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. 2002 Sep 16;196(6):731–741. doi: 10.1084/jem.20020223

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Copyright © 2002, The Rockefeller University Press

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Figure 4. — Minigene-induced T cells inhibit IgG anti-DNA Ab production in vitro and in vivo. 3-mo-old BWF1 mice were vaccinated with three weekly intradermal injections (three short arrows in panel B) of pCD.III (▴), pCD (▪), or PBS (⋄) (n = 15 per group). (A) Six mice in each group were killed 10 d after the last injection, and their splenic T cells were cocultured with B cells from 9-mo-old untreated BWF1 mice for 5 d in the absence or presence of three peptides. IgG anti-DNA Abs in culture supernatants are shown as mean ± SD U/ml. *P < 0.05, pCD.III versus control (pCD or PBS) groups (Student's t test). (B) The remaining nine mice were challenged once subcutaneously (a long arrow) with 25 μg each of peptide VH1, VH2, and VH3 together in a 1:1 emulsion with CFA 2 wk after the last injection. Their monthly sera were tested for anti-dsDNA Abs (left panel) or total IgG (right panel). Results are expressed as the mean ± SD. *P = 0.01 to 0.003, pCD.III versus both control groups; **P = 0.04, pCD.III versus pCD group (Student's t test). Results are from one of two similar experiments. (C) 10-wk-old BWF1 mice were inoculated thrice on alternate days with pCD.III or pCD (n = 8 per group). A week after the last injection, spleens were harvested and single cell suspensions were cultured with IL-2 and 10 μg/ml each of the three MHC class I–binding epitopes shown in Fig. 1C (cells from pCD.III-injected mice) or with IL-2 alone (cells from pCD-injected mice). After 3 to 7 d in culture, cells were washed, purified as live cells using Ficoll-Hypaque, and then enriched for T cells using MACS. The purified T cells (5 × 10⁶ or 5 × 10³) were injected intravenously twice at 3-d intervals into 22-wk-old BWF1 mice (n = 6 mice per group). 5 d after the second transfer, recipient animals were killed and their spleen cells were tested for IgG anti-DNA Ab forming cells in an ELISPOT assay. Results are shown as the mean ± SD IgG anti-DNA Ab forming cells per 10⁶ spleen cells. *P = 0.02, recipients of T cells from pCD.III-vaccinated mice (▴) versus recipients of T cells from pCD-injected mice (▪).

Figure 4. — Minigene-induced T cells inhibit IgG anti-DNA Ab production in vitro and in vivo. 3-mo-old BWF1 mice were vaccinated with three weekly intradermal injections (three short arrows in panel B) of pCD.III (▴), pCD (▪), or PBS (⋄) (n = 15 per group). (A) Six mice in each group were killed 10 d after the last injection, and their splenic T cells were cocultured with B cells from 9-mo-old untreated BWF1 mice for 5 d in the absence or presence of three peptides. IgG anti-DNA Abs in culture supernatants are shown as mean ± SD U/ml. *P < 0.05, pCD.III versus control (pCD or PBS) groups (Student's t test). (B) The remaining nine mice were challenged once subcutaneously (a long arrow) with 25 μg each of peptide VH1, VH2, and VH3 together in a 1:1 emulsion with CFA 2 wk after the last injection. Their monthly sera were tested for anti-dsDNA Abs (left panel) or total IgG (right panel). Results are expressed as the mean ± SD. *P = 0.01 to 0.003, pCD.III versus both control groups; **P = 0.04, pCD.III versus pCD group (Student's t test). Results are from one of two similar experiments. (C) 10-wk-old BWF1 mice were inoculated thrice on alternate days with pCD.III or pCD (n = 8 per group). A week after the last injection, spleens were harvested and single cell suspensions were cultured with IL-2 and 10 μg/ml each of the three MHC class I–binding epitopes shown in Fig. 1C (cells from pCD.III-injected mice) or with IL-2 alone (cells from pCD-injected mice). After 3 to 7 d in culture, cells were washed, purified as live cells using Ficoll-Hypaque, and then enriched for T cells using MACS. The purified T cells (5 × 10⁶ or 5 × 10³) were injected intravenously twice at 3-d intervals into 22-wk-old BWF1 mice (n = 6 mice per group). 5 d after the second transfer, recipient animals were killed and their spleen cells were tested for IgG anti-DNA Ab forming cells in an ELISPOT assay. Results are shown as the mean ± SD IgG anti-DNA Ab forming cells per 10⁶ spleen cells. *P = 0.02, recipients of T cells from pCD.III-vaccinated mice (▴) versus recipients of T cells from pCD-injected mice (▪).