Acute rotavirus infection is associated with the induction of circulating memory CD4+ T cell subsets

Figures

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  Figure 1. RV-IgA responses in children with rotavirus-positive diarrhoea. RV-IgA was measured from the plasma of the children with laboratory-confirmed rotavirus diarrhoea using a standardized quantitative ELISA. (A) RV-IgA geometric mean concentration (U/mL) for children with rotavirus-positive diarrhoea at acute and convalescent phases of infection. (B) Paired sample analysis for children with rotavirus-positive diarrhoea at acute and convalescent phases of infection. CI confidence interval, GMC geometric mean concentration.

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  Figure 2. CD4+ T subsets in children with or without rotavirus diarrhoea. The cells were stained with the following fluorochrome-conjugated antibodies CD3 PerCP-CY5.5, CD4 APC-CY7, CCR7 AF700, CD45RA FITC, CRTH2 APC, CXCR3 Qdot 605, PD1 PE CY7 and BCL6 PE. Central memory T cells were defined as CD4+, CCR7+ and CD45RA-, effector memory T cells as CD4+, CD45RA− and CCR7−, terminally differentiated as CD4+, CD45RA+ and CCR7− and naïve T cells as CD4+, CCR7+ and CD45RA+. Th1 was defined as CD4+ and CXCR3+, Th2 as CD4+ and CRTH2+ and Tfh as CD4+ BCL6+ and PD1hi. Central memory Th1 was defined as CD4+, CXCR3+, CD45RA− and CCR7+, effector memory Th1 as CD4+, CXCR3+, CD45RA− and CCR7−, Central memory Th2 was defined as CD4+, CRTH2+, CD45RA− and CCR7+ and effector memory Th1 as CD4+, CRTH2+ CD45RA− and CCR7−. Mann–Whitney U test was used to compare the proportion of the T cells between children with rotavirus-positive and rotavirus-negative at both acute and convalescent phases of infection. Differences after comparisons were considered statistically significant if p-values were less than 0.05. (A) Representative flow cytometry plot from a child with rotavirus diarrhoea peripheral whole blood…

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  Figure 3. CD4+ T stimulations in children with or without rotavirus diarrhoea. The cells were stained with the following fluorochrome-conjugated antibodies CD3 PerCP-CY5.5, CD4 BV 421, CD8 APC-CY7, TNF-α FITC, IFN-γ PE-CF594 and CD69 AF700. Mann–Whitney U test was used to compare the proportion of the T cells between children with rotavirus-positive and rotavirus-negative at both acute and convalescent phases of infection. Differences after comparisons were considered statistically significant if p-values were less than 0.05. (A) Representative FACS plot from children with rotavirus diarrhoea peripheral whole blood showing CD4 + T cells producing TNF-α and IFN-γ cells from PMA + ION (Mitogen) and VP6 protein responses and the unstimulated control. (B) Proportion of CD4 + producing TNF-α or IFN-y and both TNF-α and IFN-γ cells for children with rotavirus rotavirus-positive and rotavirus-negative diarrhoea at the acute phase of infection. (C) Proportion of CD4 + producing TNF-α or IFN-y and both TNF-α and IFN-γ cells for children with rotavirus-positive and rotavirus-negative diarrhoea at the convalescent phase of infection.

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  Figure 4. CD4+ T cells mitogenic cytokine profiles in children with or without rotavirus diarrhoea. The cells were stimulated with either PMA + ION or VP6 protein for 18 hours, with the unstimulated control in all experiments. The cells were stained with the following fluorochrome-conjugated antibodies CD3 PerCP-CY5.5, CD4 BV421, CD8 APC-CY7, TNF-α FITC, IFN-γ PE-CF594 and CD69 AF700. Single producers were either CD4 + T cells producing TNF-α or CD4 + T cells producing IFN-γ and double producers were CD4 + T cells producing TNF-α and IFN-γ. Mann–Whitney U test was used to compare the proportion of the T cells between children with rotavirus-positive and rotavirus-negative at both acute and convalescent phases of infection. Differences after comparisons were considered statistically significant if p-values were less than 0.05. (A) Frequency of CD4 + T cells producing TNF-α and IFN-γ, activated (CD69+) or not activated (CD69−) in children with rotavirus-positive and rotavirus-negative diarrhoea at the acute phase of infection. (B) Frequency of CD4 + T cells producing TNF-α and IFN-γ, activated (CD69+) or not activated (CD69−) in children with rotavirus-positive and rotavirus-negative…