Effect of RIP3 expression on apoptosis of mouse macrophages infected with BCG

AIM:To investigate the function of receptor-interacting proteins 3 (RIP3) in regulating Bacillus Calmette-Guérin (BCG)-induced apoptosis of mouse macrophages (RAW264.7 cells). METHODS:The RIP3 adenovirus interference vector was constructed and used to infect the RAW264.7 cells, and then the RAW264.7 cells were infected with BCG. The cell viability was measured by MTT assay. The apoptotic rate, mitochondrial membrane potential and production of reactive oxygen species (ROS) were determined by flow cytometry analysis. The protein levels of RIP3 and apoptosis-associated proteins were examined by Western blot. RESULTS:The viability of RAW264.7 cells was decreased after BCG infection. In the meantime, the expression of RIP3 was up-regulated significantly (P<0.01). Compared with BCG infection group, the apoptotic rate and ROS level in BCG and RIP3 adenovirus interference vector co-infection group were significantly decreased (P<0.01). Importantly, RIP3 was able to further promote apoptosis in BCG-infected RAW264.7 cells in part by increasing mitochondrial membrane potential (P<0.01). In addition, Western blot analysis further demonstrated that RIP3 was involved in BCG-induced apoptosis partly through down-regulation of anti-apoptotic protein Bcl-2, and up-regulation of Bax and cleaved caspase-3 (P<0.01). CONCLUSION:RIP3 is involved in BCG-induced apoptosis of RAW264.7 cells, and this process may be achieved by the mitochondrial pathway.