Effects of puerarin on intracellular Ca²⁺ and cell viability of MDCK renal tubular cells

Puerarin is a natural compound and has been used as herb medication in a number of countries, especially in Asia. The effect of puerarin on Ca²⁺ signaling is unknown in renal cells. This study examined whether puerarin affected Ca²⁺ physiology in MDCK renal tubular cells. Cytosolic free Ca²⁺ levels ([Ca²⁺]_i) were measured using the fluorescent dye fura-2. Cell viability was examined by using WST-1 assay. Puerarin induced [Ca²⁺]_i rises and the response was reduced by removing extracellular Ca²⁺. Puerarin-induced Ca²⁺ entry was not altered by protein kinase C (PKC) activity, but was inhibited by nifedipine. In Ca²⁺-free medium, treatment with the endoplasmic reticulum Ca²⁺ pump inhibitor 2,5-di-tert-butylhydroquinone (BHQ) or thapsigargin partly inhibited puerarin-evoked [Ca²⁺]_i rises. Inhibition of phospholipase C (PLC) with U73122 did not change puerarin-induced [Ca²⁺]_i rises. Puerarin at 25–50 μM caused cytotoxicity, which was not reversed by pretreatment with the Ca²⁺ chelator 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid-acetoxymethyl ester (BAPTA/AM). Collectively, in MDCK cells, puerarin induced [Ca²⁺]_i rises by evoking PLC-independent Ca²⁺ release from the endoplasmic reticulum and other unknown stores, and Ca²⁺ entry via nifedipine-sensitive, PKC-insensitive Ca²⁺ entry pathways. Puerarin also caused Ca²⁺-independent cell death.