A pan caspase inhibitor decreases caspase-1, IL-1α and IL-1β, and protects against necrosis of cisplatin-treated freshly isolated proximal tubules

Abstract

Caspase-1, IL-1α, and IL-1β are known to be activated in the NLRP3 inflammasome. The inflammasome is activated mostly in inflammatory cells. The presence of inflammasome proteins in proximal tubules (PTs) and the effect of cisplatin-treatment or caspase inhibition on inflammasome proteins in PTs are not known. The aim of this study was to investigate the effect of cisplatin on inflammasome proteins in freshly isolated PTs and also to determine the effect of caspase inhibition on inflammasome proteins and PT injury. PTs were isolated using collagenase digestion and Percoll centrifugation. After recovery period, freshly isolated PTs were incubated with vehicle, 50μM cisplatin or 50μM cisplatin plus 50μM pan caspase inhibitor, QVD-OPH. PTs treated with 50μM cisplatin showed Propidium Iodide staining indicative of necrosis. Necrotic cells (%) were 2.2 in Vehicle-treated, 37.7 in Cisplatin-treated (p<0.05 vs. Vehicle), and 3.3 in QVD-treated (p<0.05 vs. Cisplatin). LDH release (%), a marker of cell membrane damage seen in necrosis was 7.1 in Vehicle-treated, 39.7 in Cisplatin-treated (p<0.05 vs. Vehicle), and 13.5 in QVD-treated (p<0.05 vs. Cisplatin). Caspase-1 activity and active caspase-1 protein (10kDa) were significantly increased in Cisplatin-treated PTs. NLRP3 was strongly expressed in PTs, but there were no significant changes between groups. Pro-apoptotic BID (22kDa) was unchanged between groups. IL-1α and IL-1β activity was increased in Cisplatin-treated PTs. QVD-OPH co-treatment decreased caspase-1, IL-1α, and IL-1β. In summary, caspase inhibition decreases caspase-1, IL-1α, and IL-1β but not NLRP3 or BID protein and protects against necrosis in cisplatin-treated freshly isolated PTs.