Differential requirements of MHC and of DCs for endogenous proliferation of different T-cell subsets in vivo

Abstract

T cells transferred into severe lymphopenic hosts undergo rapid proliferation known as "endogenous proliferation" that are distinct from conventional homeostatic proliferation. Unlike homeostatic proliferation, cytokines, such as IL-7 are dispensable, yet TCR:MHC interaction is essential for this process to occur. However, cell types inducing the proliferation have not formally been addressed. In this study, we report that CD11c+ conventional DCs play irreplaceable roles in inducing endogenous proliferation of both naive and memory phenotype CD4 T cells via TCR-MHC II interaction. By contrast, CD8 T-cell endogenous proliferation was independent of MHC I or CD11c+ DCs. Interestingly, MHC II was necessary to support naive CD8 T-cell proliferation within MHC I-deficient hosts. Depletion of both B cells and DCs was sufficient to abrogate the proliferation of naive but not of memory CD8 T cells. These results suggest that depending on the T-cell lineages, as well as the differentiation status, different mechanisms control endogenous proliferation, revealing in vivo complexity of T-cell proliferation under lymphopenic conditions.

Fig. 1.

Roles of MHC in naive T-cell EP. (A) CFSE labeled naive Thy1.1 T cells were transferred into groups of TCRβ−/−, MHC II−/− TCRβ−/−, and β2m−/− TCRβ−/− recipients. CFSE profiles were examined 7 days after the transfer. The results shown are representative of six individually tested recipients. (B and C) Lethally irradiated TCRβ−/− mice were reconstituted with BM cells from TCRβ−/−, MHC II−/− TCRβ−/−, and β2m−/− TCRβ−/− donor mice. CFSE labeled naive Thy1.1 T cells were then transferred into the reconstituted recipients after 6 weeks of BM transfer. CFSE profiles were examined 7 days post transfer. Total donor cell recovery from the spleen was examined 16 days post transfer. The mean ± SD of donor T-cell proportion that fully diluted the CFSE (EP) is indicated. (D) MHC I−/− CD8 T cells were generated by bone marrow reconstitution. Naive CD44^low MHC I−/− CD8 T cells isolated from the reconstituted recipients were CFSE labeled and transferred into groups of β2m−/− TCRβ−/− recipients. Shown are the CFSE profiles of TCRβ+/CD8+ gated cells determined 7 days post transfer. The mean ± SD of donor T-cell proportion that fully diluted the CFSE (EP) is indicated.

Fig. 2.

CD11c+ DC dependent CD4 T-cell EP. Groups of TCRβ−/− and DTR Tg TCRβ−/− mice were transferred with Thy1.1 naive T cells and treated with DTX and/or PDCA Ab as described in Methods. CFSE profiles of Thy1.1 donor T cells from the indicated tissues were examined 5 days after the transfer. The results shown are representative of four to six individually tested mice from two independent experiments. The mean ± SD of donor T-cell proportion that fully diluted the CFSE (EP) is indicated.

Fig. 3.

Role of B cells during naive T-cell EP following DC depletion. (A) Groups of DTR Tg TCRβ−/− mice were injected with 2B8 or 18B12 Abs. Six days later the recipients received CFSE labeled Thy1.1 naive T cells and then were subsequently treated with DTX or PBS (days −1 and 2). Recipients were killed 5 days after T-cell transfer. CFSE profiles in the indicated tissues were determined. The mean ± SD of donor T-cell proportion that fully diluted the CFSE (EP) is indicated. (B) Groups of DTR Tg TCRβ−/− mice were injected with 2B8, 18B12, or DTX, as described above. At day 0, 1 × 10⁶ Thy1.1 naive CD8 T cells were transferred into the treated recipients. At day 5 after the transfer surface activation markers (Ly6C, CD44, CD122, and CD127) of Thy1.1+ donor CD8 T cells were determined by FACS analysis. The histograms shown are representative of 3–10 individually tested recipients. The mean ± SD of Ly6C^high (%), CD44^high (MFI), CD122 (MFI), and CD127 (MFI) are indicated. (C and D) Cells from the indicated tissues of the groups described in (B) were stimulated with PMA/Ionomycin and cytokine expression was determined by intracellular staining. Shown are the mean ± SD of three to 10 individually tested mice. *, P < 0.05; **, P < 0.01; ***, P < 0.001.

Fig. 4.

MHC II-dependent CD8 T-cell EP. CFSE labeled naive Thy1.1 CD8 T cells were transferred (1 × 10⁶ per recipient) into groups of TCRβ−/−, TCRβ−/− β2m−/−, and TCRβ−/− β2m−/− MHC II−/− recipients. The recipients were killed 7 days post transfer and CFSE profiles of the donor CD8 T cells were determined by FACS analysis. (A) Histogram profiles shown are representative of two to three individually tested mice. (B) Total Thy1.1 CD8 T cells were enumerated. *, P < 0.05; **, P < 0.01; ***, P < 0.001

Fig. 5.

Role of APCs during memory T-cell EP following DC depletion. Groups of DTR Tg TCRβ−/− mice were injected with 2B8 or 18B12 Abs. Six days later the recipients received CFSE labeled CD44^high CD8 T cells and then were subsequently treated with DTX or PBS (days −1 and 2). Recipients were killed 5 days after T-cell transfer and CFSE profiles were examined. Shown are CFSE profile representatives of three individually tested mice. The mean ± SD of donor T-cell proportion that fully diluted the CFSE (EP) is indicated.