Genome-wide expression profiling deciphers host responses altered during dengue shock syndrome and reveals the role of innate immunity in severe dengue

Abstract

Background: Deciphering host responses contributing to dengue shock syndrome (DSS), the life-threatening form of acute viral dengue infections, is required to improve both the differential prognosis and the treatments provided to DSS patients, a challenge for clinicians.

Methodology/principal findings: Based on a prospective study, we analyzed the genome-wide expression profiles of whole blood cells from 48 matched Cambodian children: 19 progressed to DSS while 16 and 13 presented respectively classical dengue fever (DF) or dengue hemorrhagic fever grades I/II (DHF). Using multi-way analysis of variance (ANOVA) and adjustment of p-values to control the False Discovery Rate (FDR<10%), we identified a signature of 2959 genes differentiating DSS patients from both DF and DHF, and showed a strong association of this DSS-gene signature with the dengue disease phenotype. Using a combined approach to analyse the molecular patterns associated with the DSS-gene signature, we provide an integrative overview of the transcriptional responses altered in DSS children. In particular, we show that the transcriptome of DSS children blood cells is characterized by a decreased abundance of transcripts related to T and NK lymphocyte responses and by an increased abundance of anti-inflammatory and repair/remodeling transcripts. We also show that unexpected pro-inflammatory gene patterns at the interface between innate immunity, inflammation and host lipid metabolism, known to play pathogenic roles in acute and chronic inflammatory diseases associated with systemic vascular dysfunction, are transcriptionnally active in the blood cells of DSS children.

Conclusions/significance: We provide a global while non exhaustive overview of the molecular mechanisms altered in of DSS children and suggest how they may interact to lead to final vascular homeostasis breakdown. We suggest that some mechanisms identified should be considered putative therapeutic targets or biomarkers of progression to DSS.

Figure 1. Unsupervised hierarchical clustering of whole blood cells expression profiles from the 48 dengue-infected children.

The clustering is based on the 2959 gene list (3515 clones, detailed in Table S2) discriminating dengue fever (DF), dengue hemorrhagic fever (DHF) and dengue shock syndrome (DSS) patients. Each row represents a single transcript and each column represents a patient's sample. Color scale indicates the range of gene expression: black indicates median expression level, red greater expression, green lower expression. The 2 patient subsets identified are indicated. PLxxx, code relative to one patient. Black star: DSS patient sampled 3 days after shock. Orange star: Patients who received perfusion of human plasma before collection of blood samples.

Figure 2. Top 30 canonical pathways identified from the DSS-gene signature using Ingenuity Pathway Analysis software.

The significance of the association between data set and canonical pathway was estimated by the p-value (Fischer's exact test; left axis) and the ratio (right axis) of genes that maps to each canonical pathway.

Figure 3. T Cell Receptor Signaling canonical pathway from Ingenuity Pathway Analysis.

Genes in green and red are respectively under- and over-expressed in the DSS-gene signature. Genes in white are other genes present in the canonical pathway but absent from the DSS-gene signature. DSS: Dengue Shock Syndrome.

Figure 4. Hypothesis of a second inflammatory amplification loop in dengue shock syndrome.

After induction of a first inflammatory and anti-viral response to dengue virus, disease resolution generally occurs around time of defervescence for most dengue-infected patients. Some patients however progress towards a life-threatening dengue shock syndrome. Results obtained in this study suggest that in those patients, a second inflammatory amplification loop, which involves a diversity of pro-inflammatory responses related to innate immunity, occurs and leads to a major inflammatory systemic syndrome and to vascular homeostasis breakdown. The putative role of different markers identified in vascular endothelial dysfunction is indicated. Thin black arrow, release of; Bold black arrow, interaction between; Punctuated black arrow, chemotactic effect; Thin red arrow, biological activity; Bold red arrow, direct activity on endothelium. DAMPs, danger-associated molecular pattern; GAG, glycosaminoglycane; ROI, reactive oxygen intermediates; TLR, Toll-like receptor.