation of signaling lymphocytic activation molecule triggers a signaling cascade that enhances Th1 responses in human intracellular infection.

T cell production of IFN-gamma contributes to host defense against infection by intracellular pathogens, including mycobacteria. Lepromatous leprosy, the disseminated form of infection caused by Mycobacterium leprae, is characterized by loss of cellular response against the pathogen and diminished Th1 cytokine production. Relieving bacterial burden in Ag-unresponsive patients might be achieved through alternative receptors that stimulate IFN-gamma production. We have previously shown that ligation of signaling lymphocytic activation molecule (SLAM) enhances IFN-gamma in mycobacterial infection; therefore, we investigated molecular pathways leading from SLAM activation to IFN-gamma production in human leprosy. The expression of the SLAM-associated protein (an inhibitory factor for IFN-gamma induction) on M. leprae-stimulated cells from leprosy patients was inversely correlated to IFN-gamma production. However, SLAM ligation or exposure of cells from lepromatous patients to a proinflammatory microenvironment down-regulated SLAM-associated protein expression. Moreover, SLAM activation induced a sequence of signaling proteins, including activation of the NF-kappaB complex, phosphorylation of Stat1, and induction of T-bet expression, resulting in the promotion of IFN-gamma production, a pathway that remains quiescent in response to Ag in lepromatous patients. Therefore, our findings reveal a cascade of molecular events during signaling through SLAM in leprosy that cooperate to induce IFN-gamma production and strongly suggest that SLAM might be a focal point for therapeutic modulation of T cell cytokine responses in diseases characterized by dysfunctional Th2 responses.

Expression of signaling lymphocytic activation molecule-associated protein interrupts IFN-gamma production in human tuberculosis.

Production of the Th1 cytokine IFN-gamma by T cells is considered crucial for immunity against Mycobacterium tuberculosis infection. We evaluated IFN-gamma production in tuberculosis in the context of signaling molecules known to regulate Th1 cytokines. Two populations of patients who have active tuberculosis were identified, based on their T cell responses to the bacterium. High responder tuberculosis patients displayed significant M. tuberculosis-dependent T cell proliferation and IFN-gamma production, whereas low responder tuberculosis patients displayed weak or no T cell responses to M. tuberculosis. The expression of the signaling lymphocytic activation molecule (SLAM)-associated protein (SAP) on cells from tuberculosis patients was inversely correlated with IFN-gamma production in those individuals. Moreover, patients with a nonfunctional SAP gene displayed immune responses to M. tuberculosis similar to those of high responder tuberculosis patients. In contrast to SAP, T cell expression of SLAM was directly correlated with responsiveness to M. tuberculosis Ag. Our data suggest that expression of SAP interferes with Th1 responses whereas SLAM expression contributes to Th1 cytokine responses in tuberculosis. The study further suggests that SAP and SLAM might be focal points for therapeutic modulation of T cell cytokine responses in tuberculosis.

La proteina asociada a SLAM (SAP) regula la expresion de IFN-gamma en lepra / The SLAM-associated protein (SAP) regulates IFN-gamma expression in leprosy

La inmunidad protectora contra Mycobacterium leprae requiere IFN-gamma. Los pacientes con lepra tuberculoide producen localmente citoquinas Th1, mientras que los pacientes lepromatosos producen citoquinas Th2. La molécula linfocitaria activadora de señales (SLAM) y la proteína aociada a SLAM (SAP) participan en la diferenciación celular que conduce a producción de patrones específicos de citoquinas. A fin de investigar la vía SLAM/SAP en la infección por M. leprae, determinamos expresión de ARN mensajero (ARNm) de SAP, IFN-gamma y SLAM en pacientes con lepra. Obeservamos que la expresión de SLAM correlacionó en forma directa con la expresión de IFN-gamma, mientras que la expresión de SAP interferiría con las respuetas de citoquinas Th1 mientras que SLAM contribuiría con la respuesta Th1 en lepra, señalando a la vía SLAM/SAP como potencial blanco modulador de citoquinas en enfermedades con respuestas Th2 disfuncionales.