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Aging is a complex process best characterized as the chronic dysregulation of cellular processes leading to deteriorated tissue and organ function. While aging cannot currently be prevented, its impact on lifespan and healthspan in the elderly can potentially be minimized by interventions that aim to return these cellular processes to optimal function. Recent studies have demonstrated that partial reprogramming using the Yamanaka factors (or a subset;OCT4, SOX2, andKLF4; OSK)can reverse age-related changesin vitroandin vivo. However, it is still unknown whether the Yamanaka factors (or a subset) are capable of extending the lifespan of aged wild type mice. Here, we show that systemically delivered AAVs, encoding an inducible OSK system, in 124-week-old mice extends the median remaining lifespan by 109% over wild-type controls and enhances several health parameters. Importantly, we observed a significant improvement in frailty scores indicating that we were able to improve the healthspan along with increasing the lifespan. Furthermore, in human keratinocytes expressing exogenous OSK, we observed significant epigenetic markers of age-reversal, suggesting a potential reregulation of genetic networks to a younger, potentially healthier state. Together, these results may have important implications for the development of partial reprogramming interventions to reverse age-associated diseases in the elderly.

The corrosion resistance of an alkyd paint was evaluated, substituting commercial anticorrosive with a thin film of NiO-ZrO2Y partially stabilized with Ytria deposited by means of the RF sputtering technique. Hot-rolled (HR) steel was used as the substrate. For the two anticorrosive coating systems, the structure, morphology, and composition were determined via the techniques of X-ray diffraction (XRD), scanning electron microscopy (SEM), energy-dispersive X-ray spectroscopy (EDX), and X-ray photoelectron spectroscopy (XPS) in order to determine the composition of the NiO-ZrO2Y film. The corrosion resistance was determined from polarization curves and weight loss by immersion in a 3.5% NaCl solution. The electrochemical and weight loss results indicated that use of the NiO-ZrO2 film shows a fourfold increase in the corrosion resistance of the material subjected to immersion in the corrosive electrolyte compared to the traditionally-used commercial anticorrosive.

Antecedentes. Puede existir comorbilidad entre el Trastorno del Espectro Autista (TEA) y el Trastorno por Déficit de Atención/Hiperactividad (TDAH), aunque hasta hace pocos años existía restricción para su diagnóstico. Objetivo. En este estudio se describe el caso de un niño de 11 años con diagnóstico dual de TEA y TDAH que es evaluado con el fin de documentar la presencia de déficits en el funcionamiento ejecutivo y adaptativo, y explorar la correspondencia entre medidas cognitivas y una medida conductual de Funciones Ejecutivas (FE). Metodología. Se administraron las pruebas M-WCST, Stroop, TMT-A y TMT-B, test de Símbolos y Dígitos, y test de Fluidez fonológica y semántica como medidas cognitivas de FE; el Brief-2 como medida de evaluación conductual de FE, y las escalas Vinneland-3 como instrumento de habilidades adaptativas. Resultados. Las medidas cognitivas de FE permitieron identificar dificultades principalmente en memoria de trabajo. Por su parte, el instrumento conductual informó rigidez cognitiva, dificultad en memoria de trabajo, pobre control inhibitorio, y problemas en la regulación emocional, cognitiva y conductual. La valoración de las habilidades adaptativas del menor pone en evidencia un nivel bajo de desarrollo de habilidades adaptativas relacionadas con el área comunicativa tanto en la expresión como en la comprensión del lenguaje, y moderadamente bajo en habilidades de socialización. Conclusiones. Se evidencia que, si bien el niño tiene un rendimiento promedio en la mayoría de las tareas cognitivas de FE, tiene dificultad en la realización de actividades de la vida cotidiana que involucran a las FE en la regulación de la conducta. En este sentido, es recomendable la complementariedad de instrumentos cognitivos de FE y medidas conductuales con valor ecológico en las evaluaciones realizadas a pacientes con comorbilidad entre TEA y TDAH.

El artículo analiza las afectaciones de la pandemia por Covid-19 en el mercado laboral, en las actividades económicas y familiares de las mujeres en la ciudad de Popayán y las empresarias del Cauca. Para ello, se estima un modelo logit, además se utilizan fuentes de información primaria y secundaria provenientes del DANE, ANIF y Confecámaras. Los resultados muestran que la brecha en participación laboral para las mujeres se incrementó en aproximadamente 1,2 puntos porcentuales, los datos obtenidos, a través de 298 empresarias encuestadas, revelan que estas han tenido que cambiar su actividad económica, hacer uso de las TIC y otras estrategias para adaptarse a las dificultades coyunturales. Finalmente se observó un importante cambio en la actividad económica de las empresas, pocos emprendimientos dirigidos por mujeres están operando a plena capacidad, mientras que la mayoría se encuentran operando con restricciones, cerrados o en proceso de cierre.

Genome mining of the bacterial strains Pseudomonas sp. SH-C52 and Pseudomonas fluorescens DSM 11579 showed that both strains contained a highly similar gene cluster encoding an octamodular nonribosomal peptide synthetase (NRPS) system which was not associated with a known secondary metabolite. Insertional mutagenesis of an NRPS component followed by comparative profiling led to the discovery of the corresponding novel linear octalipopeptide thanafactin A, which was subsequently isolated and its structure determined by two-dimensional NMR and further spectroscopic and chromatographic methods. In bioassays, thanafactin A exhibited weak protease inhibitory activity and was found to modulate swarming motility in a strain-specific manner.

