Fig. 3

Lymphocytes cytotoxicity mediated by AdCD3-HAC was enhanced by 5-FU through upregulation of CAR and αvβ3. a ELISA analyses for IFN-γ on supernatants from 231.CD3scfv cells incubated with PBMCs for 3 days followed by that floating cells were harvested for second-round co-incubation with or without HAC for 5 days. b Jurkat T cells were incubated either alone or in co-culture with various virus-loaded MDA-MB-231 cells at the ratio of 1:10 for 10 h. Apoptosis of Jurkat cells was determined by flow cytometry using FITC-Annexin V. c Representative images show the percentages of apoptotic Jurkat cells. d Cytotoxicity of PBMCs to MDA-MB-231 cells infected with different low MOIs of AdCD3-HAC (E:T = 10:1) with or without 5-FU. e Cytokines including IL-2, IFN-γ, and TNF-α in the corresponding co-culture supernatants from d were detected by ELISA. f Flow cytometry analysis performed on the infection efficiencies of adenovirus to MDA-MB-231 cells, which were pretreated with 5-FU (0, 0.25, and 0.5 μg/mL) for 48 h, at different MOIs. g–i Flow cytometry analysis on the expression level of CAR, αvβ3, and PD-L1 on the surface of MDA-MB-231 cells and MCF-7 cells treated with low doses of 5-FU for 48 h. j Mean fluorescence intensity of PD-L1 on MDA-MB-231 cells treated with 5-FU. SD shown, n ≥ 3. Two-tailed unpaired t test for a and j and one-way ANOVA with Tukey’s post-test for b, e, f, g, h, and i. *P < 0.05; **P < 0.01; ***P < 0.001;****P < 0.0001