The effect of gallic acid on cytotoxicity, Ca2+ homeostasis and ROS production in DBTRG-05MG human glioblastoma cells and CTX TNA2 rat astrocytes

Abstract

Gallic acid, a polyhydroxylphenolic compound, is widely distributed in various plants, fruits and foods. It has been shown that gallic acid passes into blood brain barrier and reaches the brain tissue of middle cerebral artery occlusion rats. However, the effect of gallic acid on Ca2+ signaling in glia cells is unknown. This study explored whether gallic acid affected Ca2+ homeostasis and induced Ca2+-associated cytotoxicity in DBTRG-05MG human glioblastoma cells and CTX TNA2 rat astrocytes. Gallic acid (20–40 μM) concentration-dependently induced cytotoxicity and intracellular Ca2+ level ([Ca2+]i) increases in DBTRG-05MG cells but not in CTX TNA2 cells. In DBTRG-05MG cells, the Ca2+ response was decreased by half by removal of extracellular Ca2+. In Ca2+-containing medium, gallic acid-induced Ca2+ entry was inhibited by store-operated Ca2+ channel inhibitors (2-APB, econazole and SKF96365). In Ca2+-free medium, pretreatment with the endoplasmic reticulum Ca2+ pump inhibitor thapsigargin abolished gallic acid-induced [Ca2+]i increases. Conversely, incubation with gallic acid also abolished thapsigargin-induced [Ca2+]i increases. Inhibition of phospholipase C with U73122 abolished gallic acid-induced [Ca2+]i increases. Gallic acid significantly caused cytotoxicity in DBTRG-05MG cells, which was partially prevented by prechelating cytosolic Ca2+ with BAPTA-AM. Moreover, gallic acid activated mitochondrial apoptotic pathways that involved ROS production. Together, in DBTRG-05MG cells but not in CTX TNA2 cells, gallic acid induced [Ca2+]i increases by causing Ca2+ entry via 2-APB, econazole and SKF96365-sensitive store-operated Ca2+ entry, and phospholipase C-dependent release from the endoplasmic reticulum. This Ca2+ signal subsequently evoked mitochondrial pathways of apoptosis that involved ROS production.