Genome mining of the bacterial strains

TiO2 is a semiconductor widely used in photocatalytic degradation of organic pollutants due to its band gap energy. However, its absorption range is restricted only to UV radiation that is less than 10% of solar light. With the aim of increasing the adsorption area on TiO2 nanoparticles a modified sol-gel method was used to produce a smaller particle size, and to extend the absorption range to the visible spectrum, TiO2 nanoparticles were synthesized with different CeO2 contents to generate semiconductor heterojunction between them. The crystallographic, morphological, and optoelectronic characteristics of these TiO2-CeO2 nanoparticles were studied, and two crystalline phases were differentiated: anatase for TiO2 and fluorite for CeO2. An increase in the CeO2 content produced crystallite sizes between 6.5 nm and 12.0 nm. TiO2-CeO2 nanoparticles showed morphological properties such as small particle size, heterogeneous surface and high BET surface area compared to bare commercial TiO2. These features involve a positive effect of CeO2 in TiO2 nanoparticles surface, thus TiO2-CeO2 nanoparticles exhibit enhanced optoelectronic properties caused by a decrease in the effective band gap and red-shift in the electromagnetic spectrum. In addition, methylene blue degradation showed that TiO2-CeO2 nanoparticles are suitable for high photocatalytic activity application under visible light.

Here, we report a 4.3-Mb draft genome sequence of a potential new Ochrobactrum species, which clarified its taxonomic position and gave insight into the complete secondary metabolite production capacity of the strain.

Here, we report a 4.3-Mb draft genome sequence of a potential new Ochrobactrum species, which clarified its taxonomic position and gave insight into the complete secondary metabolite production capacity of the strain.

Pseudomonas fluorescens DSM 11579 is known to be a producer of the lipopeptides brabantamide and thanamycin. Its draft genome gives insight into the complete secondary metabolite production capacity of the strain and builds the basis for a comparative study with Pseudomonas sp. strain SH-C52, a lipopeptide-producing strain involved in natural disease-suppressive soils.

Pseudomonas fluorescens DSM 11579 is known to be a producer of the lipopeptides brabantamide and thanamycin. Its draft genome gives insight into the complete secondary metabolite production capacity of the strain and builds the basis for a comparative study with Pseudomonas sp. strain SH-C52, a lipopeptide-producing strain involved in natural disease-suppressive soils.

Covering: 2014 to 2019 Nonribosomal peptide synthetases (NRPSs) and polyketide synthases (PKSs) have been the subject of engineering efforts for multiple decades. Their modular assembly line architecture potentially allows unlocking vast chemical space for biosynthesis. However, attempts thus far are often met with mixed success, due to limited molecular compatibility of the parts used for engineering. Now, new engineering strategies, increases in genomic data, and improved computational tools provide more opportunities for major progress. In this review we highlight some of the challenges and progressive strategies for the re-design of NRPSs & type I PKSs and survey useful computational tools and approaches to attain the ultimate goal of semi-automated and design-based engineering of novel peptide and polyketide products.

Summary Streptomyces sp. NTK937, producer of benzoxazole antibiotic caboxamycin, produces in addition a methyl ester derivative, O-methylcaboxamycin. Caboxamycin cluster, comprising one regulatory and nine structural genes, has been delimited, and each gene has been individually inactivated to demonstrate its role in the biosynthetic process. The O-methyltransferase potentially responsible for O-methylcaboxamycin synthesis would reside outside this cluster. Five of the genes, cbxR, cbxA, cbxB, cbxD and cbxE, encoding a SARP transcriptional regulator, salicylate synthase, 3-oxoacyl-ACP-synthase, ACP and amidohydrolase, respectively, have been found to be essential for caboxamycin biosynthesis. The remaining five structural genes were found to have paralogues distributed throughout the genome, capable of partaking in the process when their cluster homologue is inactivated. Two of such paralogues, cbxC’ and cbxI’, coding an AMP-dependent synthetase-ligase and an anthranilate synthase, respectively, have been identified. However, the other three genes might simultaneously have more than one paralogue, given that cbxF (DAHP synthase), cbxG (2,3-dihydro-2,3-dihydroxybenzoate dehydrogenase) and cbxH (isochorismatase) have three, three and five putative paralogue genes, respectively, of similar function within the genome. As a result of genetic manipulation, a novel benzoxazole (3′-hydroxycaboxamycin) has been identified in the salicylate synthase-deficient mutant strain ΔcbxA. 3′-hydroxycaboxamycin derives from the cross-talk between the caboxamycin and enterobactin pathways.

Streptomyces sp. strain Z26 exhibited antifungal activity and turned out to be a producer of the secondary metabolites novonestmycin A and B. The 6.5-Mb draft genome gives insight into the complete secondary metabolite production capacity and builds the basis to find and locate the biosynthetic gene cluster encoding the novonestmycins.

