Fig 3

NK cells inhibit CD4⁺ T cells and Tfh cells in vitro. a Rat splenocytes were co-cultured with NK cells or not. Representative flow cytometric plots of Tfh cells (CD4⁺CXCR5⁺ICOS⁺) among CD4⁺ T cells (left). Frequencies of Tfh cells in CD4⁺ T cells were downregulated by NK cells. b, c Mouse splenocytes were co-cultured with NK cells or not. CD4⁺ T cell and Tfh (CD4⁺CXCR5⁺PD1⁺) cell numbers were counted by flow cytometry and shown in relative forms (b, left and median). The Tfh cell frequencies in CD4⁺ T cells were not changed (b, right). The apoptosis of CD4⁺ T cells was determined by staining cells with CD4 and Annexin V (c). d, e Splenic CD3⁺ T were co-cultured with NK cells or not. The CD4⁺ T cells and Tfh cells were determined by flow cytometry (d). The apoptosis of CD4⁺ T cells and Tfh cells was determined by staining cells with Annexin V (e). f, g Mouse splenocytes were cultured with IL-15 or not. IL-15 increased NK cells dramatically in vitro (f, left). IL-15 treatment decreased CD4⁺ T cell numbers and Tfh cell frequencies in CD4⁺ T cells (f, median and right) and induced apoptosis of CD4⁺ T cells (g). h Mouse splenocytes were labeled with CFSE and cultured with IL-15 or not. Cells were then stained with CD19 to detect the proliferation of B cells. Representative flow cytometric histogram of CFSE dilutions of B cells. Data were from one (a, h) or two independent experiments (b–g). Data were presented as mean ± SEM. Unpaired Student’s t test was used. ns means not significant, *p < 0.05, **p < 0.01, ***p < 0.001