Cytokine and chemokine kinetics in natural human dengue infection as predictors of disease outcome

Abstract

Dengue is an important tropical disease with considerable global impact. Despite this, there remains an urgent need for reliable biomarkers to predict disease severity, as well as effective antiviral drugs and targeted treatments. In this study, we conducted a comprehensive profiling of 41 plasma mediators in patients with asymptomatic dengue (AD) and symptomatic dengue (SD), which includes mild dengue fever (DF) and severe dengue hemorrhagic fever (DHF). Our findings revealed that the levels of nearly all measured mediators were consistently lower in AD compared to SD patients, suggesting a potential protective cytokine response signature. Time-course cytokine analysis in SD shown significantly elevated levels of pro-inflammatory cytokines and chemokines associated with inflammation and viral clearance upon the acute phase, while various growth factors were elevated during the convalescence. Notably, we identified elevated IL-15 levels in DHF patients three days before fever subsidence, highlighting its potential as an early prognostic biomarker for severe disease outcomes. Furthermore, prolonged high levels of IL-8 and IP-10 in DHF during the critical period may contribute to dengue immunopathogenesis. This study advances the understanding of cytokine dynamics in the natural course of human dengue infection, providing valuable insights for the development of targeted treatments and prognostic biomarkers.

Fig. 1

Asymptomatic infection has lower cytokine secretion than symptomatic infection. (a) Comparable viremia level between AD (n = 5), DF (n = 32) and DHF (n = 10) sample. (b) PCA and (c) PCA biplot of the total 41 humoral mediators clustered by severity (Total n = 47). Each circle corresponds to one patient. (d) Levels of pro-inflammatory cytokines: TNF-α, IL-1α, IL-15, MIP-1α, and MIP-1β. (e) Level of antiviral cytokines: IFN-α2 and IFN-γ. (f) Levels of anti-inflammatory cytokines: IL-10 and IL-1RA. (g) Levels of chemokines: IL-8, IP-10, and Eotaxin. Each dot represents the level of cytokine from each sample. Kruskal–Wallis test was used to compare 3 groups of independent data (p value indicated in each graph). Kruskal–Wallis statistics with Dunn’s multiple comparisons test were used to compare between 2 groups indicated. *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001. Median and interquartile ranges were shown in each group.

Fig. 2

Overall expression of the 41 cytokines over the time course in symptomatic DENV infection. (a) Diagram shows the 4 time points analyzed during the course of DENV infection. (b) Principal component analysis (PCA) plot and (c) PCA Biplot of total 55 dependent samples clustered by timepoints. (d) PCA plot clustered by severities; DF and DHF. (DF n = 15, DHF n = 41).

Fig. 3

Cytokines associated with acute dengue infection. The level of (a) proinflammatory cytokines, (b) Anti-inflammatory cytokines, (c) Chemoattractant cytokines and (d) Antiviral cytokines. Each dot represents the level of cytokine from each sample. The 3 left columns include all samples, Kruskal–Wallis test was used to compare 4 groups of independent data (p value indicated in each graph). Kruskal–Wallis statistics with Dunn’s multiple comparisons test were used to compare between 2 groups indicated. *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001. Median and interquartile ranges were shown in each group. Left most column: combined DF and DHF, second column: DHF only, third column: DF only. The 3 right columns include only dependent data from the same patients (each line connects between paired samples). Wilcoxon matched-pairs signed rank test was used to compare between 2 groups indicated. *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001. The 4th column: combined DF and DHF, 5th column: DHF only, 6th column: DF only.

Fig. 3

Cytokines associated with acute dengue infection. The level of (a) proinflammatory cytokines, (b) Anti-inflammatory cytokines, (c) Chemoattractant cytokines and (d) Antiviral cytokines. Each dot represents the level of cytokine from each sample. The 3 left columns include all samples, Kruskal–Wallis test was used to compare 4 groups of independent data (p value indicated in each graph). Kruskal–Wallis statistics with Dunn’s multiple comparisons test were used to compare between 2 groups indicated. *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001. Median and interquartile ranges were shown in each group. Left most column: combined DF and DHF, second column: DHF only, third column: DF only. The 3 right columns include only dependent data from the same patients (each line connects between paired samples). Wilcoxon matched-pairs signed rank test was used to compare between 2 groups indicated. *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001. The 4th column: combined DF and DHF, 5th column: DHF only, 6th column: DF only.

Fig. 4

Cytokines associated with convalescent dengue infection. Each dot represents the level of cytokine from each sample. The 3 left columns include all samples, Kruskal–Wallis test was used to compare 4 groups of independent data (p value indicated in each graph). median with interquartile range were shown in each group. Kruskal–Wallis statistics with Dunn’s multiple comparisons test were used to compare between 2 groups indicated. *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001. Left most column: combined DF and DHF, second column: DHF only, third column: DF only. The 3 right columns include only dependent data from the same patients (line connect between paired samples). Wilcoxon matched-pairs signed rank test was used to compare between 2 groups indicated. *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001. The 4th column: combined DF and DHF, 5th column: DHF only, 6th column: DF only.

Fig. 5

Cytokines are associated with dengue severity. (a) Level of IL-15 at D-3 time-point. (b) Levels of IL-8 and IP-10 at D0 time-point. Each dot represents the level of cytokine from each sample. The Mann Whitney test was used to compare the difference between DF and DHF samples (p value indicated in each graph). Median and interquartile ranges are shown in each plot. (c) ROC curve of IL-15 level between DF and DHF samples at D-3 time-point. AUC; area under the curve, *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001.