METHODS: AgNP-SC was synthesised by reducing silver nitrate with an aqueous extract of S. crispus, with nanoparticle formation indicated by the appearance of a dark brown colour. Cytotoxicity was assessed using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. Apoptotic changes were examined through acridine orange/propidium iodide (AO/PI) staining, while cell cycle distribution was analysed using flow cytometry. Gene expression of apoptotic markers was analysed using reverse transcription polymerase chain reaction (RT-PCR).

RESULTS: MTT analysis demonstrated a significant, concentration-dependent decrease in CT26 cell viability following AgNP-SC treatment, with cell viability reduced to below 50% at concentrations ≥ 25 μg/mL (P < 0.05). AO/PI staining revealed classical apoptotic features, including chromatin condensation, membrane blebbing, and nuclear fragmentation. Flow cytometric analysis showed a significant accumulation of cells in the G1 phase accompanied by a marked reduction in the S-phase population in treated groups compared to untreated controls (P < 0.05), indicating G1 cell cycle arrest and inhibition of DNA synthesis. RT-PCR analysis further demonstrated significant upregulation of pro-apoptotic genes (CED-3 and CED-4) and downregulation of the anti-apoptotic gene CED-9 (P < 0.05), confirming activation of the intrinsic mitochondrial apoptotic pathway.