The β1 isoform of protein kinase C mediates the protective effects of epidermal growth factor on the dynamic assembly of F-actin cytoskeleton and normalization of calcium homeostasis in human colonic cells

被引:24
作者
Banan, A
Fields, JZ
Farhadi, A
Talmage, DA
Zhang, L
Keshavarzian, A
机构
[1] Rush Univ, Med Ctr, Div Digest Dis, Dept Internal Med,Sect Gastroenterol & Nutr, Chicago, IL 60612 USA
[2] Rush Univ, Med Ctr, Dept Pharmacol, Chicago, IL 60612 USA
[3] Rush Univ, Med Ctr, Dept Mol Physiol, Chicago, IL 60612 USA
[4] Columbia Univ, Inst Human Nutr, New York, NY 10032 USA
关键词
D O I
10.1124/jpet.301.3.852
中图分类号
R9 [药学];
学科分类号
1007 ;
摘要
Using intestinal monolayers, we showed that F-actin cytoskeletal stabilization and Ca2+ normalization contribute to epidermal growth factor (EGF)-mediated protection against oxidant injury. However, the intracellular mediator responsible for these protective effects remains unknown. Since the protein kinase C-beta1 (PKC-beta1) isoform is abundant in our naive (N) cells, we hypothesized that PKC-beta1 is essential to EGF protection. Monolayers of N Caco-2 cells were exposed to H2O2+/-EGF, PKC, or Ca2+ modulators. Other cells were transfected to over-express PKC-beta1 or to inhibit its expression and then pretreated with low or high doses of EGF or a PKC activator, OAG (1-oleoyl-2-acetyl-sn-glycerol), before H2O2. In N monolayers exposed to oxidant, pretreatment with EGF or PKC activators activated PKC-beta1, enhanced Ca-45(2+) efflux, normalized Ca2+, decreased monomeric G-actin, increased stable F-actin, and protected the cytoarchitecture of the actin. PKC inhibitors prevented these protective effects. Transfected cells stably overexpressing PKC-beta1 (+3.1-fold) but not N cell monolayers were protected from injury by even lower doses of EGF or OAG. EGF or OAG rapidly activated the over-expressed PKC-beta1. Antisense inhibition of PKC-beta1 expression (-90%) prevented all measures of EGF protection. Inhibitors of Ca2+-ATPase prevented EGF protection in N cells as well as protective synergism in transfected cells. EGF protects the assembly of the F-actin cytoskeleton in intestinal monolayers against oxidants in large part through the activation of PKC-beta1. EGF normalizes Ca2+ by enhancing Ca2+ efflux through PKC-beta1. We have identified novel biologic functions, protection of actin and Ca2+ homeostasis, among the classical isoforms of PKC.
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收藏
页码:852 / 866
页数:15
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