Fig. 1. Metformin inhibits both PD-L1 expression on cancer cells and PD-1 expression on infiltrating immune cells in the TME. (A) Schematic diagram of the experimental procedures. B16F10 murine melanoma cells were subcutaneously injected into C57BL/6 mice. From day 3 after tumor implantation, the mice received an intraperitoneal injection of metformin or PBS daily. On day 20, after tumor implantation, the mice were sacrificed. (B) Representative images of B16F10 tumors. Scale bar, 1 cm. (C) Growth curve of subcutaneous B16F10 tumors derived from metformin-treated (red line; N=5) and PBS-treated (blue line; N=5) mice. (D) Tumor weight of subcutaneous B16F10 tumors on day 20 derived from metformin-treated (red line; N=5) and PBS-treated (blue line; N=5) mice. (E) Cell-surface analysis of PD-L1 protein expression in B16F10 single-cell suspensions derived from B16F10 tumors using flow cytometry. A representative histogram (left panel) and graph (right panel) are shown. (F) Cell-surface analysis of PD-1 protein expression in NK or CD8⁺ T single-cell suspensions derived from B16F10 tumors using flow cytometry. A representative histogram (left panel) and graph (right panel) are shown. *P<0.05; **P<0.01; ***P<0.001, Student’s t-test.
Abbreviations: PD-L1, programmed death ligand 1; PD-1, programmed death 1; TME, tumor microenvironment; PBS, phosphate-buffered saline; NK, natural killer.

Ann Lab Med 2024;44:426~436 https://doi.org/10.3343/alm.2023.0443