Infection with protozoan parasite Trypanosoma cruzi results in activation of nucleotide-binding domain and leucine-rich repeat containing receptors (NLRs). NLR activation leads to inflammasome formation, the activation of caspase-1, and the subsequent cleavage of IL-1β and IL-18. Considering that inflammasome activation and IL-1β induction by macrophages are key players for an appropriate T cell response, we investigated the relevance of NLR pyrin domain-containing 3 (NLRP3) and caspase-1/11 to elucidate their roles in the induction of different T cell phenotypes and the relationship with parasite load and hepatic inflammation during T. cruzi-Tulahuen strain acute infection. We demonstrated that infected nlrp3-/- and C57BL/6 wild type (WT) mice exhibited similar parasitemia and survival, although the parasite load was higher in the livers of nlrp3-/- mice than in those of WT mice. Increased levels of transaminases and pro-inflammatory cytokines were found in the plasma of WT and nlrp3-/- mice indicating that NLRP3 is dispensable to control the parasitemia but it is required for a better clearance of parasites in the liver. Importantly, we have found that NLRP3 and caspase-1/11-deficient mice differentially modulate T helper (Th1, Th2, and Th17) and cytotoxic T lymphocyte phenotypes. Strikingly, caspase-1/11-/- mice showed the most dramatic reduction in the number of IFN-γ- and IL-17-producing CD4+ and CD8+ T cells associated with higher parasitemia and lower survival. Additionally, caspase-1/11-/- mice demonstrated significantly reduced liver inflammation with the lowest alanine aminotransferase (ALT) levels but the highest hepatic parasitic load. These results unequivocally demonstrate that caspase-1/11 pathway plays an important role in the induction of liver adaptive immunity against this parasite infection as well as in hepatic inflammation. Fil: Paroli, Augusto Fabián. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Córdoba. Centro de Investigaciones en Bioquímica Clínica e Inmunología; Argentina Fil: Gonzalez, Patricia V.. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Córdoba. Centro de Investigaciones en Bioquímica Clínica e Inmunología; Argentina Fil: Díaz Luján, Cintia María. Universidad Nacional de Cordoba. Facultad de Medicina. Instituto de Biologia Celular; Argentina Fil: Onofrio, Luisina Inés. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Córdoba. Centro de Investigaciones en Bioquímica Clínica e Inmunología; Argentina Fil: Arocena, Alfredo Raul. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Córdoba. Centro de Investigaciones en Bioquímica Clínica e Inmunología; Argentina Fil: Cano, Roxana Carolina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Córdoba. Centro de Investigaciones en Bioquímica Clínica e Inmunología; Argentina Fil: Carrera Silva, Eugenio Antonio. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Medicina Experimental. Academia Nacional de Medicina de Buenos Aires. Instituto de Medicina Experimental; Argentina Fil: Gea, Susana. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Córdoba. Centro de Investigaciones en Bioquímica Clínica e Inmunología; Argentina

Chronic obesity and Chagas disease (caused by the protozoan Trypanosoma cruzi) represent serious public health concerns. The interrelation between parasite infection, adipose tissue, immune system and metabolism in an obesogenic context, has not been entirely explored. A novel diet-induced obesity model (DIO) was developed in C57BL/6 wild type mice to examine the effect of chronic infection (DIO+I) on metabolic parameters and on obesity-related disorders. Dyslipidemia, hyperleptinemia, and cardiac/hepatic steatosis were strongly developed in DIO mice. Strikingly, although these metabolic alterations were collectively improved by infection, plasmatic apoB100 levels remain significantly increased in DIO+I, suggesting the presence of pro-atherogenic small and dense LDL particles. Moreover, acute insulin resistance followed by chronic hyperglycemia with hypoinsulinemia was found, evidencing an infection-related-diabetes progression. These lipid and glucose metabolic changes seemed to be highly dependent on TLR4 expression since TLR4-/- mice were protected from obesity and its complications. Notably, chronic infection promoted a strong increase in MCP-1 producing macrophages with a M2 (F4/80+CD11c-CD206+) phenotype associated to oxidative stress in visceral adipose tissue of DIO+I mice. Importantly, infection reduced lipid content but intensified inflammatory infiltrates in target tissues. Thus, parasite persistence in an obesogenic environment and the resulting host immunometabolic dysregulation may contribute to diabetes/atherosclerosis progression. Fil: Cabalén, María Eugenia. Universidad Católica de Córdoba; Argentina Fil: Cabral, Maria Fernanda. Universidad Católica de Córdoba; Argentina Fil: Sanmarco, Liliana Maria. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Córdoba. Centro de Investigaciones en Bioquímica Clínica e Inmunología; Argentina Fil: Andrada, Marta Cecilia. Universidad Católica de Córdoba; Argentina Fil: Onofrio, Luisina Inés. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Córdoba. Centro de Investigaciones en Bioquímica Clínica e Inmunología; Argentina Fil: Ponce, Nicolás Eric. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Córdoba. Centro de Investigaciones en Bioquímica Clínica e Inmunología; Argentina Fil: Aoki, Maria del Pilar. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Córdoba. Centro de Investigaciones en Bioquímica Clínica e Inmunología; Argentina Fil: Gea, Susana. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Córdoba. Centro de Investigaciones en Bioquímica Clínica e Inmunología; Argentina Fil: Cano, Roxana Carolina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Córdoba. Centro de Investigaciones en Bioquímica Clínica e Inmunología; Argentina

Streptomyces sp. Tü 6176, producer of cytotoxic benzoxazoles AJI9561, nataxazole, and 5-hydroxy-nataxazole, has been found to produce a fourth benzoxazole, UK-1. All derive from 3-hydroxy-anthranilate synthesized by the nataxazole biosynthesis machinery. However, biosynthesis of AJI9561, nataxazole, and 5-hydroxy-nataxazole requires 6-methylsalicylic acid also provided by nataxazole biosynthesis pathway, while biosynthesis of UK-1 utilizes salicylic acid produced by a salicylate synthase from the coelibactin biosynthesis pathway. This clearly suggests crosstalk between nataxazole and coelibactin pathways. Overproduction of UK-1 was obtained by growing a nataxazole non-producing mutant (lacking 6-methylsalicylate synthase, NatPK) in a zinc-deficient medium. Furthermore, Streptomyces sp. Tü 6176 also produces the siderophore enterobactin in an iron-free medium. Enterobactin production can be induced in an iron-independent manner by inactivating natAN, which encodes an anthranilate synthase involved in nataxazole production. The results indicate a close relationship between nataxazole, enterobactin and coelibactin pathways through the shikimate pathway, the source of their common precursor, chorismate.

