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. 2023 May 25;14:1192333. doi: 10.3389/fimmu.2023.1192333

Figure 1.

Figure 1

Selection and in vitro characterization of functional CD146.CARCIK cells. (A) Schematic diagram of CD146.CAR structure, including VL and VH domains of single chain fragment variable (scFv), the spacer sequences: CH2mutCH3 (Long), CH3 (Medium) and CD8stalk (Short), the CD28-OX40 co-stimulatory domains and CD3ζ signaling domain. (B) Percentage of expression of CD146.CAR molecules by CIK cells at day21 of culture, determined by FACS analysis (n=2). (C) Short-term cytotoxicity of untransduced (NT) and CD146.CAR CIK cells co-cultured with CD146+ adherent cell line (TIME) at effector:target (E:T) 5:1 ratio (n=3). (D) Cytokines production (IFN-γ and IL-2) by CD146.CAR CIK cells when engaged with TIME cells. IFN-γ and IL-2 were detected by intracellular staining after 5-hours co-culture at 1:3 E:T ratio(n=3). (E) Proliferation determined by intracellular Ki-67 staining of NT and CD146.CAR CIK cells after co-culture with TIME cells for 72 hours at 1:1 E:T ratio (n=3). (*P<0.05, **P<0.01, Two-way ANOVA analysis with Benjamini and Hochberg’s multiple comparisons test).