Immature intestinal NF-kB regulation, probiotics, and necrotizing enterocolitis

描述（由申请人提供）：坏死性小肠结肠炎（NEC）是一种病因不明、危及生命的炎症性肠病，约10%的出生体重<1500克的早产儿会受到影响。早产是最大的危险因素，而不是任何特定的侮辱，这表明肠道不成熟是一个根本问题。然而，不成熟对NEC的贡献的确切方面却知之甚少。我发现未成熟和成熟肠上皮细胞（IEC）发育差异的一个关键点是核因子kappaB的调控。针对RFA-HD-07-08“预防和治疗坏死性小肠结肠炎的新方法”，我建议验证肠上皮细胞NF-kB调节的内在不成熟导致夸大的炎症反应，使早产儿易患NEC的假设。此外，我推测益生菌分泌的可溶性因子可以调节NF-kB的调节，保持肠道屏障功能，从而减少炎症，预防NEC。本提案的初步数据表明，与成熟的IEC相比，未成熟的IEC在抑制性kappaB α和A20 -关键NF-kB下调蛋白的调控上存在差异。此外，我们的初步数据表明，在动物模型中，益生菌分泌的产物可以降低NF-kB活性并预防NEC。益生菌是一种细菌，它对健康的影响超出了其固有的营养价值。最近的临床试验表明，益生菌可能对NEC有一定的保护作用。这个建议的目的是…评估不成熟IEC中通过IkBa和A20下调NF-kB信号的可能机制。2. 在不引入活菌的动物模型中，确定植物乳杆菌、嗜酸乳杆菌和婴儿双歧杆菌分泌的细菌产物对坏死性小肠结肠炎的保护能力。实验设计采用从断奶前小鼠分离的未成熟IEC - IEC和人胎小肠细胞系H4两种模型。常规小鼠和无菌小鼠都将被使用。为了建立疾病模型，将使用已建立的NEC大鼠Caplan模型。这一建议将提供有关肠道不成熟方面的重要信息，这些信息有助于NEC，并与了解任何拟议治疗的效果有关。此外，了解无微生物益生菌条件培养基的潜在作用可能会提供一种方法来管理益生菌的有益作用而不危及活生物体，从而为治疗或预防高危婴儿NEC提供一种新的方法。公共卫生相关性：坏死性小肠结肠炎是一种知之甚少的、危及早产儿生命的炎症性肠病。尽管20-30%的患者死亡，幸存者面临严重的肠道和神经发育后果的风险，但目前尚无已知的特异性治疗方法。本研究将探讨肠道不成熟的哪些方面会导致这种疾病，并确定某些有益菌或益生菌分泌的因素是否会影响这些方面，从而降低易感婴儿对坏死性小肠结肠炎的易感性。

DESCRIPTION (provided by applicant): Necrotizing enterocolitis (NEC) is a life-threatening inflammatory bowel disorder of unknown cause that affects approximately 10% of premature infants born <1500gm. Prematurity is the greatest risk factor rather than any particular insult, suggesting that intestinal immaturity is a fundamental issue. However, the exact aspects of immaturity contributing to NEC are poorly understood. I have found that a key point of developmental difference between immature and mature intestinal epithelial cells (IEC) is regulation of nuclear factor kappaB. In response to RFA-HD-07-08 "New Approaches for the prevention and treatment of necrotizing enterocolitis," I propose to test the hypothesis that an intrinsic immaturity of intestinal epithelial cell NF-kB regulation leads to an exaggerated inflammatory response predisposing the preterm infant to NEC. Furthermore I hypothesize that soluble factors secreted by probiotic bacteria can modulate NF-kB regulation and preserve intestinal barrier function, thus decreasing inflammation and protecting against NEC. Preliminary data for this proposal demonstrate that compared to mature IEC, immature IEC have differences in regulation of both inhibitory kappaB alpha and A20 - key NF-kB down-regulatory proteins. Furthermore, our preliminary data demonstrate that secreted products from probiotic bacteria can decrease NF-kB activity and protect against NEC in an animal model. Probiotics are bacteria which have beneficial health effects beyond their inherent nutritive value. Recent clinical trials suggest that probiotics may confer some protection against NEC. This proposal is designed to 1. Assess possible mechanisms behind deficient down-regulation of NF-kB signaling via IkBa and A20 in immature IEC. 2. Determine the ability of secreted bacterial products from the probiotic organisms Lactobacillus plantarum, Lactobacillus acidophilus, and Bifidobacterium infantis to protect against necrotizing enterocolitis in an animal model without the introduction of live organisms. The experimental design uses two models of immature IEC - IEC isolated from pre-weaned mice and the human fetal small intestinal cell line H4. Both conventional and germ-free mice will be used. To model disease, the well established Caplan rat model of NEC will be used. This proposal will yield important information regarding aspects of intestinal immaturity which contribute to NEC and which are relevant to understanding the effect of any proposed treatment. Furthermore understanding the potential role of microbe free probiotic conditioned media may allow a means to administer the beneficial effects of probiotics without the risk of live organisms, thus providing a novel approach to treating or preventing NEC in at risk infants. PUBLIC HEALTH RELEVANCE: Necrotizing Enterocolitis is a poorly understood, life threatening inflammatory bowel disease of premature infants. Although 20-30% of patients die and survivors are at risk for significant intestinal and neurodevelopmental consequences, there is no known specific treatment. This proposal will investigate what aspects of intestinal immaturity contribute to this disease and determine if factors secreted by certain beneficial or probiotic bacteria can influence these aspects, thus decreasing the susceptibility of vulnerable infants to necrotizing enterocolitis.