Astaxanthin (ASX), a red‑colored xanthophyll
carotenoid, functions as an
antioxidant or pro‑oxidant. ASX displays anticancer effects by reducing or increasing oxidative stress.
Reactive oxygen species (ROS) promote
cancer cell death by necroptosis mediated by receptor‑interacting
protein kinase 1 (RIP1) and RIP3.
NADPH oxidase is a major source of ROS that may promote necroptosis in some
cancer cells. The present study aimed to investigate whether ASX induces necroptosis by increasing
NADPH oxidase activity and ROS levels in
gastric cancer AGS cells. AGS cells were treated with ASX with or without
ML171 (
NADPH oxidase 1 specific inhibitor), N‑acetyl
cysteine (NAC;
antioxidant), z‑VAD (pan‑caspase inhibitor) or Necrostatin‑1 (Nec‑1; a specific inhibitor of RIP1). As a result, ASX increased
NADPH oxidase activity, ROS levels and cell death, and these effects were suppressed by
ML171 and NAC. Furthermore, ASX induced RIP1 and RIP3 activation, ultimately inducing mixed lineage
kinase domain‑like
protein (MLKL) activation,
lactate dehydrogenase (LDH) release and cell death. Moreover, the ASX‑induced decrease in cell viability was reversed by Nec‑1 treatment and RIP1
siRNA transfection, but not by z‑VAD. ASX did not increase the ratio of apoptotic Bax/anti‑apoptotic Bcl‑2, the number of Annexin V‑positive cells, or caspase‑9 activation, which are apoptosis indices. In conclusion, ASX induced necroptotic cell death by increasing
NADPH oxidase activity, ROS levels, LDH release and the number of
propidium iodide‑positive cells, as well as activating necroptosis‑regulating
proteins, RIP1/RIP3/MLKL, in
gastric cancer AGS cells. The results of this study demonstrated the necroptotic effect of ASX on
gastric cancer AGS cells, which required
NADPH oxidase activation and RIP1/RIP3/MLKL signaling in vitro.