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Article Figures & Data

Figures

  • Figure 1.

    CD44hiCD8+T lymphocytes preferentially expand in DNRII Tg mice. (A) Lymphocytes from 6-week-old DNRII Tg mice or nontransgenic littermates (C57BL/6) were analyzed for CD44hi cell-surface expression (MFI, greater than 100) on CD4+ or CD8+ T-cell subsets. Data are represented as the average percentage of CD44hi cells in each T-cell subset, with error bars representing the SD in each group. (B) Lymphocytes from the same 6-week-old DNRII Tg mice or nontransgenic littermates (C57BL/6) were analyzed for the percentages of CD4+ or CD8+ T-cell subsets. Data are represented as the average percentage of each T-cell subset among the total T-cell population, with error bars representing the SD in each group. (C) Lymphocytes from 6-week-old DNRII Tg mice were gated into CD8+ T-cell subsets (left) or CD4+ T-cell subsets (right). Data in the top 2 panels represent the percentage of each T-cell subset with respect to total T cells in the sample. The bottom 6 panels represent flow cytometric analysis of surface receptor expression in each of the 2 gated T-cell subsets (filled histograms). Data represent the percentages of indicated surface receptors within each gated T-cell subset and are typical of these strains of mice at 6 weeks of age. Open histograms indicate nonspecific control staining. (A-B) Average of 5 or more mice per group; experiments repeated 5 times.

  • Figure 2.

    Antigenic requirements for CD8+ T-cell expansion in DNRII Tg mice. (A) Lymphocytes from 8- to 10-week-old female (i) and male (ii) DNRII Tg, HY TCR Tg, or DNRII, HY TCR double-transgenic mice were analyzed for CD44hi (MFI, greater than 100) cell-surface expression on the CD8+ T-cell population. Data are represented as the average percentage of CD44hi cells in each T-cell subset; error bars represent the SD in each group (each group included 3 or more mice). (B) Lymphocytes from 8- to 10-week-old female DNRII Tg, 2C TCR Tg, or DNRII, 2C TCR double-transgenic mice were analyzed for CD44hi (MFI, greater than 100) cell-surface expression on the CD8+ T-cell population. Data are represented as the average percentage of CD44hi cells in each T-cell subset; error bars represent the SD in each group (each group included 3 or more mice). (C) Purified, CFSE-labeled DNRII Tg T cells from 10-week-old homozygous Tg mice were injected into TAP-1 KO or age-matched C57BL/6 hosts. Hosts were killed, and lymphocytes from lymph nodes were pooled and analyzed for CFSE+ T cells 1 week after injection. Data are represented as the average percentage of CFSE+, CD8+ T cells in each cell division group. Error bars represent the SD in each group (n = 3), with asterisks denoting significant (P < .05) differences between C57BL/6 and Tap KO hosts. The experiment was performed 3 times with similar results (total n = 10/group).

  • Figure 3.

    Interdependence of CD4+ and CD8+ T-cell subsets. (A) Purified, CFSE-labeled, DNRII Tg T cells from 8-week-old mice were injected into CD4 KO hosts (□) or age-matched C57BL/6 hosts (▪). Hosts were killed, and lymphocytes from lymph nodes were pooled and analyzed for CFSE+ T cells 10 days after injection. Data are represented as the average percentage of CFSE+, CD8+ T cells in each cell division group. Error bars represent the SD in each group (n = 5), and asterisks denote significant (P < .05) differences between C57BL/6 and CD4 KO hosts. (B) Purified, CFSE-labeled DNRII Tg T cells from 8-week-old mice were injected into MHC class II KO hosts (□) or age-matched C57BL/6 hosts (▪). Hosts were killed, and lymphocytes from lymph nodes were pooled and analyzed for CFSE+ T cells 10 days after injection. Data are represented as the average percentage of CFSE+, CD8+ T cells in each cell division group. Error bars represent the SD in each group (n = 5), and asterisks denote significant (P < .05) differences between C57BL/6 and MHC class II KO hosts.

  • Figure 4.

    Flow cytometric analysis of CD8+ T cells from CD8β KO × DNRII Tg mice. Lymphocytes from a representative CD8β KO × DNRII Tg mouse (right panel), a CD8β KO mouse (center panel), and a DNRII Tg mouse (left panel) were analyzed for CD8α and CD8β chains (top panels) or CD44 surface expression on the CD8α-gated lymphocytes (bottom panels). Percentages of gated CD8α+ cells (top panels) are denoted by the numbers adjacent to the indicated oval gates and are represented as percentages of total lymphocyte numbers. Percentages of CD44hi cells in the CD8α population (bottom panels) are indicated to the left of the CD44hi gates. Data represent typical (n > 10) flow cytometric analysis of CD8α+ cells from these strains of mice. All mice studied were older than 16 weeks of age.

  • Figure 5.

    Factor-dependent expansion of CD8+ DNRII Tg T cells. (A) Purified, CFSE-labeled DNRII Tg T cells from 8- to 10-week-old mice were injected into DNRII Tg (right panels) or age-matched C57BL/6 host (left panels). Hosts were killed, and lymphocytes from lymph nodes were pooled and analyzed for CFSE+ T cells 2 weeks after injection. Data shown are from 2 representative mice. Mean of the percentage of CD8+ T cells in each peak is shown below the last row of profiles. (B) Purified, CFSE-labeled DNRII Tg T cells from 8- to 10-week-old mice were injected into IL-15 KO (right panels) or age-matched C57BL/6 host (left panels). Hosts were killed, and lymphocytes from lymph nodes were pooled and analyzed for CFSE+ T cells 2 weeks after injection. Lymphocytes from lymph nodes were analyzed by flow cytometric analysis for CFSE staining. Data shown are for CD8+ T cells from 3 representative mice. Mean of the percentage of CD8+ T cells in each peak is shown below the last row of profiles. (C) Purified, CFSE-labeled DNRII Tg T cells from 8- to 10-week-old mice were injected into IL-15 KO (right panels) or age-matched C57BL/6 hosts (left panels) with subcutaneously implanted mini-osmotic pumps containing either PBS (top panels) or IL-7 (bottom panels). Hosts were killed, and lymphocytes from lymph nodes were pooled and analyzed for CFSE+ T cells 2 weeks after injection. Data shown are for CD8+ T cells from 2 representative mice per group. Mean of the percentage of CD8+ T cells in each peak is shown below the last row of profiles. All experiments were repeated at least 3 times with similar results.

  • Figure 6.

    TGF-β regulation of IL-2/IL-15Rβ expression on CD8+ memory T cells. Purified CD8+ T cells from 8- to 10-week-old C57BL/6 mice were incubated in complete mouse media (gray lines), +IL-15 (black lines), or +IL-15 plus TGF-β (dotted lines) for various times. Cell-surface expression of IL-2/IL-15Rβ (first row), IL-15Rα (second row), and TNFRII (third row) were determined by flow cytometric analysis on the CD8+, CD44hi T-cell population (top panels) and the CD8+CD44lo T-cell population (bottom panels). CFSE-labeled CD8+ T cells were included in each experiment to measure cell division (bottom row). Experiments were repeated 4 times with similar results.