Table 1.
PLP:139-151-induced co-expression of IL-2, IL-3, IL-4, and IFN-γ in freshly isolated memory CD4 T cells. SJL mice were immunized with PLP:139-151 peptide and at the time points indicated, CD4 cells were isolated from the dLN or the spleen. CD4 cells were tested in serial dilutions starting at 3 × 105 cells per well with 5 × 105 APC (irradiated naive SJL spleen cells) for the recall response to PLP:139-151 (20 μM) performing the specified double color cytokine ELISA spot assays. Double color ELISPOT assays were of 48 h duration. Data for individual mice representative of groups of 4–24 mice tested per time point are shown. The numbers of single-positive and double-positive (DP) cells were measured by image analysis The % DP was calculated as the percentage of double-cytokine-producing cells from the total number of cells producing both cytokines: % DP = DP × 100%/(DP + Cytokine 1 only + Cytokine 2 only). The mean spot numbers of 3–4 replicate wells are shown for 3 × 105 cells per well with SD < 10%. The standard deviation for all % DP calculated was less than % DP ± 1%.
| Time | Disease | Organ | IFN-γ/IL-2 assay | IFN-γ /IL-3 assay | IFN-γ/IL-4 assay | |||||||||
|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
| (days) | state | IFN-γ | IL-2 | DP | %DP | IFN-γ | IL-3 | DP | %DP | IFN-γ | IL-4 | DP | %DP | |
| 2 | No disease | LN | 4 | 29 | 3 | 8.3 | 5 | 6 | 1 | 8.3 | 5 | 9 | 0 | 0.0 |
| Spleen | 2 | 6 | 0 | 0.0 | 4 | 2 | 0 | 0.0 | 3 | 47 ** | 0 | 0.0 | ||
| CNS * | 0 | 0 | 0 | 0.0 | 0 | 0 | 0 | 0.0 | 0 | 0 | 0 | 0.0 | ||
| 3 | No disease | LN | 41 | 104 | 12 | 7.6 | 59 | 62 | 8 | 6.2 | 51 | 14 | 0 | 0.0 |
| Spleen | 6 | 23 | 1 | 3.3 | 4 | 7 | 0 | 0.0 | 9 | 127 ** | 0 | 0.0 | ||
| CNS * | 0 | 0 | 0 | 0.0 | 0 | 0 | 0 | 0.0 | 0 | 0 | 0 | 0.0 | ||
| 7 | No disease | LN | 224 | 213 | 21 | 4.6 | 232 | 62 | 8 | 2.6 | 240 | 9 | 1 | 0.0 |
| Spleen | 356 | 217 | 23 | 3.9 | 335 | 86 | 9 | 2.1 | 349 | 218 ** | 7 | 0.0 | ||
| CNS * | 1 | 2 | 0 | 0.0 | 3 | 0 | 0 | 0.0 | 1 | 0 | 0 | 0.0 | ||
| 12 | First paralysis | LN | 46 | 183 | 7 | 3.0 | 43 | 36 | 5 | 5.9 | 52 | 7 | 0 | 0.0 |
| Spleen | 408 | 210 | 22 | 3.4 | 397 | 114 | 13 | 2.5 | 429 | 236 ** | 4 | 0.0 | ||
| CNS* | 932 | 374 | 49 | 3.6 | 971 | 372 | 47 | 3.4 | 938 | 8 | 1 | 0.0 | ||
| 21 | Second paralysis | LN | 45 | 139 | 4 | 2.1 | 52 | 53 | 6 | 5.4 | 47 | 6 | 0 | 0.0 |
| Spleen | 116 | 202 | 7 | 2.2 | 115 | 71 | 9 | 4.6 | 121 | 223 ** | 2 | 0.0 | ||
| CNS * | 307 | 108 | 11 | 2.6 | 288 | 103 | 10 | 2.5 | 296 | 14 | 1 | 0.0 | ||
| 55 | After three relapses | LN | 47 | 76 | 5 | 3.9 | 49 | 33 | 2 | 2.4 | 41 | 3 | 0 | 0.0 |
| Spleen | 140 | 201 | 12 | 3.4 | 151 | 74 | 11 | 4.7 | 148 | 192 ** | 2 | 0.1 | ||
| CNS * | 119 | 62 | 8 | 4.2 | 107 | 42 | 5 | 3.2 | 111 | 4 | 0 | 0.0 | ||
* Spinal cord mononuclear cells (frequencies shown per 3 × 105) were tested in the presence of 5 × 105 exogenous splenic APC;
** Bystander IL-4 production by non-T/non-B splenic cells activated in the presence of antigen-stimulated CD4 T cells [37].