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. 2021 Mar 25;12:625808. doi: 10.3389/fimmu.2021.625808

Figure 2.

Figure 2

VSIG3 protein inhibits T cell function. (A) VSIG3 expression by CHO cells was detected by western blot after 2-8 days of culturing. (B) The VSIG3 protein was eluted with different concentrations of imidazole solutions (20 mM, 40 mM, 60 mM, 80 mM, 250 mM, 500 mM) (Left); Coomassie blue stained PAGE of concentrated VSIG3 protein (Right). (C) VSIG3 inhibits the cytokine secretion of activated PBMCs. Anti-human CD3 antibody (1 μg/mL) and different concentrations of VSIG3-ECD protein were coated on 96-well plates. PBMCs (1×105 cells/well) were added for 48 hours to detect the secretion of IFN-γ and TNF-α in the supernatant. **p<0.01 and ***p<0.001 vs. control. (D) VSIG3 inhibits the secretion of IFN-γ and TNF-α in activated human CD4+ T cells. Anti-human CD3 antibody (1 μg/mL) and different concentrations of VSIG3-ECD protein were coated on 96-well plates. Human CD4+ T cells (1×105 cells/well) were added for 48 hours to detect the secretion of IFN-γ and TNF-α in the supernatant. **p<0.01, ***p<0.001 and ****p<0.0001 vs. control. (E) VSIG3 inhibits the proliferation of activated PBMCs. CFSE-labeled PBMCs (1×105 cells/well) were stimulated by plate-bound anti-human-CD3 (1 μg/mL), and VSIG3 protein (10 μg/mL) was added. The proliferation levels of PBMCs were measured by flow cytometry on day 5. Representative results from three independent experiments are shown.