Cholesterol secoaldehyde induces apoptosis in H9c2 cardiomyoblasts through reactive oxygen species involving mitochondrial and death receptor pathways.

Cholesterol secoaldehyde (ChSeco), a putative product of the reaction of ozone with cholesterol in aqueous environments, has been shown to induce apoptosis in H9c2 cardiomyoblasts. This study further investigated the involvement of apoptotic-related proteins and gene expression using RT-PCR, Western blot, and appropriate biochemical assays. The RT-PCR analysis revealed that ChSeco activates the expression of genes involved in the death receptor (extrinsic) pathway. The significance of this pathway was also evident from the increased activity of caspase-8. The overexpression of Apaf-1, loss of mitochondrial transmembrane potential, release of cytochrome c, and increased activity of caspase-9 provide further evidence for the involvement of a mitochondrial (intrinsic) pathway. Time-course analysis of ChSeco-exposed H9c2 cells showed an upstream increase in the generation of reactive oxygen species (ROS) and an associated decrease in the intracellular glutathione. N-acetyl-L-cysteine and Trolox significantly attenuated the ChSeco-induced ROS formation and cytotoxicity and also down-regulated the expression of the genes of all the players in either pathway. This study clearly shows that ChSeco induces apoptosis in H9c2 cells through ROS generation and the activation of both the intrinsic and the extrinsic pathway.