Induction of ROS-Dependent Mitochondria-Mediated Intrinsic Apoptosis in MDA-MB-231 Cells by Glycoprotein from Codium decorticatum

Marine macroalgae consist of a range of bioactive molecules exhibiting different biological activities, and many of these properties are attributed to sulfated polysaccharides, fucoxanthin, phycobiliproteins, and halogenated compounds. In this study, a glycoprotein (GLP) with a molecular mass of ∼48 kDa was extracted and purified from Codium decorticatum and investigated for its cytotoxic properties against human MDA-MB-231 breast cancer cells. The IC₅₀ values of GLP against MDA-MB-231 and normal breast HBL-100 cells (control) were 75 ± 0.23 μg/mL (IC₂₅), 55 ± 0.32 μg/mL (IC₅₀), and 30 ± 0.43 μg/mL (IC₇₅) and 90 ± 0.57 μg/mL (IC₂₅), 80 ± 0.48 μg/mL (IC₅₀), and 60 ± 0.26 μg/mL (IC₇₅), respectively. Chromatin condensation and poly(ADP-ribose) polymerase (PARP) cleavage studies showed that the GLP inhibited cell viability by inducing apoptosis in MDA-MB-231 cells. Induction of mitochondria-mediated intrinsic apoptotic pathway by GLP was evidenced by the events of loss of mitochondrial membrane potential (ΔΨ(m)), bax/bcl-2 dysregulation, cytochrome c release, and activation of caspases 3 and 9. Apoptosis-associated factors such as reactive oxygen species (ROS) formation and loss of ΔΨ(m) were evaluated by DCFH-DA staining and flow cytometry, respectively. Cell cycle arrest of G₂/M phase and expression of apoptosis associated proteins were determined using flow cytometry and Western blotting, respectively.