TGF-β-Mediated Sustained ERK1/2 Activity Promotes the Inhibition of Intracellular Growth of Mycobacterium avium in Epithelioid Cells Surrogates

Transforming growth factor beta (TGF-β) has been implicated in the pathogenesis of several diseases including infection with intracellular pathogens such as the Mycobacterium avium complex. Infection of macrophages with M. avium induces TGF-β production and neutralization of this cytokine has been associated with decreased intracellular bacterial growth. We have previously demonstrated that epithelioid cell surrogates (ECs) derived from primary murine peritoneal macrophages through a process of differentiation induced by IL-4 overlap several features of epithelioid cells found in granulomas. In contrast to undifferentiated macrophages, ECs produce larger amounts of TGF-β and inhibit the intracellular growth of M. avium. Here we asked whether the levels of TGF-β produced by ECs are sufficient to induce a self-sustaining autocrine TGF-β signaling controlling mycobacterial replication in infected-cells. We showed that while exogenous addition of increased concentration of TGF-β to infected-macrophages counteracted M. avium replication, pharmacological blockage of TGF-β receptor kinase activity with SB-431542 augmented bacterial load in infected-ECs. Moreover, the levels of TGF-β produced by ECs correlated with high and sustained levels of ERK1/2 activity. Inhibition of ERK1/2 activity with U0126 increased M. avium replication in infected-cells, suggesting that modulation of intracellular bacterial growth is dependent on the activation of ERK1/2. Interestingly, blockage of TGF-β receptor kinase activity with SB-431542 in infected-ECs inhibited ERK1/2 activity, enhanced intracellular M. avium burden and these effects were followed by a severe decrease in TGF-β production. In summary, our findings indicate that the amplitude of TGF-β signaling coordinates the strength and duration of ERK1/2 activity that is determinant for the control of intracellular mycobacterial growth

Transforming Growth Factor-beta(TGF-β)and ERK1/2 activation are involved in Mycobacterium avium control replication in Epithelioid-like Cells

BV UNIFESP: Teses e dissertaçõe

Trypanosoma cruzi cell invasion and traffic: Influence of Coxiella burnetii and pH in a comparative study between distinct infective forms

Previous studies have shown that Coxiella burnetii, an intracellular bacterium that resides within acidified vacuoles with secondary lysosomal characteristics, is an effective modulator of the intracellular traffic of trypomastigote forms of Trypanosoma cruzi.. in addition, vacuolar and cellular pH are related to fusion events that result in doubly infected phagosomes. T cruzi, the etiological agent of Chagas' disease, occurs as different strains grouped in two major phylogenetic lineages: T cruzi I, associated with the sylvatic cycle, and T cruzi II, linked to the human disease. in this work we compared extracellular amastigotes (EA), metacyclic trypomastigotes (NIT) and tissue Culture derived trypomastigotes (TCT) belonging to T. cruzi I or T. cruzi II for their ability to invade and escape from their parasitophorous vacuole (PV), in Vero cells or Vero cells harboring the bacterium, C burnetti. Distinct invasion patterns were observed between different infective stages and between infective forms of different strains. Studies on the transference kinetics revealed that pH modulates the intracellular traffic of each infective stage, but this influence is not exclusive for each phylogenetic group. Endosomal to lysosomal sequential labeling with EEA-1 and LAMP-1 of the PV formed during the entry of each infective form revealed that the phagosome maturation processes are distinct but not strain-dependent. Due to their low hemolysin and trans-sialidase activities, MTs are retained for longer periods in LAMP-1 positive vacuoles. Our results thus suggest that despite the contrasting invasion capabilities, parasites of distinct phylogenetic group behave in similar fashion once inside the host cell. (c) 2007 Elsevier B.V. All rights reserved.Universidade Federal de São Paulo, Escola Paulista Med, Dept Microbiol Immunol & Parasitol, BR-04023062 São Paulo, BrazilUniversidade Federal de São Paulo, Escola Paulista Med, Dept Microbiol Immunol & Parasitol, BR-04023062 São Paulo, BrazilWeb of Scienc

Mammalian cell invasion and intracellular trafficking by Trypanosoma cruzi infective forms

Trypanosoma cruzi, the etiological agent of Chagas’ disease, occurs as different strains or isolates that may be grouped in two major phylogenetic lineages: T. cruzi I, associated with the sylvatic cycle and T. cruzi II, linked to the human disease. In the mammalian host the parasite has to invade cells and many studies implicated the flagellated trypomastigotes in this process. Several parasite surface components and some of host cell receptors with which they interact have been identified. Our work focused on how amastigotes, usually found growing in the cytoplasm, can invade mammalian cells with infectivities comparable to that of trypomastigotes. We found differences in cellular responses induced by amastigotes and trypomastigotes regarding cytoskeletal components and actin-rich projections. Extracellularly generated amastigotes of T. cruzi I strains may display greater infectivity than metacyclic trypomastigotes towards cultured cell lines as well as target cells that have modified expression of different classes of cellular components. Cultured host cells harboring the bacterium Coxiella burnetii allowed us to gain new insights into the trafficking properties of the different infective forms of T. cruzi, disclosing unexpected requirements for the parasite to transit between the parasitophorous vacuole to its final destination in the host cell cytoplasm