Abstract
BackgroundSaffron extract, a natural product, has been shown to induce apoptosis in several tumor cell lines. Nevertheless, the p53-dependency of saffron’s mechanism of action in colon cancer remains unexplored.Material and methodsIn order to examine saffron’s anti-proliferative and pro-apoptotic effects in colorectal cancer cells, we treated two p53 isogenic HCT116 cell lines (HCT wildtype and HCT p53−/−) with different doses of the drug and analyzed cell proliferation and apoptosis in a time-dependent manner. MTT viability and crystal violet assays were performed in order to determine the effective dose of saffron on both cell lines. The cell cycle progress was examined by Flow cytometric analysis. Apoptosis was assessed using Annexin-PI-staining and Western Blotting for caspase 3 and PARP cleavage. Autophagy was determined by Western Blotting of the light chain 3 (LC3)-II and Beclin 1 proteins. The protein content of phospho-H2AX (γH2AX), a sensor of DNA double strand breaks, was also analyzed by Western Blotting.ResultsSaffron extract induced a p53-dependent pattern of cell cycle distribution with a full G2/M stop in HCT116 p53 wildtype cells. However, it induced a remarkable delay in S/G2 phase transit with entry into mitosis in HCT116 p53 −/− cells. The apoptotic Pre-G1 cell fraction as well as Annexin V staining and caspase 3 cleavage showed a more pronounced apoptosis induction in HCT116 p53 wildtype cells. Obviously, the significantly higher DNA-damage, reflected by ɣH2AX protein levels in cells lacking p53, was coped by up-regulation of autophagy. The saffron-induced LC3-II protein level was a remarkable indication of the accumulation of autophagosomes, a response to the cellular stress condition of drug treatment.ConclusionsThis is the first study showing the effect of saffron in HCT116 colorectal cancer cells with different p53 status. Saffron induced DNA-damage and apoptosis in both cell lines. However, autophagy has delayed the induction of apoptosis in HCT116 p53 −/− cells. Considering the fact that most tumors show a functional p53 inactivation, further research is needed to elucidate the long-term effects of saffron in p53 −/− tumors.
Highlights
Saffron extract, a natural product, has been shown to induce apoptosis in several tumor cell lines
This is the first study showing the effect of saffron in HCT116 colorectal cancer cells with different p53 status
Autophagy has delayed the induction of apoptosis in HCT116 p53 −/− cells
Summary
A natural product, has been shown to induce apoptosis in several tumor cell lines. Colorectal cancer is one of the most common malignancies in the world with approximately one million new cases diagnosed every year [1]. Natural products encompass three main categories of compounds, phenylpropanoids, isoprenoids, and alkaloids, which are widely distributed in plant foods and medicinal herbs [3,4,5]. This large array of molecules is crucial to human nutrition and health. The dried styles (stigmata) of Crocus sativus L. flowers, is widely known as a spice and its uses in traditional medicine are well established and date back nearly 3000 years, spanning many continents, civilizations, and cultures [7,8]
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