The Impact of Circadian Misalignment on Colonic Barrier Homeostasis in Ulcerative Colitis

摘要 最近有令人信服的证据表明炎症性肠病(IBD)患者，无论是克罗恩病 疾病(CD)和溃疡性结肠炎(UC)，通常有睡眠障碍，睡眠障碍与 疾病爆发的风险。睡眠/清醒周期、免疫功能、新陈代谢和多种生物过程 都是由昼夜节律编排的。大脑中的中央生物钟之间的昼夜节律失调 已发现环境对多种代谢和胃肠道(Git)有促进作用。 疾病。然而，昼夜节律失调的发生率及其对IBD疾病活动性和胃粘膜的影响 炎症尚未得到证实。我们研究的长期目标是调查 IBD患者昼夜节律失调加重胃粘膜炎症和病程的假说。为了测试 在这一假设下，我们将在实验室与人类受试者进行一项前瞻性研究，以探讨昼夜节律的影响 左侧轻度至中度溃疡性结肠炎。我们将执行两个昼夜节律测量阶段 在严格控制的实验室条件下进行评估：1)基线，2)昼夜节律失调后 在非活动期UC患者和健康对照受试者模拟夜班三天后。要测试 假设UC患者(与健康受试者对照组相比)对昼夜节律失调的恢复能力较差 我们将通过以下方式评估：我们的协议对昼夜节律的时间影响(目标1)： 夹带[从暗光褪黑素开始到睡眠开始的时间](Aim1a)；外周昼夜节律 乙状结肠镜检查结肠组织和受试者外周血中时钟基因的表达 单个核细胞(Aim1b)；以及结肠钟基因表达和PER2：：Luc活性在24小时内利用 结肠有机化合物的体外模型(Aim1c)。接下来，我们将检验这样的假设：昼夜节律失调将 增加UC患者的结肠通透性和粘膜炎症(与对照组相比)(AIM2) 以下指标：内窥镜评分、粪便钙化保护素、结肠屏障(通透性、AJC蛋白)和 系统屏障功能和炎症(目标2a)；对微生物群落产生不利影响 结构/功能(目标2b)；并利用2D结肠有机化合物单分子膜探讨体外屏障功能。 用肿瘤坏死因子-α/干扰素-γ培养(目标2c)。这一创新提案的结果将大大增加我们的 了解昼夜节律失调在UC疾病活动和结肠中的重要作用 炎症，并确定新的昼夜节律调节的UC治疗靶点。

ABSTRACT Recently there has been compelling evidence that inflammatory bowel disease (IBD) subjects, both Crohn's disease (CD) and Ulcerative Colitis (UC), commonly have disrupted sleep, and disrupted sleep correlates with the risk of disease flare. Sleep/wake cycle, immune function, metabolism and multiple biological processes are all orchestrated by circadian rhythms. Circadian misalignment between the central circadian clock in the brain and environment has been found to contribute to a variety of metabolic and gastrointestinal tract (GIT) diseases. Yet, the prevalence and impact of circadian misalignment on IBD disease activity and GIT mucosal inflammation have not been established. The long-term objective of our research is to investigate the hypothesis that circadian malalignment worsens GIT mucosal inflammation and disease course in IBD. To test this hypothesis, we will conduct a prospective in lab study with human subjects on the impact of circadian misalignment in left sided mild to moderate Ulcerative Colitis. We will perform two circadian measures phase assessments under strictly controlled laboratory conditions at: 1) baseline, and after 2) circadian misalignment after three days of simulated night shifts in both inactive UC patients and healthy control subjects. To test the hypothesis that UC patients (compared to healthy subject controls) are less resilient to circadian misalignment we will assess: time impact of our protocol on circadian rhythms (Aim 1) by the following: phase angle of entrainment [time from dim light melatonin onset to sleep onset] (Aim1a); peripheral circadian rhythms by clock gene expression in colonic tissue from a flexible sigmoidoscopy and in subjects peripheral blood mononuclear cells (Aim1b); and colonic clock gene expression and Per2::Luc activity over 24 hours utilizing an in-vitro model of colonic organoids (Aim1c). Next, we will test the hypothesis that circadian misalignment will increase colonic permeability and mucosal inflammation in UC patients (vs. Controls) (Aim2) through the following: endoscopy score, stool calprotectin, colonic barrier (permeability, AJC proteins), and markers of systemic barrier function and inflammation (Aim 2a); adversely impacting microbial community structure/function (Aim 2b); and use 2D colonic organoid monolayers to explore ex-vivo barrier function by co- culturing with TNF-α/INF-γ (Aim 2c). The results of this innovative proposal will greatly increase our understanding of the important role circadian misalignment may have in UC disease activity and colonic inflammation, and identify new circadian regulated targets for treatment in UC.