Lipopolysaccharide-Induced Lysosomal Cell Death Through Reactive Oxygen Species in Rat Liver Cell Clone 9

In sepsis, bacterial components, particularly lipopolysaccharide (LPS), trigger organ injuries such as liver dysfunction. Although sepsis induces hepatocyte damage, the mechanisms underlying sepsis-related hepatic failure remain unclear. In this study, we demonstrated that the LPS-treated rat hepatocyte cell line Clone 9 not only induced reactive oxygen species (ROS) generation and apoptosis but also increased the expression of the autophagy marker proteins LC3-II and p62, and decreased the expression of intact Lamp2A, a lysosomal membrane protein. Additionally, LPS increased lysosomal membrane permeability and galectin-3 puncta formation, and promoted lysosomal alkalization in Clone 9 cells. Pharmacological inhibition of caspase-8 and cathepsin D (CTSD) suppressed the activation of caspase-3 and rescued the viability of LPS-treated Clone 9 cells. Furthermore, LPS induced CTSD release associated with lysosomal leakage and contributed to caspase-8 activation. Pretreatment with the antioxidant N-acetylcysteine (NAC) not only diminished ROS generation and increased the cell survival rate, but also decreased the expression of activated caspase-8 and caspase-3 and increased the protein level of Lamp2A in LPS-treated Clone 9 cells. These results demonstrate that LPS-induced ROS causes lysosomal membrane permeabilization and lysosomal cell death, which may play a crucial role in hepatic failure in sepsis. Our results may facilitate the development of new strategies for sepsis management.