Ginsenoside Ro improves <i>Salmonella</i> Typhimurium‐induced colitis through inhibition of the virulence factors <scp>SopB</scp> and <scp>SopE2</scp> via the <scp>RAC1</scp> / <scp>CDC42</scp> / <scp>ARP2</scp> /3 pathway

Salmonella enterica serovar Typhimurium (S. Typhimurium) poses a serious threat to human and animal health, and there is an urgent need to develop new therapeutic agents. In our in vivo study, ginsenoside Ro (Ro) reduced the mortality rate of S. Typhimurium-infected mice by effectively improving three key disease activity index (DAI) indicators. In particular, ginsenoside Ro inhibited S. Typhimurium-induced colitis by reversing colon length shortening; alleviating pathological damage to the colon; decreasing the levels of IL-1β, TNF-α, IFN-γ, and IL-6; and decreasing the activities of MPO and EPO, while increasing the levels of IL-10 as well as the colon epithelial barrier and tight junction-related genes (Mucin 1, Mucin 2, Occludin, Claudin-3, and ZO-1). Furthermore, ginsenoside Ro reduced CFUs in the liver, spleen, colon, and feces. In a mechanistic in vitro study, ginsenoside Ro reduced CFUs in HeLa and Raw264.7 cells, which was associated with ginsenoside Ro inhibition of the recruited S. Typhimurium-containing vacuole (SCV) biomarkers LC3, Rab7, GAL8, and NDP52. Molecular docking results revealed that the binding energies of ginsenoside Ro and SopB and ginsenoside Ro and SopE2 were as high as -11.3 and -9.7 kcal/mol, respectively, as verified by CETSA and DARTS assays. Moreover, ginsenoside Ro at 100 and 200 μM significantly decreased the enzyme activities and expression of SopB and SopE2. Finally, ginsenoside Ro inhibited the membrane ruffling caused by SopB-regulated Arf6/Cyth2/Arf1-, RAC1-, and CDC42-driven Arp2/3-dependent actin polymerization and the SopE2-regulated CDC42/Arp2/3 signaling pathway. In summary, our findings suggest that ginsenoside Ro is a potential lead compound for therapeutic use against S. Typhimurium infection, and these findings lay a foundation for its further development.