Creeping fat and Crohn's disease associated strictures

摘要 超过一半的克罗恩病（CD）患者发展为狭窄引起的肠梗阻，并且最终 需要手术肠狭窄形成的机制的理解是强制性的，以开发新的 预防和治疗方法。固有肌层（MP）增生，而非细胞外增生 基质（ECM）沉积是肠壁增厚并因此肠腔变窄的主要原因。在 受CD影响的肠段，通常观察到肠系膜脂肪包裹肠，称为 “爬行脂肪”。这是CD的特异性，与MP增生和狭窄的存在高度相关 疾病（有或没有内部穿透性疾病）。基本上没有机械数据将爬行 脂肪与肠狭窄形成或解释蠕动脂肪形成。初步结果显示 肠系膜脂肪衍生脂质选择性诱导人肠MP肌细胞显著增殖 （HIMC）通过长链游离脂肪酸（LC-FFA）代谢并通过线粒体摄取 转运蛋白肉毒碱棕榈酰转移酶1A（CPT-1A）。ECM主要由活化的HIMC释放 纤连蛋白（FN）选择性地促进原代人肠系膜脂肪细胞（Ad）和Pre-Ad的迁移。这 类似于肠段周围的蠕动脂肪的形成。因此，我们提出以下假设： CD中的狭窄形成是反馈回路的结果，其中蠕动脂肪非免疫细胞来源的 因子诱导平滑肌细胞增生，导致ECM分泌增加， 进一步形成爬行脂肪。这一假设将通过三个具体目标进行检验：具体目标1。定义 爬行脂肪诱导平滑肌细胞增生的机制。我们会鉴别出爬行脂肪 介质和他们的细胞来源负责HIMC增殖，重点是FFA信号通路， 增殖、线粒体功能和增殖途径的调节。具体目标2。确定 HIMC衍生的ECM分子在整合素介导的脂肪细胞迁移中的作用。我们将调查 HIMC衍生的ECM导致脂肪迁移，使用功能丧失和功能获得方法， 目的是确定特定的整合素信号通路。具体目标3。研究肠系膜脂肪的作用 缺失和线粒体肌肉代谢对体内肠平滑肌增生的影响。我们将诱导 在两种转基因小鼠品系中的实验性纤维化，其1）表现出可以暂时控制的脂肪缺失 和2）在诱导实验性肠纤维化之前和之后删除线粒体转运蛋白CPT-1 尤其是在β-SMA阳性肌细胞中。此外，我们提出了第一个爬行脂肪的小鼠模型， 是在反复的肠损伤后形成的我们将通过时间评估爬行脂肪的形成和消退， 在β-SMA阳性肌细胞中选择性地控制FN缺失。如果成功，这一提议将挑战 纯粹免疫驱动ECM沉积驱动狭窄形成的范例，并提供了新的 预防或治疗CD患者狭窄相关性肠梗阻的机制。

ABSTRACT More than half of Crohn’s disease (CD) patients develop stricture induced intestinal obstruction and ultimately requiring surgery. A mechanistic understanding of intestinal stricture formation is mandatory to develop novel preventive and therapeutic approaches. Hyperplasia of the muscularis propria (MP) rather than extracellular matrix (ECM) deposition is a major contributor to intestinal wall thickening and hence gut luminal narrowing. In intestinal segments affected by CD, wrapping of mesenteric fat around the bowel is typically observed, called ‘creeping fat’. This is specific for CD and highly associated with the presence of MP hyperplasia and stricturing disease (with or without internal penetrating disease). There are essentially no mechanistic data linking creeping fat with intestinal stricture formation or explaining creeping fat formation. Preliminary results show that mesenteric fat derived lipids selectively induce remarkable proliferation of human intestinal MP muscle cells (HIMC) via long-chain free fatty acids (LC-FFAs) metabolism and uptake into mitochondria through the transporter carnitine palmitoyltransferase 1A (CPT-1A). ECM released by activated HIMC, predominantly fibronectin (FN), selectively promotes migration of primary human mesenteric adipocytes (Ad) and Pre-Ad. This resembles formation of creeping fat around intestinal segments. Hence, we propose the following hypothesis: stricture formation in CD is the result of a feedback loop where creeping fat non-immune cell-derived factors induce smooth muscle cell hyperplasia leading to increased secretion of ECM which promotes further creeping fat formation. This hypothesis will be tested by three specific aims: Specific Aim 1. Define the mechanisms of creeping fat-induced smooth muscle cell hyperplasia. We will identify creeping fat derived mediators and their cellular source responsible for HIMC proliferation, focusing on FFA signaling pathways, proliferation, mitochondrial function and modulation of the proliferation pathways. Specific Aim 2. Determine the role of HIMC-derived ECM molecules in integrin-mediated adipocyte migration. We will investigate mechanisms of HIMC-derived ECM leading to fat migration using a loss-of-function and gain-of-function approach with the goal to identify specific integrin signaling pathways. Specific Aim 3. Investigate the effect of mesenteric fat deletion and mitochondrial muscle metabolism on intestinal smooth muscle hyperplasia in vivo. We will induce experimental fibrosis in two transgenic mouse strains that 1) exhibit fat deletion that can be temporally controlled and 2) delete the mitochondrial transporter CPT-1 prior to and after induction of experimental intestinal fibrosis specifically in -SMA positive muscle cells. In addition, we present the first mouse model for creeping fat, developing after repeated intestinal injury. We will assess creeping fat formation and resolution by temporally controlled deletion of FN selectively in -SMA positive muscle cells. If successful, this proposal will challenge the paradigm of purely immune-driven ECM deposition driving stricture formation and provide novel mechanisms to prevent or treat stricture associated intestinal obstruction in CD patients.