The mechanism of safrole-induced [Ca ²⁺ ]i rises and Non-Ca ²⁺ -triggered cell death in SCM1 human gastric cancer cells

Safrole is a carcinogen found in plants. The effect of safrole on cytosolic free Ca ²⁺ concentrations ([Ca ²⁺ ]i) and viability in SCM1 human gastric cancer cells was explored. The Ca ²⁺ -sensitive fluorescent dye fura-2 was applied to measure [Ca ²⁺ ]i. Safrole at concentrations of 150-450 μM induced a [Ca ²⁺ ]i rise in a concentration-dependent manner. The response was reduced by 60% by removing extracellular Ca ²⁺ . Safrole-evoked Ca ²⁺ entry was not altered by nifedipine, econazole, SKF96365, and protein kinase C activator or inhibitor. In Ca ²⁺ -free medium, treatment with the endoplasmic reticulum Ca ²⁺ pump inhibitor thapsigargin or 2,5-di-tert-butylhydroquinone (BHQ) abolished safrole-evoked [Ca ²⁺ ]i rises. Conversely, treatment with safrole abolished thapsigargin or BHQ-evoked [Ca ²⁺ ]i rises. Inhibition of phospholipase C (PLC) with U73122 abolished safrole-induced [Ca ²⁺ ]i rises. At 250-550 μM, safrole decreased cell viability concentration-dependently, which was not reversed by chelating cytosolic Ca ²⁺ with 1,2-bis(2-aminophenoxy)ethane-N,N,N’,N’-tetraacetic acid/acetoxy methyl (BAPTA/AM). Annexin V/propidium iodide staining data suggest that safrole (350-550 μM) induced apoptosis concentration-dependently. These studies suggest that in SCM1 human gastric cancer cells, safrole induced [Ca ²⁺ ]i rises by inducing PLC-dependent Ca ²⁺ release from the endoplasmic reticulum and Ca ²⁺ influx via non-store-operated Ca ²⁺ entry pathways. Safrole-induced cell death may involve apoptosis.