Fasciola hepatica Infection in Cattle: Analyzing Responses of Peripheral Blood Mononuclear Cells (PBMC) Using a Transcriptomics Approach - PubMed (original) (raw)
Fasciola hepatica Infection in Cattle: Analyzing Responses of Peripheral Blood Mononuclear Cells (PBMC) Using a Transcriptomics Approach
Andres Garcia-Campos et al. Front Immunol. 2019.
Abstract
The parasitic helminth Fasciola hepatica (liver fluke) causes economic loss to the livestock industry globally and also causes zoonotic disease. New control strategies such as vaccines are urgently needed, due to the rise of drug resistance in parasite populations. Vaccine development requires a comprehensive understanding of the immunological events during infection. Previous in vivo studies by our group have investigated global differentially expressed genes (DEGs) in ovine peripheral blood mononuclear cells (PBMC) in response to both acute and chronic F. hepatica infection. This work demonstrated that pathways involved in the pathogenesis of ovine fasciolosis included fibrosis, inhibition of macrophage nitric oxide production, and antibody isotype switching, among others. Transcriptomic changes in PBMC populations following F. hepatica infection in cattle, in which the disease phenotype is quite different, have not yet been examined. Using RNA sequencing we investigated gene expression changes in PBMC isolated from 9 non-infected and 11 _F. hepatica_-experimentally-infected calves immediately before infection, at 1 and at 14 weeks post-infection. Longitudinal time-course comparisons between groups revealed 21 and 1,624 DEGs driven exclusively by F. hepatica infection in cattle at acute and chronic stages, respectively. These results show that fewer DEGs at the acute stage of infection can be identified in cattle, as compared with sheep. In addition, the log2 fold-changes of these DEGs were relatively low (-1 to 3) reflecting the different clinical presentation of F. hepatica infection in cattle. Gene pathways for hepatic fibrosis and hepatic cholestasis along with apoptosis of antigen-presenting cells were enriched at chronic stages. Our results reflect the major differences in the disease phenotype between cattle and sheep and may indicate pathways to target in vaccine development.
Keywords: Fasciola; PBMCs; apoptosis; cattle; immunoregulation; transcriptomics; vaccines.
Figures
Figure 1
Summary of the comparisons used for the differential expression analysis. Non-paired comparisons between uninfected and infected groups at week 0 (W0), 1 week (W1), and 14 weeks (W14) post-experimental infection (A). Longitudinal time-course analysis of DEGs for each group between week 1 and week 0 (W1 vs. W0), week 14 and week 1 (W14 vs. W1), week 14 and week 0 (W14 vs. W0) (B). DEGs obtained exclusively in the infected group were employed for IPA downstream analysis.
Figure 2
Evaluation of anti Fasciola hepatica Cathepsin L1 IgG1 antibodies in uninfected (shapes in gray) and infected (shapes in pink) animals by ELISA at week 0 (W0), 1 week (W1), and 14 weeks (W14) post-experimental infection. Each dot and star represents optical density (O.D.) values at 450 nm for each Holstein-Friesian crossbred and Holstein-Friesian individual, respectively. Horizontal lines represent the mean for each group and time point (A). Fluke burden found in livers and bile ducts of uninfected and infected animals 15 weeks post-infection. Each star represents numbers of F. hepatica flukes for each Holstein-Friesian animal. Each dot represents numbers of F. hepatica flukes for each Holstein-Friesian crossbred animal. Horizontal lines represent the mean for each group (B). Dynamics of the different PBMC sub-populations (lymphocytes, monocytes, and eosinophils) in uninfected (gray) and infected (pink) animals at W0, W1, and W14. Each dot and star represent absolute values that are expressed in cells × 109/L for each Holstein-Friesian crossbred and Holstein-Friesian individual, respectively. Horizontal line represents the mean for each group and time point. Dots and stars located above the horizontal line indicate that values are above the standard reference range (C). *P = 0.08 **P = 0.009.
Figure 3
Multidimensional scaling of uninfected and infected Holstein-Friesian and Holstein-Friesian crossbred animals at week 0 (W0), week 1 (W1), and week 14 (W14) post F. hepatica infection. The BCV distance 1 (x-axis) and 2 (y-axis) represent the directions that separate the gene expression of the samples to the greatest extent.
Figure 4
Volcano plots showing the extent of log2 fold-change (x-axis) and their −log10 FDR values (y-axis) for the DEGs identified in PBMC and their direction of expression driven specifically by F. hepatica infection (dark blue). Genes that did not pass the FDR threshold and those DEGs that were also identified in the uninfected group are represented in light blue. Comparison of DEGs at W1 vs. W0 (A), W14 vs. W0 (B), and W14 vs. W1 (C).
Figure 5
Venn diagrams showing the numbers of DEGs identified in PBMC and their direction of expression driven specifically by F. hepatica infection. Comparison of DEGs from W1 vs. W0 with W14 vs. W0 (A) and W1 vs. W0 with W14 vs. W1 (B). Downregulated genes are shown in gray and pink, upregulated genes in green and blue.
Figure 6
Interaction of the upstream regulators for the activation of apoptosis of antigen presenting cells (A), migration of mononuclear leukocytes (B), recruitment of myeloid cells (C), recruitment of phagocytes (D), chemotaxis (E), and migration of phagocytes (F) in PBMC of _F. hepatica_-infected animals at the W14 vs. W1 timepoint. STAT3 and APP are indicated as activated (orange), whereas mir-155 and DUSP1 are inhibited (blue). Downstream target genes are highlighted as upregulated (red) or downregulated (green) with color intensity increasing with degree of log2 fold-change. Arrowheads at the end of interactions (dotted lines) indicate activation, while bars indicate inhibitory effects. The color of lines represents predicted relationships based on gene expression, including orange (activation), blue (inhibition), yellow (findings inconsistent with state of downstream molecule), and gray (effect not predicted). © 2000–2019 QIAGEN. All rights reserved.
Figure 7
Prediction of TNF and IL1B as the upstream regulator cascade involved in hepatic fibrosis in animals experimentally infected with F. hepatica at 14 weeks post-infection. TNF is indicated as activated (orange square). Downstream target genes are highlighted as upregulated (red) or downregulated (green) with color intensity increasing with degree of log2 fold-change. Genes are illustrated according to the subcellular location of proteins translated from respective genes. Downstream target genes that do not have an effect in liver fibrosis are shaded in the background. Arrowheads at the end of interactions (dotted lines) indicate activation, while bars indicate inhibitory effects. The colors of lines represent predicted relationships based on gene expression, including orange (activation), blue (inhibition), yellow (findings inconsistent with state of downstream molecule), and gray (effect not predicted). © 2000–2019 QIAGEN. All rights reserved.
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