Fig. 4

Trypanosoma cruzi infection alters ECM components expression by CFs. a, d, g Immunoblotting performed to ECM proteins in CFs cultures and their densitometric analyses plotted as graphs. a-c Fibronectin (FN) expression was increased after 6, 24 and 48 h of T. cruzi infection, followed by a decrease at 72 hpi. Immunofluorescence for FN (green) showed that at 24 hpi T. cruzi-infected cultures (c) had higher deposition of this protein (arrows) when compared to uninfected dishes (b). d-f Laminin (LN) is increased after 6 and 24 h of infection as shown by immunoblot. e and f show immunolocalization of LN in CF. In non-infected cultures LN showed a cytoplasmic distribution (in green). At 24 hpi, infected cultures displayed patches of cells with a more intense signal for LN (arrows), that corresponded with parasitized cells (arrowheads for intracellular amastigotes). g-i Collagen IV (COL) is transiently increased by T. cruzi infection. Immunoblot for COL showed a 1.5-fold increase at 6 hpi compared to uninfected cultures (g). COL had a fibrillar pattern in uninfected cultures (arrows in h) while in T. cruzi infection (6 hpi), COL immunoreactivity was increased. GAPDH was used as loading control. DAPI stained the DNA of host cells and of intracellular parasites (arrowheads) and is shown in red (pseudo-color). Values are expressed as fold change of means of the analyzed samples by means of their respective controls (NI) ± SEM. Representative data from three independent experiments. *P < 0.05 (stimulus), **P < 0.01 (stimulus), ***P < 0.001 (stimulus), &&& P < 0.001 (inhibition). One-way ANOVA with Tukey’s post-hoc test. Abbreviations: NI, non-infected cultures; Y, T. cruzi-infected cultures. Scale-bars: 20 μm