Differential cardiomyocyte transcriptomic remodeling during in vitro Trypanosoma cruzi infection using laboratory strains provides implications on pathogenic host responses

Fig. 1. Difference in cardiomyocyte infectivity among Trypanosoma cruzi infection conditions. Percentage of T. cruzi-infected cells (containing intracellular amastigotes) at 24- and 48-h post-infection. hpi hours post-infection

Fig. 2. Cardiomyocyte infectivity among different Trypanosoma cruzi infection conditions. Intracellular amastigote counts per infected cell are plotted at A 24- and B 48-h post-infection. Pairwise comparisons were performed with Mann–Whitney test. Black horizontal bars indicate the mean and SD

Fig. 3. Remodeling of gene expression profiles in cardiomyocytes infected with three different Trypanosoma cruzi strains at two time points post-infection. Y axes indicate the gene expression level of the non-infected control cardiomyocyte, while X-axes indicate gene expression levels of Trypanosoma cruzi-infected cardiomyocytes. The black dashed line, and upper and lower red dashed lines show the line of identity, and the 0.5- and twofold change thresholds, respectively. Each plot represents a differentially expressed gene (DEG). Blue and pink shaded areas represent down- and up-regulated DEGs, respectively. hpi hours post-infection

Fig. 4. Identification of differentially expressed genes (DEGs) of the host cardiomyocyte upon Trypanosoma cruzi infection. Numbers of up- (left column) and down-regulated (right column) DEGs at 24- (upper row) and 48-h post-infection (lower row) are shown in the respective Venn diagrams. hpi hours post-infection

Fig. 5. Principal component analysis plot of the transcriptional profile of the in vitro cardiomyocyte infection models. Principal component analysis plot, based on the fold-change (vs control) values of the 10,784 valid genes. Each dot represents a biological condition at 24- or 48-h post-infection. The principal components 1 and 2 together describe 58.9% of the total variance in the dataset. Dim dimension

Fig. 6. Top differentially expressed genes of the cardiomyocyte upon Trypanosoma cruzi infection. Extensively A upregulated (fold-change > 5) and B downregulated (fold-change < 0.2) genes of the cardiomyocyte at 24-h post-infection are listed. Δ Gene expression, the fold-change or ratio between gene expression level of the infected cardiomyocytes and the non-infected negative control. Arrdc4 arrestin domain containing 4, Txnip thioredoxin interacting protein, Gdf15 growth differentiation factor 15, Atf3 activating transcription factor 3, Soat2 sterol o-acyltransferase 2, Dusp8 dual specificity phosphatase 8, Chac1 cation transport regulator 1, Gtpbp2 GTP binding protein 2, Capn5 Calpain 5, Dhcr24 24-dehydrocholesterol reductase, Lgals9 lectin galactose binding soluble 9, Acat2 acetyl-coenzyme A acetyltransferase 2, Hmgcs1 3-hydroxy-3-methylglutaryl-Coenzyme A synthase 1, Mvd mevalonate (diphospho) decarboxylase, Tpcn1 two pore channel 1, Medag mesenteric estrogen dependent adipogenesis, Sulf2 sulfatase 2, Fam64a family with sequence similarity 64 member A, Midn midnolin, Ldlr low density lipoprotein receptor, Acat3 acetyl-coenzyme A acetyltransferase 3, Triobp TRIO and F-actin binding…

Fig. 7. Signatures of the common cardiomyocyte response to Trypanosoma cruzi infection. A, B ‘WikiPathways’ and C, D ‘Biological Process gene ontology Terms’ enriched by commonly A, C upregulated and B, D downregulated genes of the host cardiomyocyte at 24-h post-infection are sorted based on P-value. Bar lengths represent enrichment Z-scores

Fig. 8. Heatmap of expression levels of inflammation-related genes in HL-1 cells infected with T. cruzi at 24-h post-infection. Genes are represented in rows and infection conditions are represented in columns. Color legend represents the logarithm 2 of gene expression level. P-values were calculated by paired T-test. Gdf-15 growth differentiation factor 15, Cxcl10 C-X-C motif chemokine ligand 10, Cxcl2 C-X-C motif chemokine ligand 2, Il-6 interleukin-6, Tnf tumor necrosis factor-alpha

Fig. 9. Signatures of transcriptomic remodeling unique to Colombian- and Y-infections. ‘WikiPathways’ (A) and 'Biological Process gene ontology Terms’ (B), enriched by the genes uniquely upregulated at 24-h post-infection in Colombian- and Y-infected cardiomyocytes are sorted based on P-value. Bar lengths represent enrichment Z-scores

Fig. 10. Elevated intracellular levels of reactive oxygen species in Trypanosoma cruzi-infected cardiomyocytes. A Signal intensity of reactive oxygen species-specific fluorescence was higher in the cardiomyocytes infected with the Y strain at 24-h post-infection, compared with non-infected control cells. Statistical comparison was performed with Mann–Whitney test. Bars represent mean fluorescence intensity and error bars represent standard deviation. B Representative fluorescence microscopy images from reactive oxygen species-specific CellRox green staining. Comparison with the non-infected (upper) control cells shows higher intensity of reactive oxygen species fluorescence signal in the cardiomyocytes 24-h post-infection with Trypanosoma cruzi Y strain (lower). Scale bars represent 100 µm