Central role of extracellular signal-regulated kinase and Toll-like receptor 4 in IL-10 production in regulatory dendritic cells induced by Trypanosoma cruzi.

Several Trypanosoma cruzi molecules that stimulate macrophages activity were described as Toll-like receptor 2 (TLR2) ligands. Besides, the models of dendritic cells (DC) are poorly characterised. We have previously demonstrated that live-trypomastigotes (Tp) plus lipopolysaccharide (LPS) induce DC with tolerogenic properties that produce high levels of interleukin (IL)-10 and an impaired capacity to induce lymphoproliferation. Here, we show that the regulatory phenotype was observed with heat-killed trypomastigotes (Tphk) stimulation, ruling out DC infection. T. cruzi induced a particular DC activation state increasing LPS-activation of extracellular regulated kinase (ERK) 1/2 and signal transducer and activator of transcription (STAT) 3. Inhibition of ERK down-regulated IL-10 production and restored DC stimulatory capacity, showing the importance of this pathway in the DC modulation. A recent work shows that signalling via TLR4 and TLR2 induces a synergism in anti-inflammatory cytokine production in murine DC. Upon TLR2 and TLR4 stimulation using Pam(3)Cys or LPS and Tphk in DC from TLR2 knock out (KO) or TLR4-mutant mice, we showed that high levels of IL-10 were independent of TLR2 but associated with TLR4 and NF-kappaB signallization. Although sialic acid has been described as a molecule responsible of DC inhibition, we determine that it is not associated with T. cruzi-IL-10 modulatory response. In conclusion, all these findings demonstrate a key role of ERK and TLR4 in association with NF-kappaB in IL-10 modulation induced by T. cruzi and suggest that this regulatory effect involves parasite-DC interactions not described yet.

Evidence for a relationship between bovine erythrocyte lipid membrane peculiarities and immune pressure from ruminal ciliates.

Erythrocytes of bovines and other ruminants have a strikingly anomalous phospholipid composition, with low or absent phosphatidylcholine (PC) together with high sphingomyelin (SM) content. Here, we report the presence in normal bovine serum of high levels of anti-phospholipid antibodies of IgM isotype against, PC and the phosphono analogue of phosphatidylethanolamine, aminoethylphosphonolipid (AEPL), normally produced by rumen ciliates. In contrast, no antibodies were detected against SM or N-acyl-phosphatidylethanolamine (NAPE), the major components of bovine erythrocytes. In addition, we found that exposure of the ciliate Tetrahymena thermophila to bovine serum results in rapid lysis, an effect that was inhibited by adsorption of the serum with SM/AEPL liposomes. Furthermore, incubation with bovine serum had a similar effect on freshly obtained ruminal ciliates, and the lytic activity was eliminated by pre-adsorption of the serum with SM/PE liposomes. The ruminant mode of life with its concomitant ciliate fauna is hereby linked to the peculiar conformation of bovine erythrocyte membranes. We propose that the unique phospholipid composition of bovine erythrocytes appears as an evolutionary adaptation to tolerate the lytic effects of anti-phospholipid antibodies generated against AEPL, a membrane component of the huge mass of ruminal ciliates, necessary commensals of this group of mammals.

Free fatty acids induce cell differentiation to infective forms in Trypanosoma cruzi.

Intestinal extracts of Triatoma infestans induce cell differentiation of Trypanosoma cruzi epimastigotes into the infective metacyclic form. Part of this effect can be explained by the presence of haemoglobin fragments, which stimulate trypanosomal adenylate cyclase. In this work we examined the metacyclogenic activity of lipids present in this intestinal extract. We found that lipid extracts of the intestinal extract have significant stimulatory effects that reside with the free-fatty-acid fraction, especially oleic acid. These compounds stimulate de novo diacylglycerol formation and protein kinase C activity in the parasite. Moreover, metacyclogenesis is stimulated by phorbol esters and cell-permeant diacylglycerol, while protein kinase C down-regulation or incubation with inhibitors of this kinase abrogates this effect. These results indicate that free fatty acids are a novel signal, inducing metacyclogenesis, acting through a pathway involving diacylglycerol biosynthesis and protein kinase C activation.