Streptomyces sp. Tü 6176 produces the cytotoxic benzoxazole nataxazole. Bioinformatic analysis of the genome of this organism predicts the presence of 38 putative secondary-metabolite biosynthesis gene clusters, including those involved in the biosynthesis of AJI9561 and its derivative nataxazole, the antibiotic hygromycin B, and ionophores enterobactin and coelibactin. The nataxazole biosynthesis gene cluster was identified and characterized: it lacks the O-methyltransferase gene required to convert AJI9561 into nataxazole. This O-methyltransferase activity might act as a resistance mechanism, as AJI9561 shows antibiotic activity whereas nataxazole is inactive. Moreover, heterologous expression of the nataxazole biosynthesis gene cluster in S. lividans JT46 resulted in the production of AJI9561. Nataxazole biosynthesis requires the shikimate pathway to generate 3-hydroxyanthranilate and an iterative type I PKS to generate 6-methylsalicylate. Production of nataxazole was improved up to fourfold by disrupting one regulatory gene in the cluster. An additional benzoxazole, 5-hydroxynataxazole is produced by Streptomyces sp. Tü 6176. 5-Hydroxynataxazole derives from nataxazole by the activity of an as yet unidentified oxygenase; this implies cross-talk between the nataxazole biosynthesis pathway and an unknown pathway.

Macrophages are the primary immune cells that reside within the myocardium, suggesting that these mononuclear phagocytes are essential in the orchestration of cardiac immunity and homeostasis. Independent of the nature of the injury, the heart triggers leukocyte activation and recruitment. However, inflammation is harmful to this vital terminally differentiated organ with extremely poor regenerative capacity. As such, cardiac tissue has evolved particular strategies to increase the stress tolerance and minimize the impact of inflammation. In this sense, growing evidences show that mononuclear phagocytic cells are particularly dynamic during cardiac inflammation or infection and would actively participate in tissue repair and functional recovery. They respond to soluble mediators such as metabolites or cytokines, which play central roles in the timing of the intrinsic cardiac stress response. During myocardial infarction two distinct phases of monocyte influx have been identified. Upon infarction, the heart modulates its chemokine expression profile that sequentially and actively recruits inflammatory monocytes, first, and healing monocytes, later. In the same way, a sudden switch from inflammatory macrophages (with microbicidal effectors) toward anti-inflammatory macrophages occurs within the myocardium very shortly after infection with Trypanosoma cruzi, the causal agent of Chagas cardiomyopathy. While in sterile injury, healing response is necessary to stop tissue damage; during an intracellular infection, the anti-inflammatory milieu in infected hearts would promote microbial persistence. The balance of mononuclear phagocytic cells seems to be also dynamic in atherosclerosis influencing plaque initiation and fate. This review summarizes the participation of mononuclear phagocyte system in cardiovascular diseases, keeping in mind that the immune system evolved to promote the reestablishment of tissue homeostasis following infection/injury, and that the effects of different mediators could modulate the magnitude and quality of the immune response. The knowledge of the effects triggered by diverse mediators would serve to identify new therapeutic targets in different cardiovascular pathologies. Fil: Sanmarco, Liliana Maria. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Córdoba. Centro de Investigaciones en Bioquímica Clínica e Inmunología; Argentina Fil: Eberhardt, Natalia. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Córdoba. Centro de Investigaciones en Bioquímica Clínica e Inmunología; Argentina Fil: Ponce, Nicolás Eric. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Córdoba. Centro de Investigaciones en Bioquímica Clínica e Inmunología; Argentina Fil: Cano, Roxana Carolina. Universidad Nacional de Córdoba; Argentina. Universidad Catolica Argentina; Argentina Fil: Bonacci, Gustavo Roberto. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Córdoba. Centro de Investigaciones en Bioquímica Clínica e Inmunología; Argentina Fil: Aoki, Maria del Pilar. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Córdoba. Centro de Investigaciones en Bioquímica Clínica e Inmunología; Argentina

Damaged cells release the pro-inflammatory signal ATP, which is degraded by the ectonucleotidases CD39 and CD73 to the anti-inflammatory mediator adenosine (ADO). The balance between ATP/ADO is known to determine the outcome of inflammation/infection. However, modulation of the local immune response in different tissues due to changes in the balance of purinergic metabolites has yet to be investigated. Here, we explored the contribution of CD73-derived ADO on the acute immune response against Trypanosoma cruzi parasite, which invades and proliferates within different target tissues. Deficiency of CD73 activity led to an enhanced cardiac microbicidal immune response with an augmented frequency of macrophages with inflammatory phenotype and increased CD8+ T cell effector functions. The increment of local inducible nitric oxide (NO) synthase (iNOS)+ macrophages and the consequent rise of myocardial NO production in association with reduced ADO levels induced protection against T. cruzi infection as observed by the diminished cardiac parasite burden compared to their wild-type (WT) counterpart. Unexpectedly, parasitemia was substantially raised in CD73KO mice in comparison with WT mice, suggesting the existence of tissue reservoir/s outside myocardium. Indeed, CD73KO liver and visceral adipose tissue (VAT) showed increased parasite burden associated with a reduced ATP/ADO ratio and the lack of substantial microbicidal immune response. These data reveal that the purinergic system has a tissue-dependent impact on the host immune response against T. cruzi infection.

