Project description:The intracellular pathogen Trypanosoma cruzi secretes an activity that blocks TGF-β-dependent induction of connective tissue growth factor (CTGF/CCN2). Here, we address the mechanistic basis for T. cruzi-mediated interference of CTGF/CCN2 expression by examining host cell signaling pathways and the global inhibitory effect on TGF-β-dependent gene expression. We show that the expression of a discrete subset of TGF-β-inducible genes involved in cell proliferation, wound repair, and immune regulation are blocked by the soluble T. cruzi activity, demonstrating that this parasite-derived activity has broad, but specific effects on fibroblast gene regulation. Primary human fibroblasts were treated with TGF-β, T. cruzi conditioned medium (PCM) and TGF-β/ PCM simultaneously. Untreated cells were also included as controls. Total RNA was extracted and gene expression levels analyzed with Affymetrix microarrays. Three independent biological replicates were included for each type of treatment.
Project description:Primary human skeletal muscle cells (Lonza) were treated with LLC1 conditioned medium, LLC1 conditioned medium plus Calcitriol, LLC1 non-conditioned medium or LLC1 non-conditioned medium plus Calcitriol for a period of 24 hours prior to isolation of RNA.
Project description:Trypanosoma cruzi is an obligate intracellular protozoan parasite that causes human Chagas’ disease, a leading cause of heart failure in Latin America. Using Affymetrix oligonucleotide arrays we screened phenotypically diverse human cells (foreskin fibroblasts, microvascular endothelial cells and vascular smooth muscle cells) for a common transcriptional response signature to T. cruzi. A common feature was a prominent type I interferon response, indicative of a secondary response to secreted cytokines. Using transwell plates to distinguish cytokine-dependent and -independent gene expression profiles in T. cruzi-infected cells, a core cytokine-independent response was identified in fibroblasts and endothelial cells that featured metabolic and signaling pathways involved in cell proliferation, amino acid catabolism and response to wounding. Significant downregulation of genes involved in mitotic cell cycle and cell division predicted that T. cruzi infection impedes cell cycle progression in the host cell.
