Effects of sanguinarine on murine myocyte gene expression

Sanguinarine (13-methyl[1,3]benzodioxolo[5,6-c]-1.3-dioxolo[4,5-i]phenanthridinium) is derived from the root of Sanguinaria canadensis L. (blood root), Chelidonium majus L. (Greater Celandine) and other poppy-fumaria species. Sanguinarine belongs to the benzophenanthridine alkaloid group, which are biosynthesized from phenylalanine in plants of the papaveraceae. This alkaloid has been reported to display a wide spectrum of biological effects, such as antifungal, antimicrobial, analgesic and moreover, anti-cancer properties via the induction of cell death. Utilizing dose-response studies with sanguinarine, we have shown that at two different concentrations sanguinarine induced over 95% apoptosis and oncosis respectively, in several cancer cell lines Also, the fact that oncosis and apoptosis are both induced in the same cell line under similar conditions and time points (with the sole exception being the concentration used) makes it a highly attractive model, because it facilitates easy and accurate comparison of both forms of cell death. These qualities make sanguinarine an ideal tool to study and compare oncosis with apoptosis. We are not aware of any other cell model system used to study oncosis that has all the above advantages.The objective of this study was to provide insight into the pathogenetic mechanisms of oncosis and apoptosis in cardiac myocytes in ischemic/reperfusion injury so that appropriate interventions could be designed to minimize injury and preserve myocardial function. We have shown here that primary cardiomyocytes, when exposed to sanguinarine at concentrations of 1.5 µg/ml (low dose) and 25 µg/ml (high dose) for 1 hour, displayed the classic biphasic pattern of apoptosis and oncosis. Using Illumina whole-genome expression microarrays, we have identified over 488 probes that were differentially expressed at p < 0.001 in cardiac myocytes between the low dose and high dose of sanguinarine after 1 hour of treatment; 78 of these changed more than 2-fold. Thus far, oncosis/necrosis has been thought of as uncontrolled and accidental, and lacking the features of apoptosis and autophagy. However, our findings and that of others reported in the literature indicate that this occurrence is not accidental but a tightly regulated physiological process. To our knowledge, this is the first report to present a model for the comparative study of apoptosis vs. oncosis in primary cardiomyocytes. Cultured murine myocytes were cultured under standard conditions and treated for 1 hour with either vehicle (EtOH), 1.5 ug/ml, or 25 ug/ml sanguinarine. 3 separate experiments were done for a total of 7 biological replicates for control, 4 for 1.5 ug/ml dose and 8 for the 25 ug/ml dose. Total RNA was extracted and processed using standard Illumina protocols.