ROCK, tight junctions and prematurity in the pathogenesis of necrotizing enterocolitis and neonatal sepsis.

摘要： 坏死性小肠结肠炎(NEC)是一种致命的新生儿疾病，影响7%的住院患者。 新生儿重症监护病房，是革兰氏阴性新生儿败血症的主要原因。脓毒症是第二种 新生儿死亡的主要原因，每年占全球死亡人数的100万人。尽管我们有 在了解母体定植模式对新生儿败血症的影响方面取得了进展，在 对调节肠道屏障功能的调节过程的理解 易患败血症。事实上，肠道是人体最大的免疫器官，负责 在其管腔内保持对微生物世界的屏障。因此，这是一个高度优先的发展 预防和治疗新生儿败血症的医疗干预。Rho激酶(ROCK)是丝氨酸/苏氨酸激酶 并参与多种细胞过程，包括调节紧密连接功能、肌动蛋白细胞骨架 收缩、炎性细胞因子和细胞死亡。我们发现Rho相关蛋白激酶(ROCK)是 在NEC期间激活，抑制ROCK提供保护，防止肠道屏障损伤和破坏 正常的TJ动态平衡。我们的数据表明，TJ调节、肠屏障功能和细胞凋亡 受岩石依赖的小窝蛋白-1介导的途径调节。此外，我们还发现了以下方面的差异 TJ亚细胞分布和小窝蛋白-1在有、无脓毒症和NEC患者中的分布 不同的孕龄。我们提议的研究的目标是确定屏障损失的机制 在NEC中发生，并利用这一点来了解上皮特异性摇滚抑制在 预防NEC和新生儿败血症。我们的中心假设是肠道屏障的反应 早产儿对损伤的改变和特征是上皮岩信号的变化，异常的TJ 调节，增加粘膜和全身炎症，败血症和上皮细胞死亡。我们将对此进行测试 通过确定是否抑制TJ蛋白的表达和重分布或小窝蛋白-1介导的假说 内吞作用可防止细胞旁通透性增加、粘膜和全身炎症反应，以及 实验性脓毒症和NEC的上皮细胞死亡。人体产生的肠道有机物质的应用 婴儿和岩石和洞穴蛋白-1基因敲除小鼠，将提供巨大的翻译能力，并使我们 探索分子、细胞、组织和生物水平上皮细胞的功能。使用药理学和 我们将确定TJ反应和ROCK信号是否以及如何增加细胞死亡 以恶化早产儿的结局，以及我们的干预措施是预防还是诱发败血症。这些发现 通过提供对机制的新理解，对人类健康产生重大的积极影响 NEC和新生儿败血症期间早产儿肠上皮屏障功能的调控。

ABSTRACT: Necrotizing enterocolitis (NEC) is a deadly disease of the newborn that affects 7% of patients admitted to the neonatal intensive care unit and is a leading cause of gram negative, neonatal sepsis. Sepsis is the second major cause of death among neonates, accounting for one million global deaths per year. Although we have made progress understanding the effect of maternal colonization patterns on neonatal sepsis, there is a gap in the understanding of the regulatory processes that regulate intestinal barrier function that contribute to susceptibility to sepsis. Indeed, the intestine is the largest immune organ in the body and is responsible for maintaining a barrier to the microbial world within its lumen. Therefore, it is a high priority for the development of medical interventions to prevent and treat neonatal sepsis. Rho kinases (ROCK) are serine/ threonine kinases and are involved in multiple cellular processes including regulating tight junction function, actin cytoskeleton contraction, inflammatory cytokines and cell death. We found that Rho-associated protein kinase (ROCK) is activated during NEC, and inhibition of ROCK confers protection against intestinal barrier injury and disruption of normal TJ homeostasis. Our data suggest that TJ regulation, intestinal barrier function and apoptosis are regulated by a ROCK dependent, caveolin-1 mediated pathway. Furthermore, we have identified differences in TJ subcellular distribution and caveolin-1 both in patients with and without sepsis and NEC, and in those of different gestational ages. The objective of our proposed study is to define the mechanism by which barrier loss occurs in NEC and to leverage this in order to understand the potential of epithelial-specific ROCK inhibition in preventing NEC and neonatal sepsis. Our central hypothesis is that the response of the intestinal barrier in premature infants to injury is altered and is characterized by changes in epithelial ROCK signaling, aberrant TJ regulation, increased mucosal and systemic inflammation, sepsis and epithelial cell death. We will test this hypothesis by determining whether inhibition of TJ protein expression and redistribution or caveolin-1-mediated endocytosis prevent paracellular permeability increases, mucosal and systemic inflammatory responses, and epithelial cell death in experimental sepsis and NEC. The use of intestinal organoids generated from human infants and from ROCK and caveolin-1 knockout mice, will provide tremendous translational power and allow us to probe epithelial functions at molecular, cellular, tissue, and organismal levels. Using pharmacological and genetic approaches, we will determine if and how TJ responsiveness and ROCK signaling increases cell death to worsen outcomes in premature infants and whether our interventions prevent or induce sepsis. These findings have a significant positive impact on human health by providing a new understanding of the mechanisms governing epithelial intestinal barrier function of the premature infant during NEC and neonatal sepsis.