Fig. 6

TGF-β sustains CD73 expression and mitigates 4-1BB-mediated expansion of CD8⁺ T cells. a Purified CD8⁺ T cells were cultured as indicated in the presence of anti-CD3/anti-CD28. After 3 d, the percentage of CD73⁺ in CD8⁺ T cells was measured by flow cytometer. b The mRNA expression of CD73 was measured by real-time RT-PCR. Splenocytes were cultured with indicated reagents in the presence of anti-CD3. After 3 days, the absolute number of total CD8⁺ T cells (c), CD73⁺ versus CD73^- subsets (d) among these treated CD8⁺ T cells were counted. Purified CD8⁺ T cells labeled with efluor450 were cultured in different doses of 5 ng/ml TGF-β with control IgG or anti-4-1BB in the presence of anti-CD3/anti-CD28. The representative flow histograms (e) and summarized results (f) of T cell proliferation indicated by efluor450 dilution were analyzed. g, h Flow analysis of percentage of Ki67⁺ in WT or CD73^−/− CD8⁺ T cells cultured with indicated reagents in the presence of anti-CD3 with/without 5 ng/ml TGF-β. Flow analysis of percentage of CD73⁺ (i, j), or Ki67⁺ (k, l) in WT or CD73^−/− CD8⁺ T cells cultured with indicated reagents in the presence of anti-CD3/anti-CD28 with/without different doses of TGF-β (0.1–10 ng/ml). *p < 0.05, **p < 0.01, ***p < 0.001. Unpaired Student’s two-tailed t test was used. Data (mean ± SEM) are representative of 2 independent experiments