Regulation of intestinal tyrosine phosphorylation and programmed cell death by peroxovanadate

Regulation of intestinal tyrosine phosphorylation and programmed cell death by peroxovanadate. Am. J. Physiol. 277 (Cell Physiol. 46): C572-C579, 1999 .-Cell suspensions of ileal mucosa undergo a rapid and synchronized form of programmed cell death when cultured in a simple medium at 37 degrees C. Be cause tyrosine phosphorylation of proteins plays a crucial role in the sign al transduction of many cellular processes, we examined its role in intesti nal programmed cell death by use of immunoblot and immunohistochemical meth ods. 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 tyro sine phosphorylation in these suspensions and decrease programmed cell deat h. A strong inhibitor of these phosphatases (peroxovanadate) but not a weak er one (sodium orthovanadate) abolished the DNA fragmentation/laddering nor mally seen in dying enterocytes. Peroxovanadate enhanced protein tyrosine p hosphorylation of many intestinal proteins, dramatically increasing the dua lly phosphorylated and active form of mitogen-activated protein kinase. Imm unohistochemistry 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 phosph atase inhibition occurred within 5 min of ex vivo culture, precisely when p rotein tyrosine phosphorylation declined. Our data suggest that tyrosine ki nase inactivation or tyrosine phosphatase activation may initiate intestina l epithelial cell death.