63,5 % de l’ensemble des syndicalistes tues au monde entre 1999 et 2009 ont trouve la mort en Colombie. L’objet de cet article n’est pas tant de chercher les causes de cette violence que de comprendre les strategies individuelles et collectives qui font perdurer le travail militant dans un tel contexte de repression sociopolitique. En examinant ces mecanismes, cet article met en lumiere les aspects les plus saillants des investissements et des retributions que les militants tirent de leur travail, ainsi que les strategies deployees par l’organisation pour encourager et conserver les engagements. La construction d’un ethos aux traits contestataires serait ainsi le fruit de la repression et la source principale a l’origine du travail militant dans un contexte de polarisation croissante.

Summary We have modelled elaidyl-sulfamide (ES), a sulfamoyl analogue of oleoylethanolamide (OEA). ES is a lipid mediator of satiety that works through the peroxisome proliferator-activated receptor alpha (PPARα). We have characterised the pharmacological profile of ES (0.3-3 mg/kg) by means of in silico molecular docking to the PPARα receptor, in vitro transcription through PPARα and in vitro and in vivo administration to obese rats. ES interacts with the PPARα binding site in a similar way to OEA, is capable of activating PPARα and also reduces feeding in a dose-dependent manner when administered to food-deprived rats. When ES was given to obese male rats for 7 days, it reduced feeding and weight gain, lowered plasma cholesterol and reduced the plasmatic activity of transaminases, indicating a clear improvement of hepatic function. This pharmacological profile is associated with the modulation of both cholesterol and lipid metabolism regulatory genes, including the sterol response element-binding proteins SREBF1/2 and their regulatory proteins INSIG1/2 in liver and white adipose tissues. ES treatment induced the expression of thermogenic regulatory genes, including the uncoupling proteins UCP1/2/3 in brown adipose tissue and UCP3 in white adipose tissue. However, its chronic administration resulted in hyperglycaemia and insulin resistance, which represent a constraint for its potential clinical development.

Local innate immunity plays a key role in initiating and coordinating homeostatic and defense responses in the heart. We have previously reported that the cardiotropic parasite Trypanosoma cruzi, the etiological agent of Chagas disease, protects cardiomyocytes against growth factor deprivation-induced apoptosis. In this study, we investigated cardiomyocyte innate immune response to T. cruzi infection and its role in cellular protection from apoptosis. We found that Toll-like receptor (TLR) 2-expressing cellswere strongly increased by the parasite in BALB/c neonatal mouse cardiomyocyte cultures. Using a dominant-negative system, we showed that TLR2 mediated cardiomyocyte survival and the secretion of interleukin (IL) 6, which acted as an essential antiapoptotic factor. Moreover, IL6 released by infected cells, as well as the recombinant bioactive cytokine, induced the phosphorylation of the signal transducers and activators of transcription-3 (STAT3) in cultured cardiomyocytes. In accord with the in vitro results, during the acute phase of the infection, TLR2 expression increased 2.9-fold and the antiapoptotic factor Bcl-2 increased 4.5-fold in the cardiac tissue. We have clearly shown a cross-talk between the intrinsic innate response of cardiomyocytes and the pro-survival effect evoked by the parasite. © Springer-Verlag 2011. Fil: Ponce, Nicolás Eric. Universidad Nacional de Cordoba. Facultad de Cs.quimicas. Departamento de Bioquimica Clinica. Cat.de Inmunologia; Argentina Fil: Cano, Roxana Carolina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Córdoba. Centro de Investigaciones en Bioquímica Clínica e Inmunología; Argentina Fil: Carrera Silva, Eugenio Antonio. University of Yale; Estados Unidos Fil: Lima, Ana Paula. Universidade Federal do Rio de Janeiro; Brasil Fil: Gea, Susana. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Córdoba. Centro de Investigaciones en Bioquímica Clínica e Inmunología; Argentina Fil: Aoki, Maria del Pilar. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Córdoba. Centro de Investigaciones en Bioquímica Clínica e Inmunología; Argentina

We evaluated the host metabolic response to chronic varied stress during infection with the fungus Candida albicans. We used four groups of female Wistar rats: normal uninfected and unstressed, stressed, C. albicans infected and infected, and stressed. Infected rats reacted with rapid metabolic adjustments, evident as anorexia and body weight loss, partly mediated by glucocorticoids and TNF-á. Higher circulating levels of IL-6 and glucose (p < 0.05) revealed the progress and catabolic effect of the inflammatory response. Infected and stressed rats instead showed anorexia associated with infection and weight loss as the result of reduced food intake. This group exhibited a prompt reduction in circulating leptin on day 3 (p < 0.05), reduction in glucose levels and depletion of hepatic glycogen depots. We also evaluated the contribution of TNF-á, glucocorticoids, and food deprivation to liver damage. Lipid peroxidation in liver detected in the infected and infected-stressed groups was exacerbated by the glucocorticoid receptor antagonist RU 486, suggesting the modulatory activity of glucocorticoids, while hepatic fat accumulation and glycogen depletion decreased with anti-TNF-á treatment. Food deprivation exacerbated liver injury while the response to stress contributed to greater fungal colonization. Our findings emphasize the impact of metabolic alterations on tissue damage when the host immune activity is modulated by stress mediators. Fil: Rodriguez Galan, Maria Cecilia. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Córdoba. Centro de Investigaciones en Bioquímica Clínica e Inmunología; Argentina Fil: Porporatto, Carina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Centro de Investigaciones y Transferencia de Villa María. Universidad Nacional de Villa María. Centro de Investigaciones y Transferencia de Villa María; Argentina Fil: Sotomayor, Claudia Elena. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Córdoba. Centro de Investigaciones en Bioquímica Clínica e Inmunología; Argentina Fil: Cano, Roxana Carolina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Córdoba. Centro de Investigaciones en Bioquímica Clínica e Inmunología; Argentina Fil: Cejas, Hugo. Universidad Nacional de Córdoba. Facultad de Medicina; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Córdoba. Centro de Investigaciones en Bioquímica Clínica e Inmunología; Argentina Fil: Correa, Silvia Graciela. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Córdoba. Centro de Investigaciones en Bioquímica Clínica e Inmunología; Argentina

