The Carcinogen Safrole Increases Intracellular Free Ca(superscript 2+) Levels and Causes Death in MDCK Cells

Wei-Chuan Chen ； He-Hsiung Cheng ； Chun-Jen Huang ； Yih-Chau Lu ； I-Shu Chen ； Shiuh-Inn Liu ； Shu-Shong Hsu ； Hong-Tai Chang ； Jong-Khing Huang ； Jin-Shyr Chen ； Chung-Ren Jan

《The Chinese Journal of Physiology》 50卷1期 (2007/02) Pp. 34-40

並列摘要

The effect of the carcinogen safrole on intracellular Ca(superscript 2+) movement in renal tubular cells has not been explored previously. The present study examined whether safrole could alter Ca(superscript 2+) handling in Madin-Darby canine kidney (MDCK) cells. Cytosolic free Ca(superscript 2+) levels ([Ca(superscript 2+)](subscript i)) in populations of cells were measured using fura-2 as a fluorescent Ca(superscript 2+) probe. Safrole at concentrations above 33 μM increased [Ca(superscript 2+)](subscript i) in a concentration-dependent manner with an EC50 value of 400 μM. The Ca(superscript 2+) signal was reduced by 90% by removing extracellular Ca(superscript 2+), but was not affected by nifedipine, verapamil, or diltiazem. Addition of Ca(superscript 2+) after safrole had depleted intracellular Ca(superscript 2+)-induced dramatic Ca(superscript 2+) influx, suggesting that safrole caused store-operated Ca(superscript 2+) entry. In Ca(superscript 2+)-free medium, after pretreatment with 650 μM safrole, 1 μM thapsigargin (an endoplasmic reticulum Ca(superscript 2+) pump inhibitor) failed to release more Ca(superscript 2+). Inhibition of phospholipase C with 2 μM U73122 did not affect safrole-induced Ca(superscript 2+) release. Trypan blue exclusion assays revealed that incubation with 650 μM safrole for 30 min did not kill cells, but killed 70% of cells after incubation for 60 min. Collectively, the data suggest that in MDCK cells, safrole induced a [Ca(superscript 2+)](subscript i) increase by causing Ca(superscript 2+) release from the endoplasmic reticulum in a phospholipase C-independent fashion, and by inducing Ca(superscript 2+) influx via store-operated Ca(superscript 2+) entry. Furthermore, safrole can cause acute toxicity to MDCK cells.

並列關鍵字

Ca(superscript 2+) ； Ca(superscript 2+) stores ； fura-2 ； MDCK ； renal tubular cells ； safrole

參考文獻

Alvarez-Barrientos, A.,O`Connor, J.E.,Nieto Castillo, R.,Moreno Moreno, A.B.,Prieto, P.(2001).Use of flow cytometry and confocal microscopy techniques to investigate early CdCl2-induced nephrotoxicity in vitro.Toxicol. In Vitro.15,407-412.

Benedetti, M.S.,Malnoe, A.,Broillet, A.L.(1977).Absorption, metabolism and excretion of safrole in the rat and man.Toxicology.7,69-83.

Bolton, J.L.,Acay, N.M.,Vukomanovic, V.(1994).Evidence that 4-allyl-o-quinones spontaneously rearrange to their more electrophilic quinone methides: potential bioactivation mechanism for the hepatocarcinogen safrole.Chem. Res. Toxicol..7,443-450.

Chang, M.J.,Ko, C.Y.,Lin, R.F.,Hsieh, L.L.(2002).Biological monitoring of environment exposure to safrole and the Taiwanese betel quid chewing.Arch. Environ. Contam. Toxicol..43,432-437.

Chen, C.L.,Chi, C.W.,Chang, K.W.,Liu, T.Y.(1999).Safrole-like DNA adducts in oral tissue from oral cancer patients with a betel quid chewing history.Carcinogenesis.20,2331-2334.