Signal transduction of epidermal growth factor on the production of nitric oxide in rat intestinal epithelial cells

表皮生长因子(EGF)是小肠移植(SBT)后肠适应的营养因子之一。最近的研究表明，一氧化氮(NO)对SBT后细菌易位(BT)具有细胞保护作用。我们假设表皮生长因子刺激移植后移植物中诱导型一氧化氮合酶(INOS)基因的表达，继而增加NO的产生，导致BT减少。在有或没有磷脂酰肌醇3-激酶(PI3K)和表皮生长因子受体激酶抑制剂(LY294002和Tyrphostin A25)的情况下，用表皮生长因子或/和白细胞介素1β(IL-1β)处理肠上皮细胞。分析了诱导型一氧化氮合酶及其信号分子，包括核因子-κB抑制蛋白(I-κB)、核转录因子-κB(NF-κB)和AKT的诱导作用。IL-1β可促进I-κB的降解和核因子-κB的活化，但对诱导型一氧化氮合酶的诱导作用不明显。表皮生长因子对NF-κB的激活和iNOS的诱导无影响，但可通过PI3K/AKT刺激I型白细胞介素1受体(IL-1R1)上调。结果表明，表皮生长因子和IL-1β分别通过PI3K/AKT上调IL-1R1和通过I-κB激酶激活NF-κb是诱导iNOS产生的两个重要信号。同时加入表皮生长因子和IL-1β可促进NO的产生，这可能是表皮生长因子对肠道损伤的细胞保护作用之一。

Epidermal growth factor (EGF) is one of trophic factors for intestinal adaptation after small bowel transplantation (SBT). Recent report indicates that nitric oxide (NO) has cytoprotective effect on bacterial translocation (BT) after SBT. We hypothesized that EGF stimulates the expression of inducible nitric oxide synthase (iNOS) gene in the graft after SBT, followed by increased production of NO, resulting in the decrease of BT.Intestinal epithelial cells (IEC-6) were treated with EGF or/and interleukin-1β (IL-1β) in the presence and absence of phosphatidylinositol 3-kinase (PI3K) and EGF receptor kinase inhibitors (LY294002 and tyrphostin A25). The induction of NO production and iNOS, and its signal molecules including the inhibitory protein of NF-κB (IκB), nuclear factor-κB (NF-κB) and Akt were analyzed.IL-1β stimulated the degradation of IκB and the activation of NF-κB, but had no effect on iNOS induction. EGF, which had no effect on the NF-κB activation and iNos induction, stimulated the up-regulation of type 1 interleukin-1 receptor (IL-1R1) through PI3K/Akt. Simultaneous addition of EGF and IL-1b stimulated synergistically the induction of iNOS, leading to the increased production of NO.Our results indicate that EGF and IL-1β stimulate two essential signals for iNOS induction in IEC-6 cells, the up-regulation of IL-1R1 through PI3K/Akt and the activation of NF-κb through IκB kinase, respectively. Simultaneous addition of EGF and IL-1β can enhance the production of NO, which may contribute to the cytoprotective effect of EGF against intestinal injury.