Department of Surgery, Kaohsiung Veterans General Hospital, Kaohsiung, Taiwan.
Hum Exp Toxicol. 2012 Dec;31(12):1251-61. doi: 10.1177/0960327112446841. Epub 2012 May 31.
The effect of the natural compound phenethyl isothiocyanate (PEITC) on cytosolic Ca(2+) concentrations (Ca(2+)) and viability in MDCK renal cells is unknown. This study explored whether PEITC changed Ca(2+) in MDCK cells using the Ca(2+)-sensitive fluorescent dye fura-2. PEITC at 200-700 μM increased Ca(2+) in a concentration-dependent manner. The signal was reduced by removing extracellular Ca(2+). PEITC-induced Ca(2+) influx was inhibited by nifedipine, econazole, SK&F 96365 and protein kinase C modulators. In Ca(2+)-free medium, treatment with the endoplasmic reticulum Ca(2+) pump inhibitor thapsigargin (TG) or 2,5-di-tert-butylhydroquinone (BHQ) inhibited PEITC-induced rise in Ca(2+). Incubation with PEITC also inhibited TG or BHQ-induced rise in Ca(2+). Inhibition of phospholipase C with U73122 abolished PEITC-induced rise in Ca(2+). At 15-75 μM, PEITC decreased viability. The cytotoxic effect of PEITC was enhanced by chelating cytosolic Ca(2+) with 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid/acetoxymethyl ester. Annexin V-FITC data suggest that 20 and 50 μM PEITC induced apoptosis. At 10 and 15 μM, PEITC did not increase reactive oxygen species (ROS) production. Together, in renal tubular cells, PEITC-induced rise in Ca(2+) by inducing phospholipase C-dependent Ca(2+) release from endoplasmic reticulum and Ca(2+) entry via store-operated Ca(2+) channels. PEITC induced apoptosis in a concentration-dependent, ROS/Ca(2+)-independent manner.
天然化合物苯乙基异硫氰酸酯（PEITC）对肾细胞溶质 Ca(2+)浓度 (Ca(2+)) 和活力的影响尚不清楚。本研究使用 Ca(2+) 敏感荧光染料 fura-2 研究了 PEITC 是否改变了 MDCK 细胞中的 Ca(2+)。PEITC 在 200-700μM 浓度范围内以浓度依赖性方式增加 Ca(2+)。去除细胞外 Ca(2+)可减少信号。硝苯地平、依康唑、SK&F 96365 和蛋白激酶 C 调节剂抑制 PEITC 诱导的 Ca(2+) 内流。在无 Ca(2+) 培养基中，内质网 Ca(2+) 泵抑制剂 thapsigargin (TG) 或 2,5-二叔丁基对苯二酚 (BHQ) 处理抑制了 PEITC 诱导的 Ca(2+) 升高。用 PEITC 孵育也抑制了 TG 或 BHQ 诱导的 Ca(2+) 升高。用 U73122 抑制磷脂酶 C 消除了 PEITC 诱导的 Ca(2+) 升高。在 15-75μM 时，PEITC 降低了细胞活力。用 1,2-双(2-氨基苯氧基)乙烷-N,N,N',N'-四乙酸/乙氧羰基甲酯螯合细胞质 Ca(2+) 增强了 PEITC 的细胞毒性作用。Annexin V-FITC 数据表明，20 和 50μM 的 PEITC 诱导了细胞凋亡。在 10 和 15μM 时，PEITC 并未增加活性氧物质 (ROS) 的产生。综上所述，在肾小管细胞中，PEITC 通过诱导内质网中依赖于磷脂酶 C 的 Ca(2+) 释放和通过储存操作的 Ca(2+) 通道的 Ca(2+) 内流来诱导 Ca(2+) 的增加。PEITC 以浓度依赖的方式、ROS/Ca(2+) 非依赖性方式诱导细胞凋亡。
