Regulation of intestinal tyrosine phosphorylation and programmed cell death by peroxovanadate.
Regulation of intestinal tyrosine phosphorylation and programmed cell death by peroxovanadate.
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过氧钒酸盐调节肠道酪氨酸磷酸化和程序性细胞死亡。
DOI：
10.1152/ajpcell.1999.277.3.c572
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发表时间：
1999
期刊：
影响因子：
--
通讯作者：
Zhang,L
中科院分区：
文献类型：
--
作者：
Scheving,LA;Thomas,JR;Zhang,L
当在37°C下在简单培养基中培养时，回肠粘膜的细胞悬浮液经历快速和同步形式的程序性细胞死亡。由于蛋白质的酪氨酸磷酸化在许多细胞过程的信号转导中起着至关重要的作用，我们通过使用免疫印迹和免疫组织化学方法研究了其在肠道程序性细胞死亡中的作用。我们观察到在肠上皮细胞培养的最初10分钟内酪氨酸磷酸化减少了50-70%。我们假设蛋白酪氨酸磷酸酶的抑制将增加这些悬浮液中的蛋白酪氨酸磷酸化，并减少程序性细胞死亡。这些磷酸酶的强抑制剂（过氧钒酸盐），但不是一个较弱的（原钒酸钠）取消了DNA片段化/梯状通常看到在垂死的肠细胞。过氧钒酸盐增强了许多肠道蛋白的蛋白酪氨酸磷酸化，显著增加了丝裂原活化蛋白激酶的双重磷酸化和活性形式。免疫组化显示，在过氧钒酸盐处理的粘膜肠隐窝中，酪氨酸磷酸化水平特别高。动力学研究表明，蛋白酪氨酸磷酸酶抑制的关键时间发生在离体培养的5分钟内，正是当蛋白酪氨酸磷酸化下降。我们的数据表明，酪氨酸激酶失活或酪氨酸磷酸酶激活可能会引发肠上皮细胞死亡。
Cell suspensions of ileal mucosa undergo a rapid and synchronized form of programmed cell death when cultured in a simple medium at 37°C. Because tyrosine phosphorylation of proteins plays a crucial role in the signal transduction of many cellular processes, we examined its role in intestinal programmed cell death by use of immunoblot and immunohistochemical methods. We observed a 50–70% reduction in tyrosine phosphorylation during the initial 10 min of intestinal epithelial cell culture. We hypothesized that the inhibition of protein tyrosine phosphatases would increase protein tyrosine phosphorylation in these suspensions and decrease programmed cell death. A strong inhibitor of these phosphatases (peroxovanadate) but not a weaker one (sodium orthovanadate) abolished the DNA fragmentation/laddering normally seen in dying enterocytes. Peroxovanadate enhanced protein tyrosine phosphorylation of many intestinal proteins, dramatically increasing the dually phosphorylated and active form of mitogen-activated protein kinase. Immunohistochemistry revealed a particularly high level of increased tyrosine phosphorylation in the intestinal crypts in peroxovanadate-treated mucosa. Kinetic studies indicated that the pivotal time for protein tyrosine phosphatase inhibition occurred within 5 min of ex vivo culture, precisely when protein tyrosine phosphorylation declined. Our data suggest that tyrosine kinase inactivation or tyrosine phosphatase activation may initiate intestinal epithelial cell death.