Hepatic Ischemia-reperfusion Injury in Mice was Alleviated by Rac1 Inhibition - More Than Just ROS-inhibition

Background and Aims: Reducing reactive oxygen species (ROS) production has proven an effective way for allevi-ating oxidative stress during ischemia-reperfusion injury (IRI). Moreover, inhibition of Rac1 could reduce ROS pro-duction and prevent oxidative stress injury. Previous stud-ies have suggested a positive interactivation feedback loop between Rac1 and hypoxia-inducible factor (HIF)-1 alpha, the latter being up-regulated early during ischemia. The posi-tive inter-activation between Rac1 and HIF-1 alpha would ag-gravate ROS production, thereby promoting IRI. This study was designed to verify the effects of Rac1 inhibition on he-patic IRI both at animal and cellular levels and to explore the interaction between Rac1 and HIF-1 alpha during hepatic IRI. Methods: C57B/6 mice and AML-12 cells were used for the construction of hepatic IRI animal and cell models. Rac1 inhibition was achieved by NSC23766 (a specific Rac1 inhibitor). Lentiviral vectors were used for Rac1 knock -down. At designated time points, serum and liver tissues were collected from the mice and treated cells were col-lected for further analysis. Results: NSC23766 treatment significantly alleviated the hepatic IRI in mice, manifesting as lower vacuolation score and less apoptosis cells, lower ROS and serum/liver alanine aminotransferase/aspartate aminotransferase levels, and fewer activated inflammatory cells. IRI of AML-12 was also alleviated by 50 mu M NSC23766 or Rac1-knockdown, manifesting as reduced cell apoptosis, less extensive interruption of mitochondrial membrane po-tential, down-regulation of apoptosis, and effects on DNA damage-related proteins. Interestingly, Rac1 knockdown also down-regulated the expression level of HIF-1 alpha. Conclusions: Our study supports a protective effect of Rac1 inhibition on hepatic IRI. Aside from the classic topics of reducing ROS production , oxidative stress, our study showed an interaction between Rac1 and HIF-1 alpha signaling during hepatic IRI.