B-221. Intracellular Salmonella Induce Apoptosis of Human Monocyte-Derived Macrophages through Both Intrinsic and Extrinsic Pathways

H-L. Chen, W-C. Tang, C-H. Chiu;
Chang Gung Children’s Hosp., Chang Gung Univ. Coll. of Med., Taoyuan, TAIWAN.

Salmonella enterica are facultative intracellular pathogens to humans. During infection, Salmonella replicate in host cells and produce cytopathology by inducing host cell death. In this study, we used the human-derived macrophage-like cell line THP-1 to investigate the mechanism involved in cell death induced by Salmonella. Flow cytometry with Annexin V-FITC/PI double staining was used to detect the cell death of macrophages caused by Salmonella. Approximately 10 to 18 % of the cells exhibited apoptosis during the time course from 2 hours to 24 hours post-infection. However, the percentage of cell death (combined necrosis and apoptosis) was increased with the same period of infection (24 to 69 %). Transmission electron microscopy demonstrated typical morphological changes of apoptosis in THP-1 cells 24 hours post-infection, including nuclear condensation, cytoplasm vacuolization and membrane blabbing. To further evaluate the mechanism of Salmonella-induced cell death, we tested the effect of different moi (10, 100 and 500) in the following experiments. By western blot hybridization, we found that the caspase-8 and BAX expressions were significantly increased only in the condition of moi 500. Higher multiplicity of bacterial infection proportionately induced more apoptosis, as shown by the increase of caspase-3 activity over time. However, this caspase-3 expression was blocked by the treatment with either Z-IETD-FMK (caspase-8 inhibitor) or Z-LEHD-FMK (caspase-9 inhibitor), with the later being more inhibitory. The results indicated that both extrinsic and intrinsic apoptosis pathways are involved in the intracellular Salmonella-induced cell death, and the later may play a more important role. Unraveling of the associated mechanisms of Salmonella-induced cell death may represent an unexploited avenue for therapeutic intervention of Salmonella infection.