Abstract
Saikosaponin a (SSa), a bioactive phytochemical from Bupleurum, triggers sequential caspase-2 and caspase-8 activation, and thereby induces caspase-mediated apoptosis in human colon carcinoma (HCC) cells. However, the upstream mechanism of caspase-2 activation remains unknown. Therefore, we investigated the signaling mechanisms underlying SSa-induced caspase activation and apoptosis in HCC cells. SSa treatment triggered marked antitumor effects, especially in HCC cells, in a cell culture model and a mouse xenograft model. SSa also induced the activation of several endoplasmic reticulum (ER) stress signals. Specifically, caspase-4, a critical regulator of ER stress-induced apoptosis, was activated significantly after SSa treatment. Mechanistically, selective inhibition of caspase-4 suppressed SSa-induced apoptosis, colony inhibition, and the activation of caspase-3, -8, and -2, but not vice versa. Consistent with the important role of caspase-2 in the DNA damage response, SSa induced DNA damage, as evidenced by a cytokinesis-block micronucleus assay, single-cell gel electrophoresis, and an increase in the levels of γ-H2AX, a DNA damage marker. Moreover, inhibition of caspase-4 activation inhibited SSa-induced histone H2AX phosphorylation. Taken together, these results suggest that caspase-4 is an upstream regulator of SSa-induced DNA damage and caspase activation in HCC cells. Given that SSa-induced apoptosis appeared to be specific to certain cell types including HCC cells, SSa may be a promising cancer therapy agent in certain types of cancer.
Highlights
Human caspase-4 and mouse caspase-12, which are localized in the endoplasmic reticulum (ER), are cleaved by ER stress-inducing agents and are involved in the ER stress response [1,2,3]
These results indicate that Saikosaponin a (SSa) can trigger apoptotic cancer cell death in a cell type-dependent manner
Because 20 μM SSa led to significant cytotoxicity in all colon cancer cells tested, we used this concentration for subsequent experiments
Summary
Human caspase-4 and mouse caspase-12, which are localized in the endoplasmic reticulum (ER), are cleaved by ER stress-inducing agents and are involved in the ER stress response [1,2,3]. Caspase-4 activation is essential for ER stressinduced apoptosis in some cell lines [1, 7,8,9]. At least one study has suggested that caspase-4 is not essential for ER stress-induced apoptosis [10]; the role of caspase-4 in ER stress-induced apoptosis remains unclear. Our group found that SSa could induce the activation of various caspases and poly (ADP-ribose) polymerase (PARP), decrease Bcl-2 and X-linked inhibitor of apoptosis (XIAP) expression, and induce apoptosis in human colon carcinoma (HCC) cells [14]. SSa has been shown to have cytotoxicity against human gastric cancer, breast cancer, and hepatoma [15,16,17,18]
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