The yeast Candida albicans belongs to the microflora of healthy individuals, although it can infect a variety of tissues ensuing changes in the host's immune status. To evaluate the effect of neuroendocrine input on the early immune response during the fungal infection, we use a 3-day paradigm of chronic varied stress in Wistar rats infected with C. albicans. We find that stress mediators contribute to the spread of the fungus and downregulate critical functions of phagocytic cells at the infection site. Phenotypic and functional alterations of effector cells account for the decreased resistance to candidiasis and condition the development of the adaptive response. Stressed hosts exhibit a higher fungal burden in kidneys and livers associated with hyphal forms. The hepatic inflammatory reaction is compromised with severe steatosis, increment of functional enzymes, marked lipid peroxidation and hepatocyte apoptosis. Moreover, infection-related sickness symptoms are significantly increased by exposure to stress with anorexia, weight loss, lack of leptin and depletion of glycogen depots. Food deprivation exacerbates the liver injury. Stress mediators perturb the complex immune and metabolic program that operates early during fungal spread and promotes severe tissue damage Fil: Rodriguez Galan, Maria Cecilia. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Córdoba. Centro de Investigaciones en Bioquímica Clínica e Inmunología; Argentina Fil: Sotomayor, Claudia Elena. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Córdoba. Centro de Investigaciones en Bioquímica Clínica e Inmunología; Argentina Fil: Cano, Roxana Carolina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Córdoba. Centro de Investigaciones en Bioquímica Clínica e Inmunología; Argentina Fil: Porporatto, Carina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Córdoba. Centro de Investigaciones en Bioquímica Clínica e Inmunología; Argentina Fil: Renna, Maria Sol. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Córdoba. Centro de Investigaciones en Bioquímica Clínica e Inmunología; Argentina Fil: Paraje, María Gabriela. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Córdoba. Centro de Investigaciones en Bioquímica Clínica e Inmunología; Argentina Fil: Cejas, Hugo. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Córdoba. Centro de Investigaciones en Bioquímica Clínica e Inmunología; Argentina Fil: Correa, Silvia Graciela. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Córdoba. Centro de Investigaciones en Bioquímica Clínica e Inmunología; Argentina

The amount of experimental restraints e.g., NOEs is often too small for calculating high quality three- dimensional structures by restrained molecular dynamics. Considering this as a typical missing value problem we propose here a model based data imputation technique that should lead to an improved estimation of the correct structure. The novel automated method implemented in AUREMOL makes a more efficient use of the experimental information to obtain NMR structures with higher accu- racy. It creates a large set of substitute restraints that are used either alone or together with the experimental restraints. The new approach was successfully tested on three examples: firstly, the Ras-binding domain of Byr2 from Schizosaccharomyces pombe, the mutant HPr (H15A) from Staphylococcus aureus, and a X-ray structure of human ubiquitin. In all three examples, the quality of the resulting final bundles was improved considerably by the use of additional substitute restraints, as assessed quanti- tatively by the calculation of RMSD values to the ''true'' structure and NMR R-factors directly calculated from the original NOESY spectra or the published diffraction data.

We previously reported the synthesis of three new opioid agonists as well as their in vitro and in vivo activity [Girón, R., Abalo, R., Goicoechea, C., Martín, M.I., Callado, L.F., Cano, C., Goya, P., Jagerovic, N. 2002. Synthesis and opioid activity of new fentanyl analogs. Life Sci. 71, 1023-1034]. One of them, N-[1-phenylpyrazol-3-yl]-N-[1-(2-phenethyl)-4-piperidyl)] propenamide (IQMF-4), showed an interesting antinociceptive activity. Intraperitoneally (i.p.) administered, it was as effective as fentanyl or morphine, being less potent than fentanyl but more so than morphine. The aim of the present work was to evaluate its antinociceptive effect by different routes of administration, using the hot plate test, and to investigate possible side effects, such as tolerance and withdrawal, in vitro, using the myenteric plexus-longitudinal muscle strip preparation from guinea pig ileum, and in vivo, using the hot plate test. IQMF-4 was more potent than morphine when administered per os (p.o.), but less potent when administered intracerebroventricularly (i.c.v.). By both routes, fentanyl is more potent that IQMF-4. When IQMF-4 was administered i.p., naloxone methiodide, a peripherally acting antagonist, was able to completely block its antinociceptive effect, whereas, after i.c.v. administration, the blockade was only partial. An interesting feature of the new compound is that it induces tolerance in vitro but not in vivo. Moreover, though in vitro withdrawal was not different from fentanyl or morphine, in vivo withdrawal symptoms were significantly less frequent in mice treated with IQMF-4 than in those treated with morphine or fentanyl. Although more assays are required, these results show that IQMF-4 appears to be a potent analgesic compound with an interesting peripheral component, and reduced ability to induce dependence. © 2008 Elsevier B.V. All rights reserved.

