The Mechanism of Ca(2+) Movement in the Involvement of Baicalein-Induced Cytotoxicity in ZR-75-1 Human Breast Cancer Cells

Baicalein (5,6,7-trihydroxyflavone) (1) has been found to be active against a wide variety of Cancer cells. However, the molecular mechanism underlying the effects of 1 on the induction of CA(2+) movement and cytotoxicity in human breast Cancer cells is unknown. This study examined the relationship between 1-induced CA(2+) signaling and cytotoxicity in ZR-75-1 human breast Cancer cells. The in vitro investigations reported herein produced the following results: (i) Compound 1 increased intracellular CA(2+) concentration ([CA(2+)]i) in a concentration-dependent manner. The signal was decreased by approximately 50% by removal of extracellular CA(2+). (ii) Compound 1-triggered [CA(2+)]i increases were significantly suppressed by store-operated CA(2+) channel blockers 2-aminoethoxydiphenyl borate (2-APB) and the PKC Inhibitor GF109203X. (iii) In CA(2+)-free medium, compound 1-induced [CA(2+)]i increases were also inhibited by GF109203X. Furthermore, pretreatment with the endoplasmic reticulum CA(2+) pump inhibitor thapsigargin (TG) or 2,5-ditert-butylhydroquinone (BHQ) abolished 1-induced [CA(2+)]i increases. Inhibition of Phospholipase C (PLC) with U73122 abolished 1-induced [CA(2+)]i increases. (iv) Compound 1 (20-40 μM) caused cytotoxicity, increased Reactive Oxygen Species (ROS) production, and activated caspase-9/Caspase-3. Furthermore, compound 1-induced Apoptosis was significantly inhibited by prechelating cytosolic CA(2+) with BAPTA-AM (1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid acetoxymethyl ester) or by decreasing ROS with the antioxidant NAC (N-acetylcysteine). Together, baicalein (1) induced a [CA(2+)]i increase by inducing PLC-dependent CA(2+) release from the endoplasmic reticulum and CA(2+) entry via PKC-dependent, 2-APB-sensitive store-operated CA(2+) channels. Moreover, baicalein (1) induced CA(2+)-associated Apoptosis involved ROS production in ZR-75-1 cells.