The involvement of mitochondrial apoptotic pathway in eugenol-induced cell death in human glioblastoma cells

- We explored the effect of eugenol on human glioblastoma cells.
- Eugenol induced Ca2+ movement including Ca2+ release and Ca2+ entry.
- Eugenol activated the mitochondrial apoptotic pathway.

Eugenol, a natural phenolic constituent of clove oil, has a wide range of applications in medicine as a local antiseptic and anesthetic. However, the effect of eugenol on human glioblastoma is unclear. This study examined whether eugenol elevated intracellular free Ca2+ levels ([Ca2+]i) and induced apoptosis in DBTRG-05MG human glioblastoma cells. Eugenol evoked [Ca2+]i rises which were reduced by removing extracellular Ca2+. Eugenol-induced [Ca2+]i rises were not altered by store-operated Ca2+ channel blockers but were inhibited by the PKC inhibitor GF109203X and the transient receptor potential channel melastatin 8 (TRPM8) antagonist capsazepine. In Ca2+-free medium, pretreatment with the endoplasmic reticulum Ca2+ pump inhibitor thapsigargin (TG) or 2,5-di-tert-butylhydroquinone (BHQ) abolished eugenol-induced [Ca2+]i rises. The phospholipase C (PLC) inhibitor U73122 significantly inhibited eugenol-induced [Ca2+]i rises. Eugenol killed cells which were not reversed by prechelating cytosolic Ca2+ with 1,2-bis(2-aminophenoxy) ethane-N,N,N′,N′-tetraacetic acid-acetoxymethyl ester (BAPTA-AM). Eugenol induced apoptosis through increasing reactive oxygen species (ROS) production, decreasing mitochondrial membrane potential, releasing cytochrome c and activating caspase-9/caspase-3. Together, in DBTRG-05MG cells, eugenol evoked [Ca2+]i rises by inducing PLC-dependent release of Ca2+ from the endoplasmic reticulum and caused Ca2+ influx possibly through TRPM8 or PKC-sensitive channels. Furthermore, eugenol induced the mitochondrial apoptotic pathway.