Citrus wastes trigger p53-mediated apoptosis in human colon cancer cells

Despite the well-documented pharmacological benefits and anticancer properties of Citrus fruits, the precise molecular mechanisms driving their cytotoxicity in cancer cells remain elusive. This study elucidates the roles of nitric oxide (NO•) and inhibitor of apoptosis proteins (IAPs) in apoptosis triggered by Citrus -derived by-products—specifically peels, immature fruits, and juice processing residues—in p53-wild-type and p53-null HCT116 colon carcinoma cells. Cellular proliferation and apoptosis were systematically assessed utilizing Trypan blue exclusion and Annexin V/propidium iodide staining. To determine NO• production and the expression profiles of key apoptotic regulators (Bcl-2, Bax, PARP, and IAPs), we employed Griess assays, quantitative RT-PCR, and western blotting, complemented by colorimetric evaluations of caspase-3 activity. Treatment with Citrus by-products effectively suppressed cell viability and induced apoptosis, strongly correlating with a dose- and time-dependent accumulation of endogenous NO•. Furthermore, both cell lines exhibited significant downregulation of anti-apoptotic Bcl-2 and critical IAPs, including survivin, cIAP-1/2, and XIAP. This was accompanied by marked PARP cleavage and enhanced caspase-3 activity, underscoring their integral roles in mediating the apoptotic cascade. Collectively, our findings demonstrate that apoptosis induced by Citrus by-products is heavily dependent on the cellular p53 status and is mechanistically driven by the modulation of endogenous NO• generation and the suppression of IAP family proteins.