Role for tyrosine kinases in carbachol-regulated Ca entry into colonic epithelial cells.
Role for tyrosine kinases in carbachol-regulated Ca entry into colonic epithelial cells.
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酪氨酸激酶在卡巴胆碱调节的 Ca 进入结肠上皮细胞中的作用。
DOI：
10.1152/ajpcell.1995.268.1.c154
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发表时间：
1995
期刊：
影响因子：
--
通讯作者：
Machen,TE
中科院分区：
文献类型：
--
作者：
Bischof,G;Illek,B;Reenstra,WW;Machen,TE
我们研究了酪氨酸激酶在调节钙进入结肠上皮细胞HT-29/B6使用数字图像处理的Fura 2荧光的可能作用。卡巴胆碱和毒胡萝卜素增加钙进入到一个类似的程度和钙内流减少酪氨酸激酶抑制剂染料木素（50 μ M）。进一步的实验进行了在含有95 mM K的溶液中，以降低膜电位，和不同的抑制剂对钙，锰，钡流入的影响进行了比较。染料木黄酮，而不是非活性类似物大豆苷元，也不是蛋白激酶C抑制剂1-（5-异喹啉磺酰基）-2-甲基哌嗪，减少进入所有三个二价阳离子的47- 59%。在高钾溶液中，卡巴胆碱或毒胡萝卜素都导致细胞内钙增加到223 +/- 19 nM的平台。酪氨酸激酶抑制剂染料木黄酮（至95 +/- 8 nM）、乳木黄酮A（至155 +/- 17 nM）和2，5-二羟基肉桂酸甲酯（至39 +/- 3 nM）降低了该平台。原钒酸盐，一种蛋白酪氨酸磷酸酶抑制剂，阻止染料木素的抑制作用。钙泵不受染料木素。卡巴胆碱增加110-，75-和70-kDa蛋白质的酪氨酸磷酸化（免疫印迹与抗磷酸酪氨酸抗体），这种磷酸化被抑制染料木素。我们的结论是卡巴胆碱和毒胡萝卜素增加钙的进入，和酪氨酸磷酸化的一些关键蛋白可能是重要的调节这一途径。
We studied a possible role of tyrosine kinases in the regulation of Ca entry into colonic epithelial cells HT-29/B6 using digital image processing of fura 2 fluorescence. Both carbachol and thapsigargin increased Ca entry to a similar extent and Ca influx was reduced by the tyrosine kinase inhibitor genistein (50 microM). Further experiments were performed in solutions containing 95 mM K to depolarize the membrane potential, and the effects of different inhibitors on influx of Ca, Mn, and Ba were compared. Genistein, but not the inactive analogue daidzein nor the protein kinase C inhibitor 1-(5-isoquinolinylsulfonyl)-2- methylpiperazine, decreased entry of all three divalent cations by 47-59%. In high-K solutions, carbachol or thapsigargin both caused intracellular Ca to increase to a plateau of 223 +/- 19 nM. This plateau was reduced by the tyrosine kinase inhibitors genistein (to 95 +/- 8 nM), lavendustin A (to 155 +/- 17 nM), and methyl-2,5-dihydroxycinnamate (to 39 +/- 3 nM). Orthovanadate, a protein tyrosine phosphatase inhibitor, prevented the inhibitory effect of genistein. Ca pumping was unaffected by genistein. Carbachol increased tyrosine phosphorylation (immunoblots with anti-phosphotyrosine antibodies) of 110-, 75-, and 70-kDa proteins, and this phosphorylation was inhibited by genistein. We conclude that carbachol and thapsigargin increase Ca entry, and tyrosine phosphorylation of some key proteins may be important for regulating this pathway.