Neferine induces p38 MAPK/JNK1/2 activation to modulate melanoma proliferation, apoptosis, and oxidative stress

Abstract

BACKGROUND: Melanoma is a malignant skin cancer that has a poor prognosis in advanced patients. The aim of the present study was to investigate the antitumor role of neferine in melanoma. METHODS: A375 and C32 cells were selected as research vectors in vitro. Cell counting Kit-8, 5-ethynyl-2'-deoxyuridine staining, transwell, and flow cytometry assay were used to examined cell malignant phenotypes. Mitochondrial dysfunction was detected by 5,50,6,60-tetrachloro-1,10,3,30-tetraethyl-imidacarbocyanine iodide staining and enzyme-linked immunosorbent assay. Reactive oxygen species (ROS) generation was measured using oxidation sensitive fluorescent probe. The phosphorylation activity of p38 and Jun-N-terminal kinase (JNK) 1/2 were examined by Western blot. A xenograft model was established via the subcutaneous injection of A375 cells into the right flank of BALB/c mice in vivo. RESULTS: Neferine (2.5, 5, or 10 µM) treatment inhibited proliferation, invasion, and enhanced apoptotic rate of A375 and C32 cells. Neferine treatment induced abnormal changes in mitochondrial membrane potential. Further studies showed that neferine could significantly increase the production of reactive oxygen species (ROS) and 3,4-methylenedioxyamphetamine (MDA) content, decreased the superoxide dismutase (SOD) level. Neferine (5, 10, or 20 mg/kg) obviously suppressed the weight and size of the xenograft tumor, the number of apoptotic cells in vivo, and the expression of Ki67(+) and survivin(+) decreased. Notably, neferine also activated the phosphorylation of p38 and JNK1/2. CONCLUSIONS: Neferine inhibits the proliferative and invasion ability of melanoma cells and promotes their apoptosis, ameliorating the malignant progression of melanoma, likely achieved by upregulating the phosphorylation levels of p38 mitogen-activated protein kinase and JNK1/2.