Increasing evidence demonstrates that generation of extracellular adenosine from ATP, which is hydrolyzed by the CD39/CD73 enzyme pair, attenuates the inflammatory response and deactivates macrophage antimicrobial mechanisms. Although CD73 is emerging as a critical pathway and therapeutic target in cardiovascular disorders, the involvement of this ectonucleotidase during myocardial infection has not been explored. Using a murine model of infection with Trypanosoma cruzi, the causal agent of Chagas cardiomyopathy, we observed a sudden switch from the classical M1 macrophage (microbicidal) phenotype toward an alternative M2 (repairing/anti-inflammatory) phenotype that occurred within the myocardium very shortly after BALB/c mice infection. The observed shift in M1/M2 rate correlated with the cardiac cytokine milieu. Considering that parasite persistence within myocardium is a necessary and sufficient condition for the development of the chronic myocarditis, we hypothesized that CD73 activity may counteract cardiac macrophage microbicidal polarization, rendering the local immune response less effective. In fact, a transient treatment with a specific CD73 inhibitor (adenosine 5′-α,β-methylene-diphosphate) enhanced the microbicidal M1 subset predominance, diminished IL-4– and IL-10–producing CD4+ T cells, promoted a proinflammatory cytokine milieu, and reduced parasite load within the myocardium during the acute phase. As a direct consequence of these events, there was a reduction in serum levels of creatine kinase muscle–brain isoenzyme, a myocardial-specific injury marker, and an improvement in the electrocardiographic characteristics during the chronic phase. Our results demonstrate that this purinergic system drives the myocardial immune response postinfection and harbors a promising potential as a therapeutic target.

Innate and adaptive immunity collaborate in the protection of intracellular pathogens including Trypanosoma cruzi infection. However, the parasite molecules that regulate the host immune response have not been fully identified. We previously demonstrated that the immunisation of C57BL/6 mice with cruzipain, an immunogenic T. cruzi glycoprotein, induced a strong specific T-cell response. In this study, we demonstrated that active immunisation with cruzipain was able to stimulate nitric oxide (NO) production by splenocytes. Immune cells also showed increased inducible nitric oxide synthase protein and mRNA expression. Spleen adherent cells secreted high levels of IFN-gamma and IL-12. Microbicidal activity in vitro was mainly mediated by reactive nitrogen intermediaries and IFN-gamma, as demonstrated by the inhibitory effects of NO synthase inhibitor or by IFN-gamma neutralisation. Specific T-cells were essential for NO, IFN-gamma and TNF-alpha production. Furthermore, we reported that cruzipain enhanced CD80 and major histocompatibility complex-II molecule surface expression on F4/80+ spleen cells. Interestingly, we also showed that cruzipain up-regulated toll like receptor-2 expression, not only in F4/80+ but also in total spleen cells which may be involved in the effector immune response. Our findings suggest that a single parasite antigen such as cruzipain, through adaptive immune cells and cytokines, can modulate the macrophage response not only as antigen presenting cells, but also as effector cells displaying enhanced microbicidal activity with reactive nitrogen intermediary participation. This may represent a mechanism that contributes to the immunoregulatory process during Chagas disease.

Resume En Colombie, les syndicalistes resistent a la repression exercee par plusieurs acteurs (et en particulier par les groupes paramilitaires), qui fait de ce pays l’endroit le plus dangereux au monde pour ces militants. Cet article adopte une approche du syndicalisme en tant que travail militant qui jette des lumieres sur les tactiques collectives pour surmonter la peur liee au risque et qui nous permet de comprendre que les syndicalistes font plus que resister : en se battant contre la repression ils se constituent en veritables sujets politiques.

We have recently reported that Trypanosoma cruzi infection protects cardiomyocytes against apoptosis induced by growth factor deprivation. Cruzipain, a major parasite antigen, reproduced this survival effect by a Bcl-2-dependent mechanism. In this study, we have investigated the molecular mechanisms of cruzipain-induced cardiomyocyte protection. Neonatal BALB/c mouse cardiac myocytes were cultured under minimum serum conditions in the presence of cruzipain or T. cruzi (Tulahuen strain). Some cultures were pretreated with the phosphatidylinositol 3-kinase (PI3K) inhibitor Ly294002 or specific inhibitors of the mitogen-activated protein kinase (MAPK) family members such as the mitogen-activated protein kinase kinase (MEK1) inhibitor PD098059, Jun N-terminal kinase (JNK) inhibitor SP600125, p38 MAPK inhibitor SB203580. Inhibition of PI3K and MEK1 but not JNK or p38 MAPK increased the apoptotic rate of cardiomyocytes treated with cruzipain. Phosphorylation of Akt, a major target of PI3K, and ERK1/2, MEK1-targets, was achieved at 15 min and 5 min, respectively. In parallel, these kinases were strongly phosphorylated by T. cruzi infection. In cultures treated with cruzipain, cleavage of caspase 3 was considerably diminished after serum starvation; Bcl-2 overexpression was inhibited by PD098059 but not by Ly294002, whereas Bad phosphorylation and Bcl-xL expression were increased and differentially modulated by both inhibitors. The results suggest that cruzipain exerts its anti-apoptotic property in cardiac myocytes at least by PI3K/Akt and MEK1/ERK1/2 signaling pathways. We further identified a differential modulation of Bcl-2 family members by these two signaling pathways.

