Mechanism of EGF-Mediated Reduction of NEC

描述（由申请人提供）：坏死性小肠结肠炎（NEC）是早产儿发病和死亡的主要原因，但病因不明，也没有有效的治疗方法。利用大鼠NEC模型，我们报道了表皮生长因子（EGF）显著降低NEC的发病率和严重程度。本提案的中心目标是阐明egf介导的NEC减少的分子机制。NEC患者肠道细胞凋亡加速，EGF可保护细胞免于凋亡。我们的初步研究表明，EGF治疗NEC可调节NEC损伤部位的促存活信号分子。特异性目标1将通过评估EGF对NEC新生大鼠回肠中促生存和促凋亡基因和蛋白质的治疗(a), (b)对wave -2小鼠（非致病性情况下异常的EGF- r信号传导）的治疗，(c)对关键促生存和促凋亡基因过度表达或被阻断的肠细胞系的治疗，以及(d)对给予EGF- r信号传导抑制剂的新生大鼠的治疗，阐明哪些促生存途径受到EGF治疗NEC的调节。最近，我们发现肠肝循环在NEC发病机制中起着至关重要的作用。我们的初步数据表明，对NEC新生大鼠给予EGF可下调管腔胆汁酸（BA）水平并调节BA转运蛋白，提示EGF在BA稳态中起重要作用。在Specific Aim 2中，我们将通过评估EGF介导的(a) NEC新生大鼠中BA运输的变化，(b) BA运输的重要成分过度表达或阻断的IEC系，以及(c) wave -2小鼠中EGF调节NEC期间肠内BA运输的机制。肝素结合egf样生长因子（HB-EGF）是egf相关肽家族的一员。我们的初步结果表明，HB-EGF降低了实验性NEC的发生率，并增强了egf介导的NEC的减少。因此，HB-EGF与EGF可能通过独特的非拮抗机制对NEC提供更好的保护。特异性目标3将通过(a)确定减少新生大鼠NEC的最佳途径和剂量，(b)确定EGF和HB-EGF减少NEC的最佳组合，以及(c)利用体外肠损伤模型评估HB-EGF介导的NEC减少机制来研究HB-EGF介导的NEC减少机制。这些研究将阐明EGF介导的NEC减少的机制，这些信息对于开始使用EGF作为人类NEC的预防或治疗方式至关重要。

DESCRIPTION (provided by applicant): Necrotizing enterocolitis (NEC) is a major cause of morbidity and mortality in premature infants but the cause is unknown and there are no effective treatments. Using the rat model of NEC, we reported that epidermal growth factor (EGF) markedly reduced the incidence and severity of NEC. The central goal of this proposal is to clarify the molecular mechanism(s) of EGF-mediated reduction of NEC. Accelerated intestinal apoptosis has been shown in patients with NEC, and EGF can protect cells from apoptosis. Our preliminary studies indicate that EGF treatment of NEC regulates pro-survival signaling molecules in the site of NEC injury. Specific Aim 1 will clarify which pro-survival pathways are regulated by EGF treatment of NEC by evaluating EGF treatment (a) on pro-survival and pro-apoptotic genes and proteins in the ileum of neonatal rats with NEC, (b) in waved-2 mice (aberrant EGF-R signaling under non-pathogenic circumstances), (c) in intestinal cell lines where crucial pro-survival and pro-apoptotic genes are over-expressed or blocked and (d) in neonatal rats given EGF-R signaling inhibitors. Recently, we have shown a crucial role of enterohepatic circulation in NEC pathogenesis. Our preliminary data indicate that administration of EGF to neonatal rats with NEC down-regulates luminal bile acid (BA) levels and regulates BA transporters, suggesting an important role of EGF in BA homeostasis. In Specific Aim 2, we will explore the mechanism by which EGF regulates BA transport in the intestine during NEC by evaluating EGF-mediated changes in (a) BA transport in neonatal rats with NEC, (b) IEC lines where essential components of BA transport are over-expressed or blocked, and (c) in waved-2 mice. Heparin-binding EGF-like growth factor (HB-EGF) is a member of the family of EGF-related peptides. Our preliminary results show that HB-EGF reduces the incidence of experimental NEC and enhances EGF-mediated reduction of NEC. Thus, HB-EGF with EGF may provide better protection against NEC through unique nonantagonistic mechanisms. Specific Aim 3 will examine the mechanisms of HB-EGF-mediated reduction of NEC by (a) identifying the optimal route and dose to reduce NEC in neonatal rats, (b) determining the optimal combination of EGF and HB-EGF to reduce NEC, and (c) utilizing an in vitro intestinal injury model to evaluate mechanisms of HB-EGF-mediated reduction of NEC. These studies will elucidate the mechanisms of EGF-mediated reduction of NEC, information that is essential to initiate use of EGF as a preventative or treatment modality for human NEC.