Objective      To determine the effect of CGI-58 on 5-fluorouracil(5-FU) induced macrophage apoptosis

and investigate the underlying mechanism.        Methods      CCK-8 assay was used to detect the effect of 5-FU on the proliferation of primary macrophage (PLKO) and CGI-58 knockdown macrophage (CGI-58 KD). Flow cytometry was used to test the cell cycle and apoptosis of PLKO and CGI-58 KD which were induced by 5-FU. Western blotting was employed to detect apoptosis protein (cleaved Caspase-3) and expression levels of 5-FU induced PLKO and CGI-58 KD. Seahorse assay was applied to test the mitochondrial function of PLKO and CGI-58 KD.       Results      CCK-8 assay found that the cell vitality of CGI-58 KD+5-FU group was reduced significantly (P0.01). Flow cytometry results showed that after 5-FU induction

the apoptotic rate of CGI-58 KD group was significantly higher than that of PLKO group (P0.01). Western blotting showed that the expression level of cleaved Caspase-3 in CGI-58 KD+5-FU group was obviously higher than that in PLKO+5-FU group (P0.01). Seahorse assay detected that the oxygen consumption rate of CGI-58 KD group was significantly decreased (P0.01)

ATP generation was reduced (P0.05)

and ROS expression was increased (P0.01). Antioxidant N-acetyl-L-cysteine could antagonize ROS increase induced by CGI-58 deletion (P0.05)

5-FU killing effect (P0.05) and expression of cleaved Caspase-3.       Conclusion      CGI-58 deletion can strengthen the killing effect of 5-FU on macrophage and damage the mitochondrial function of macrophage through increasing ROS.