BALB/c mice immunized with cruzipain, a major Trypanosoma cruzi antigen, produce specific and autoreactive immune responses against heart myosin, associated with cardiac functional and structural abnormalities. Preferential activation of the Th2 phenotype and an increase in cell populations expressing CD19+, Mac-1+ and Gr-1+ markers were found in the spleens of these mice. The aim of the present study was to investigate whether cardiac autoimmunity could be induced by cruzipain immunization of C57BL/6 mice and to compare the immune response elicited with that of BALB/c mice. We demonstrate that immune C57BL/6 splenocytes, re-stimulated in vitro with cruzipain, produced high levels of IFNgamma and low levels of IL-4 compatible with a Th1 profile. In contrast to BALB/c mice, spleens from cruzipain immune C57BL/6 mice revealed no significant changes in the number of cells presenting CD19+, Mac-1+ and Gr-1+ markers. An increased secretion of TGFbeta and a greater number of CD4+ TGFbeta+ cells were found in immune C57BL/6 but not in BALB/c mice. These findings were associated with the lack of autoreactive response against heart myosin and a myosin- or cruzipain-derived peptide. Thus, the differential immune response elicited in C57BL/6 and BALB/c mice upon cruzipain immunization is implicated in the resistance or pathogenesis of experimental Chagas' disease.

The oxalate salts and free bases of fentanyl and N-[1-(2-phenylethyl)-4-piperidyl]-N-(1-phenyl-4-pyrazolyl)propanamide, a new lead compound for long-acting analgesia, have been characterized by (1)H- and (13)C-NMR spectroscopy. The crystal structure of the hydrochloride of N-[1-(2-phenylethyl)-4-piperidyl]-N-(1-phenyl-4-pyrazolyl)propanamide monohydrate has been determined. Two centrosymmetrically related cations, joined through C(phenyl)-H em leader pi contacts, encapsulate a large void that contains pairs of anions and bridged water molecules into a zero-dimensional (0D) supramolecular motif. The cations are linked to this framework via N(+)H em leader Cl(-) contacts. GIAO/B3LYP calculations have been carried out to compare the experimental (13)C chemical shifts with the absolute shieldings thus calculated. The protonation of both molecules takes place on the piperidine ring (axial protonation), as has been verified both in the solid state (X-ray) and in solution (NMR).

Background The immune mechanisms underlying experimental non-alcoholic steatohepatitis (NASH), and more interestingly, the effect of T. cruzi chronic infection on the pathogenesis of this metabolic disorder are not completely understood. Methodology/Principal Findings We evaluated immunological parameters in male C57BL/6 wild type and TLR4 deficient mice fed with a standard, low fat diet, LFD (3% fat) as control group, or a medium fat diet, MFD (14% fat) in order to induce NASH, or mice infected intraperitoneally with 100 blood-derived trypomastigotes of Tulahuen strain and also fed with LFD (I+LFD) or MFD (I+MFD) for 24 weeks. We demonstrated that MFD by itself was able to induce NASH in WT mice and that parasitic infection induced marked metabolic changes with reduction of body weight and steatosis revealed by histological studies. The I+MFD group also improved insulin resistance, demonstrated by homeostasis model assessment of insulin resistance (HOMA-IR) analysis; although parasitic infection increased the triglycerides and cholesterol plasma levels. In addition, hepatic M1 inflammatory macrophages and cytotoxic T cells showed intracellular inflammatory cytokines which were associated with high levels of IL6, IFNγ and IL17 plasmatic cytokines and CCL2 chemokine. These findings correlated with an increase in hepatic parasite load in I+MFD group demonstrated by qPCR assays. The recruitment of hepatic B lymphocytes, NK and dendritic cells was enhanced by MFD, and it was intensified by parasitic infection. These results were TLR4 signaling dependent. Flow cytometry and confocal microscopy analysis demonstrated that the reactive oxygen species and peroxinitrites produced by liver inflammatory leukocytes of MFD group were also exacerbated by parasitic infection in our NASH model. Conclusions We highlight that a medium fat diet by itself is able to induce steatohepatitis. Our results also suggest a synergic effect between damage associated with molecular patterns generated during NASH and parasitic infection, revealing an intense cross-talk between metabolically active tissues, such as the liver, and the immune system. Thus, T. cruzi infection must be considered as an additional risk factor since exacerbates the inflammation and accelerates the development of hepatic injury., Author Summary Chagas disease caused by the protozoan parasite T. cruzi is a neglected tropical disease widespread in Latin America, and non-alcoholic steatohepatitis constitutes a prominent health concern with increasing incidence of obesity and diabetes worldwide. Parasitic infection induced marked metabolic changes, improved insulin resistance, but it increased the triglycerides and cholesterol plasma levels. Our findings demonstrate for the first time a synergic effect between the hepatic damage caused during the steatohepatitis process (generated by a medium fat diet) and the exacerbated inflammation triggered by the chronic parasitic infection, revealing an intense cross-talk between the liver, a metabolically active tissue, and the immune system. We propose that T. cruzi infection must be considered as an additional risk factor for this metabolic disorder. These findings may provide new insight for the American Trypanosomiasis associated with the steatohepatitis.