Host cell infection by the intracellular pathogen, Trypanosoma cruzi, involves activation of signaling pathways, cytoskeletal reorganization, and targeted recruitment of host cell lysosomes. To determine the consequences of T. cruzi invasion on host cell gene expression, high density microarrays consisting of ϳ27,000 human cDNAs were hybridized with fluorescent probes generated from T. cruzi-infected human fibroblasts (HFF) at early time points following infection (2-24 h). Surprisingly, no genes were induced >2-fold in HFF between 2 and 6 h post-infection (hpi) in repeated experiments while immediate repression of six host cell transcripts was observed. A significant increase in transcript abundance for 106 host cell genes was observed at 24 hpi. Among the most highly induced is a set of interferon-stimulated genes, indicative of a type I interferon (IFN) response to T. cruzi. In support of this, T. cruzi-infected fibroblasts begin to secrete IFN␤ at 18 hpi following the induction of IFN␤ transcripts. As compared with global transcriptional responses evoked by other intracellular pathogens, T. cruzi is a stealth parasite that elicits few changes in host cell transcription during the initiation of infection.Global transcriptional responses elicited in mammalian cells by pathogenic organisms are predicted to provide a unique signature of the particular interaction (1). The recent application of oligonucleotide and cDNA microarray technologies toward the study of host-pathogen interactions has permitted rapid and unbiased examination of changes in expression of a large number of genes at the transcript level (2). Microarray analysis of host cell gene expression following infection with viral (3-7), bacterial (8 -12), fungal (13), and protozoan (14) pathogens has revealed complex and diverse transcriptional responses to these infectious agents. Data bases generated from these, and future, studies will provide an invaluable resource for the functional characterization of host cell pathways required to facilitate pathogen survival and for the further understanding of host defense mechanisms (1,15,16).Trypanosoma cruzi is a hemoflagellate protozoan parasite that causes Chagas' disease in humans. Key steps in the pathogenesis of disease include host cell penetration by T. cruzi and replication in the host cell cytoplasm. Host cell invasion, which is initiated following attachment of motile T. cruzi trypomastigotes to the host cell surface, is a slow, active process requiring ϳ5-10 min for completion (17). Parasite internalization coincides with the formation of a nascent parasitophorous vacuole, generated as a result of targeted fusion of host cell lysosomes with the plasma membrane (18). Signaling pathways that are rapidly activated in both the host cell and the parasite are known to regulate T. cruzi entry into mammalian cells. To further dissect the molecular events regulating early T. cruzi-host cell interactions, we have employed cDNA microarray hybridization to define the temporal host cell transcriptional respon...

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We have previously reported that ACTH activates a phospholipase C that hydrolyzes glycosylphosphatidylinositol (GPI), which would release inositolphosphoglycan (IPG) to the extracellular medium, and that an IPG purified from Trypanosoma cruzi is able to inhibit ACTH-mediated steroid production in adrenocortical cells. In the present paper, it was found that anti-inositolphosphoglycan antibodies (anti-CRD) increased ACTH-mediated corticosterone production, which indicates that an endogenous IPG is a physiological inhibitor of ACTH response. On the other hand, we investigated the release to the extracellular medium of the GPI-anchored enzyme, alkaline phosphatase, by ACTH. We found that: (a) the released enzyme appeared in the aqueous phase after Triton X-114 partitioning, consistent with loss of the GPI, (b) the phospholipase C inhibitor, U73122, impaired the release of the enzyme by the hormone and (c) two inhibitors of IPG uptake, inositol 2-monophosphate and 2 M NaCl, increased the amount of alkaline phosphatase in the extracellular medium. These results suggest that ACTH releases alkaline phosphatase by activation of a phospholipase C. Dibutyryladenosine-3',5'-cyclic monophosphate (db-cAMP) was able to increase the release of alkaline phosphatase from adrenocortical cells and this effect was inhibited by U73122, suggesting that cAMP is involved in the activation of phospholipase C. In addition, it was found that a pertussis-toxin sensitive G-protein is required for ACTH- and db-cAMP-mediated release of alkaline phosphatase and that incorporation of anti-Gi antibodies in adrenocortical cells inhibited the release of alkaline phosphatase by ACTH. Our results suggest that ACTH increases the release of alkaline phosphatase by activation of a phospholipase C through cAMP and Gi which would contribute to produce IPG It was also found that the two inhibitors of IPG uptake, inositol-2-monophosphate and 2 M NaCl, increased the amount of alkaline phosphatase in the extracellular medium of ACTH-treated cells more than in control cells, indicating that ACTH also stimulates the uptake of IPG These data support a role of GPI and the involvement of Gi in ACTH action.