An alpha D-globin fragment from Triatoma infestans hindgut stimulates Trypanosoma cruzi adenylyl cyclase and promotes metacyclogenesis

A peptide from hindguts of the Triatoma infestans, the hematophagous Chagas' insect vector, activates adenylyl cyclase activity in Trypanosoma cruzi epimastigote membranes and stimulates the in vitro differentiation of epimastigotes (proliferative and non-infectious forms) to metacyclic trypomastigotes (non-proliferative and infectious forms). The peptide was purified from hindguts of insects fed two days before with chicken blood. After purification, the peptide showed upon SDS-PAGE a single band of about 10 kDa. The sequence for 20 residues of the amino terminus of this peptide was: H2N-Met-Leu-Thr-Ala-Glu-Asp-Lys-Lys-Leu-Ile-Gln-Gln-Ala-Trp-Glu-Lys-Ala- Ala-Ser-His. This sequence corresponds to the amino terminus of chicken alpha D-globin. A synthetic peptide with the sequence of the 40 amino acids corresponding to the amino terminus of alpha D-globin, also stimulated T. cruzi adenylyl cyclase activity and promoted metacyclogenesis

Caracterización inmunobiologica de una cepa presuntamente atipica de Trypanosoma cruzi / Immunobiological characterization of a presumed atypical straim of Trypanosoma cruzi

Se analisaron para la cepa LP de Trypanosoma cruzi algunas características biológicas e inmunológicas y se las comparó con las presentadas por cepas de referencia, RA y CA-I. La cepa LP produjo niveles de parasistemia próximoss a los inducidos por la RA pero, asemejándose a la cepa CA-I, la elevada parasitemia fue persistente y los ratones sobrevivieron a la infección aguda; sin embargo, alrededor de 40-45 días pi los animales inoculados con LP presentaron una caída en crisis de los parásitos circulantes. Dosis supresoras de infurtimox retardaron la positivación de la parasitemia en los 3 casos y los valores máximos alcanzados fueron una unidad logarítmica menor comparado con los controles sin tratamiento. La respuesta inmune estimulada por la cepa LP en conejo y en ratón fue positiva para las pruebas serológicas convencionales (IFA y AD) al igual que lo ocurrido con las otras 2 cepas. En el caso de la IFA todos los inmunosueros fueron reactivos con cualquiera de los 4 antígenos utilizados (epimastigotes de LP, RA, CA-I y Tulahuén). La cepa LP también estimuló la formación de anticuerpos capaces de interactuar ccon la forma circulante del parásito modificando su virulencia o produciendo su lisis; estos anticuerpos se detectaron tardíamente, lo que podría justificar la prolongada parasitemia presentada por los ratones inoculados con LP. por los resultados aquí obtenidos el comportamiento atípico comunicado previamente para esta cepa dependería de la asociación hospedeiro-parásito más que de características propias de la población parasitaria

Caracterización inmunobiologica de una cepa presuntamente atipica de Trypanosoma cruzi / Immunobiological characterization of a presumed atypical straim of Trypanosoma cruzi

Se analisaron para la cepa LP de Trypanosoma cruzi algunas características biológicas e inmunológicas y se las comparó con las presentadas por cepas de referencia, RA y CA-I. La cepa LP produjo niveles de parasistemia próximoss a los inducidos por la RA pero, asemejándose a la cepa CA-I, la elevada parasitemia fue persistente y los ratones sobrevivieron a la infección aguda; sin embargo, alrededor de 40-45 días pi los animales inoculados con LP presentaron una caída en crisis de los parásitos circulantes. Dosis supresoras de infurtimox retardaron la positivación de la parasitemia en los 3 casos y los valores máximos alcanzados fueron una unidad logarítmica menor comparado con los controles sin tratamiento. La respuesta inmune estimulada por la cepa LP en conejo y en ratón fue positiva para las pruebas serológicas convencionales (IFA y AD) al igual que lo ocurrido con las otras 2 cepas. En el caso de la IFA todos los inmunosueros fueron reactivos con cualquiera de los 4 antígenos utilizados (epimastigotes de LP, RA, CA-I y Tulahuén). La cepa LP también estimuló la formación de anticuerpos capaces de interactuar ccon la forma circulante del parásito modificando su virulencia o produciendo su lisis; estos anticuerpos se detectaron tardíamente, lo que podría justificar la prolongada parasitemia presentada por los ratones inoculados con LP. por los resultados aquí obtenidos el comportamiento atípico comunicado previamente para esta cepa dependería de la asociación hospedeiro-parásito más que de características propias de la población parasitaria (AU)