Proteasome inhibition promotes autophagy and protects from endoplasmic reticulum stress in rat alveolar macrophages exposed to hypoxia-reoxygenation injury.

Alveolar macrophages play vital roles in acute lung injury, and macrophage response to hypoxia play relevant roles to disease mechanisms. There is growing evidence that cell death pathways play crucial roles in physiological and pathological settings and that the ubiquitin-proteasome system is involved in the regulation of these processes. However, the functional role of proteasome in alveolar macrophages exposed to hypoxia-reoxygenation (H/R) injury is unknown. We aimed to investigate the function of proteasome on alveolar macrophages exposed to H/R and the underlying mechanisms. NR8383 cells were pretreated with proteasome activator sulforaphane (SFN) or inhibitor MG-132 for 1 hr, and then submitted to 2/6 hr, 4/6 hr, and 6/6 hr H/R treatment. Cell viability was assessed with MTT assay. Autophagy was monitored using electron transmission microscope and flow cytometry and western blotting. The endoplasmic reticulum (ER) stress and unfolded protein response (UPR) pathways were equally analyzed by western blotting. Cell apoptosis was detected by immunohistochemistry, caspase3/7 activity, and western blotting. The viability of NR8383 cells exposed to H/R was affected by proteasome activity and proteasome inhibition significantly inhibited cell death. Treatment with MG-132 led to autophagy activation and induced the survival of NR8383 cells exposed to H/R. Pretreatment with SFN significantly decreased cell autophagy and induced cell death. ER stress was activated in H/R-treated NR8383 cells, and SFN further promoted ER stress whereas proteasome inhibition led to contrary results. Proteasome inhibtion hindered cell apoptosis as demonstrated by decreased caspase-3/7 activity, immunolabelling, and western blot results. Proteasome inhibition might be a promising approach for treating H/R injury-related lung diseases.