Caffeine causes glycerophosphorylcholine accumulation through ryanodine-inhibitable increase of cellular calcium and activation of phospholipase A2 in cultured MDCK cells
Caffeine causes glycerophosphorylcholine accumulation through ryanodine-inhibitable increase of cellular calcium and activation of phospholipase A2 in cultured MDCK cells
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DOI：
10.1038/emm.1998.22
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发表时间：
1998-09-30
影响因子：
12.8
通讯作者：
Jung, KY
中科院分区：
文献类型：
--
作者：
Kim, DK;Jung, KY
甘油磷胆碱 (GPC) 是一种肾髓相容性有机渗透剂，通过磷脂酰胆碱衍生自胆碱，部分由磷脂酶 A(2) (PLA(2)) 催化，并被 GPC:胆碱磷酸二酯酶 (GPC:胆碱 PDE) 降解。我们发现咖啡因升高了培养的 MDCK 细胞、犬肾上皮细胞中的细胞内游离钙 ([Ca2+](i)) 和 GPC 水平，并提出了可能的生化机制。当 MDCK 细胞与 1 至 10 mM 咖啡因一起孵育 3 小时时，细胞 GPC 以剂量依赖性方式升高，并且这种情况的发生与细胞外渗透压无关。咖啡因刺激 [C-14] 胆碱掺入 [C-14]GPC 和 PLA2 活性的速率。而GPC：胆碱PDE活性则伴随着较小的增加。这些酶的变化表明 MDCK GPC 的净合成增加。为了确定触发 PLA2 激活的因素，使用荧光染料 Fura-P 测量了 [Ca2+](i)。咖啡因 (10 mM) 会导致 MDCK [Ca2+](i) 浓度出现典型的短暂增加，并且用 10 mM ryanodine 预处理 MDCK 细胞 5 分钟可大大抑制这种增加。 Ryanodine (10 mM) 还可以抑制咖啡因诱导的 PLA2 活性刺激。这些发现提供了第一个证据，证明 MDCK 细胞中的咖啡因会导致兰尼碱抑制的 [Ca2+](i) 和 PLA2 活性增加，从而导致细胞 GPC 积累。
Glycerophosphrylocholine (GPC) is a renal medullary compatible organic osmolyte that is derived from choline via phosphatidylcholine, which is catalyzed in part by phospholipase A(2) (PLA(2)) and its degradation by GPC:choline phosphodiesterase (GPC:choline PDE). We found that caffeine elevated intracellular free calcium ([Ca2+](i)) and GPC level in cultured MDCK cells, canine kidney epithelial cells, and propose a possible biochemical mechanism. When MDCK cells were incubated for 3 h with 1 to 10 mM caffeine, cellular GPC was elevated in a dose-dependent manner, and this occurred independently of the extracellular osmolality. Caffeine stimulated the rate of [C-14]choline incorporation into [C-14]GPC and PLA2 activity. Whereas, GPC:choline PDE activity was accompanied by less of increase. These enzyme changes demonstrate the increased net synthesis of MDCK GPC. In order to identify what triggers the PLA2 activation, [Ca2+](i) was measured by using a fluorescence dye, Fura-P. Caffeine (10 mM) resulted in a typical transient increase in MDCK [Ca2+](i) concentration, and this increase was greatly inhibited by pretreatment of MDCK cells with 10 mM ryanodine for 5 min. Ryanodine (10 mM) also inhibited the caffeine-induced stimulation of PLA2 activity. These findings provide the first evidence that caffeine in MDCK cells causes a ryanodine-inhibitable increase of [Ca2+](i) and PLA2 activity, resulting in cellular GPC accumulation.