Immunopathology in RSV Infection Is Mediated by a Discrete Oligoclonal Subset of Antigen-Specific CD4+ T Cells  Steven M Varga, Xiaoting Wang, Raymond M Welsh, Thomas J Braciale  Immunity  Volume 15, Issue 4, Pages 637-646 (October 2001) DOI: 10.1016/S1074-7613(01)00209-6

Figure 1 Intracellular IFN-γ Expression of RSV G Peptide-Specific Vβ14+ CD4+ T Cells Lung mononuclear cells from a pool of four vvβgal or four vvG-primed mice 5 days after RSV infection were stimulated with RSV G 183-195 peptide or left unstimulated in the presence of brefeldin A for 5 hr. Representative Vβ14/IFN-γ costaining for the CD4-gated populations are shown. Percentages in the indicated quadrants represent the percentage of gated CD4+ cells that fall within each quadrant. Data shown are representative of three separate experiments with a pool of four mice per experiment Immunity 2001 15, 637-646DOI: (10.1016/S1074-7613(01)00209-6)

Figure 2 Intracellular IFN-γ and TNF-α Expression of Lung-Derived Memory Effector CD4+ T Cells Lung mononuclear cells from a pool of four vvG-primed mice 5 days after RSV infection were stimulated with RSV G 183-195 peptide in the presence of brefeldin A for 5 hr. Cells were stained for expression of CD4, Vβ14, IFN-γ, and TNF-α. Representative CD4/Vβ14 costaining is shown. The right panels represent IFN-γ and TNF-α costaining on the two gated populations. Percentages in the indicated quadrants represent the percentage of the gated cells that fall within each quadrant. Data shown are representative of three separate experiments with a pool of four mice per experiment Immunity 2001 15, 637-646DOI: (10.1016/S1074-7613(01)00209-6)

Figure 3 Th1 and Th2 Cytokine Production by Vβ14+ CD4+ T Cells Lung mononuclear cells were isolated from four individual vvG-primed mice 5 days after RSV infection and stimulated in vitro with platebound anti-Vβ14 or anti-TCRβ mAb for 48 hr. The cytokine concentration in the supernatant was determined for the given cytokines by ELISA. One representative from three separate experiments is shown Immunity 2001 15, 637-646DOI: (10.1016/S1074-7613(01)00209-6)

Figure 4 CDR3-Length Spectratypes of Vβ14+ Cells and Dominant Usage of T Cell Repertoire in BALB/c Mice (A) CDR3 profiles of the Vβ14-Jβ runoff products from a G-primed, RSV-infected mouse. Total RNAs were extracted from lung lymphocytes 5 days after infection and subjected to spectratyping analysis. Fluorescent intensity (y axis) was plotted against relative CDR3 size (x axis). (B) Similar Vβ14-Jβ1.1 TCR sequences after infection. PCR products from eight individual mice were sequenced. Amino acid sequences of the Vβ14-Jβ1.1 were deduced from DNA sequences. In mouse #6, the DNA sequence data showed that either of two amino acids were encoded by what were probably at least two clones, and they are designated in the A/B format Immunity 2001 15, 637-646DOI: (10.1016/S1074-7613(01)00209-6)

Figure 5 Vβ14 Depletion at the Time of RSV Challenge Infection Prevents RSV Vaccine-Enhanced Disease Mice were primed with vvG and challenged with RSV 21 days later. One group of four mice received anti-Vβ14 mAb at days −1 and +2 relative to RSV challenge, whereas a second group received purified rat IgM mAb as a control. Representative data from one of three separate experiments with four individual mice per experiment are shown. (A) Mean illness scores ± SD from four individual mice per group. (B) Weight loss was monitored daily following RSV challenge. The percent weight loss was calculated on subsequent days after RSV challenge on the basis of the original weight of each mouse. (C) Mean BAL eosinophil percentages ± SD from four individual mice per group Immunity 2001 15, 637-646DOI: (10.1016/S1074-7613(01)00209-6)

Figure 6 Intracellular IFN-γ Expression of RSV G Peptide-Specific Vβ14+ CD4+ T Cells in Rat IgM or Vβ14-Treated Mice Lung mononuclear cells isolated 5 days after RSV infection from four individual G-primed mice that had received either anti-Vβ14 or rat IgM as a control. (A) Representative Vβ14/IFN-γ costaining on the CD4-gated population following stimulation with the RSV G 183-195 peptide in the presence of brefeldin A for 5 hr. (B) Representative IFN-γ staining for RSV G 183-195 epitope-specific CD4+ T cells. (C) Representative IFN-γ staining following PMA and ionomycin activation of the CD4+ T cells. Contour plot analyses are shown using 5% probability curves. Data shown are representative of three separate experiments with four mice/group per experiment Immunity 2001 15, 637-646DOI: (10.1016/S1074-7613(01)00209-6)

Figure 7 Vβ14 Depletion during G Priming Prevents the Subsequent Development of RSV Vaccine-Enhanced Disease Mice were primed with vvG and challenged with RSV 21 days later. One group of four mice received anti-Vβ14 mAb at days −1, +4, and +8 relative to scarification with vvG, whereas a second group received purified rat IgM mAb as a control. Representative data from one of three separate experiments with four individual mice per experiment are shown. (A) Mean illness scores ± SD from four individual mice per group. (B) Weight loss was monitored daily following RSV challenge. The percent weight loss was calculated on subsequent days after RSV challenge on the basis of the original weight of each mouse. (C) Mean BAL eosinophil percentages ± SD from four individual mice per group Immunity 2001 15, 637-646DOI: (10.1016/S1074-7613(01)00209-6)