Fig. 4

Cytotoxicity of T cells towards MM.1S cells mediated by Tandab (CD3/BCMA) secreted from UCMSCs. In order to functionally validate the MSC-secreted Tandab (CD3/BCMA), a co-culture system using transwell plates with a 0.4-μm pore-size membrane was established. MSCs were plated into 24-well plates at a density of 2 × 10⁴ cells per well after transduction with the lentiviral vector. Seventy-two hours later, MM.1S cells were labeled with CFSE. Then, PBMCs and labeled MM.1S cells (effector to target (E:T) ratio of 10:1) were added to the equilibrated inserts. After co-culture for 24 h, cells in the inserts were labeled with 7-AAD and detected by flow cytometry. (A) Specific lysis of MM.1S cells. (B) Activation surface markers CD25, CD69, and CD107a of T cells. (C) Cytokines including IL-2, IFN-γ, and TNF-α in the supernatant were measured using ELISA kits. (D) Representative flow cytometry analysis of the proliferation of PBMCs (labeled with CellTrace™ Far Red) after co-culture with MM.1S at an E:T ratio of 10:1 in the collected supernatants of the infected UCMSCs for 72 h (left) and statistical analysis of the data (right). *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001 compared with the corresponding MSC-EV group. The data represent the means ± SD from three repeated experiments