A probiotic-derived protein regulates epigenetic programming in intestinal epithelial cells for long-term prevention of colitis

益生菌衍生的蛋白质调节肠上皮细胞的表观遗传编程，以长期预防结肠炎

• 批准号: 10386935
• 负责人: FANG YAN
• 金额: $ 44.15万
• 依托单位: VANDERBILT UNIVERSITY MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-04-01 至 2024-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10386935
• 关键词: Address; Adult; Bacteria; Binding; CD4 Positive T Lymphocytes; Cell Line; Cell division; Cell physiology; Child; Chromatin; Clinical Trials; Colitis; Complex; Data; Deposition; Dimerization; Early Intervention; Ecosystem; Enhancers; Epidermal Growth Factor Receptor; Epigenetic Process; Epithelial; Epithelial Cells; Exposure to; Foundations; Funding; Gene Expression; Gene Proteins; Genes; Health; Health Promotion; Histone H3; Histones; Human; Incidence; Individual; Infant; Inflammation; Inflammatory Bowel Diseases; Inherited; Intervention; Intestines; Lactobacillus casei rhamnosus; Lamina Propria; Life; Longevity; Lysine; Mediating; Methyltransferase; Modeling; Mus; Natural regeneration; Neonatal; Outcome; Predisposition; Prevention; Prevention strategy; Prevention therapy; Probiotics; Production; Proteins; Public Health; Regulatory T-Lymphocyte; Reporting; Research; Research Support; Signal Transduction; Supplementation; Testing; Tight Junctions; Transactivation; Transcriptional Activation; Transforming Growth Factor beta; Translational Research; Up-Regulation; Work; antibody inhibitor; autoinflammatory; dimer; dysbiosis; epigenetic memory; gut inflammation; gut microbiome; gut microbiota; high risk; histone methyltransferase; histone modification; imprint; intestinal epithelium; intestinal homeostasis; knock-down; microbiome; mouse model; neonatal period; neonate; neutralizing antibody; novel; novel therapeutic intervention; novel therapeutics; postnatal development; preservation; prevent; programs; promoter; receptor; response; secretory protein; stem cells; transcription factor

PROJECT SUMMARY Dysbiosis in infants and children is associated with increased susceptibility to inflammatory bowel disease (IBD) in adults. However, the mechanisms whereby the gut microbiota colonization in early life confers health outcomes throughout the lifespan remain unclear. Studies from the previous funding period isolated and cloned a Lactobacillus rhamnosus GG (LGG)-derived secretory protein, p40, and demonstrated that neonatal p40 supplementation prevents colitis in adult mice. Our preliminary studies discovered that p40 interacts with two transcriptional factors, Mga and Max, to regulate expression of Setd1β gene, which encodes a methyltransferase for catalyzing mono and trimethylation of histone 3-lysine 4 (H3K4me1/3). We identified TGFβ as a potential target of p40-up-regulated Setd1β. IECs are rapidly renewed and continuously regenerated from intestinal stem cells (ISCs). We found that p40 modulates H3K4m31/3 in ISCs in early stage, and p40 treatment in neonates, but not adult mice, stimulates sustained increase in TGFβ gene expression in IECs. TGFβ has been shown to have multiple functions against inflammation. Thus, we hypothesize that p40 up-regulates Setd1β gene expression in IECs through increasing Mga:Max dimerization. Supplementation with p40 in early life stimulates Setd1β-dependent H3K4me1/3 deposition at the TGFβ locus in ISCs, which is inherited by IECs to enable the sustained increase in TGFβ production, and subsequently prevention of intestinal inflammation in adulthood. In Aim 1, we will determine whether p40-stimulated Setd1β gene expression and H3K4me1/3 are required to drive the increase in expressing TGFβ gene in IECs, and elucidate whether the interaction between p40 and the Mga:Max dimers mediates up-regulation of Setd1β production. In Aim 2, we will identify the temporal window of p40 exposure in early life that causes the sustained increase in TGFβ production in IECs, and define whether p40-regulated sustained increase in TGFβ gene expression in IECs requires the increase of Setd1β gene expression inISCs in early life. Human and mouse enteroids and colonoids and 2D cultures and cell lines with silencing Setd1β or Mga genes, and mouse models of constitutive and inducible Setd1β gene knock down in IECs or ISCs will be utilized for these two aims. In Aim 3, we will determine whether the sustained increase in TGFβ production after p40 supplementation in early life is required to prevent colitis in adult mice. We will use neutralizing antibodies and inhibitors to block TGFβ function in mice with induced and spontaneously developed colitis. We will also determine whether sustained increase in TGFβ production enhances Tregs induction in the intestine and protective epithelial responses for the prevention of colitis in adulthood. This aim will be tested in mice with TGFβ receptor II deletion in CD4+ T cells and Smad4 deletion in IECs. Together, our proposed research will elucidate a novel mechanism underlying the consequences of long-lasting effects of p40 on prevention of colitis, and lay the foundation for developing early intervention with p40 as a novel therapy for prevention of IBD in adults.

项目概要 婴儿和儿童的菌群失调与炎症性肠病的易感性增加有关 然而，生命早期肠道菌群定植的机制导致了成人疾病（IBD）。 整个生命周期的健康结果仍不清楚。 克隆了鼠李糖乳杆菌 GG (LGG) 衍生的分泌蛋白 p40，并证明新生儿 我们的初步研究发现，p40 补充剂可预防成年小鼠的结肠炎。 两个转录因子 Mga 和 Max，调节 Setd1β 基因的表达，该基因编码 我们鉴定出催化组蛋白 3-赖氨酸 4 (H3K4me1/3) 单甲基化和三甲基化的甲基转移酶。 TGFβ 作为 p40 上调 Setd1β 的潜在靶标，正在快速更新并持续。 我们发现 p40 在早期阶段调节 ISC 中的 H3K4m31/3。 新生小鼠（而非成年小鼠）的 p40 治疗可刺激小鼠中 TGFβ 基因表达的持续增加 IECs 已被证明具有多种抗炎症功能，因此我们研究了 p40。 通过增加 Mga:Max 二聚化来上调 IEC 中的 Setd1β 基因表达。 生命早期的 p40 会刺激 ISC 中 TGFβ 位点上依赖 Setd1β 的 H3K4me1/3 沉积，这是 IEC 继承的技术能够持续增加 TGFβ 的产生，并随后预防 在目标 1 中，我们将确定 p40 是否刺激 Setd1β 基因。 表达和 H3K4me1/3 是驱动 IEC 中 TGFβ 基因表达增加所必需的，并阐明 p40 和 Mga:Max 二聚体之间的相互作用是否介导 Setd1β 产生的上调。 目标 2，我们将确定生命早期 p40 暴露的时间窗口，该窗口导致 IEC 中 TGFβ 的产生，以及是否定义了 p40 调节的 TGFβ 基因表达的持续增加 IEC 需要在生命早期增加 ISC 中的 Setd1β 基因表达。 具有沉默 Setd1β 或 Mga 基因的结肠和 2D 培养物和细胞系，以及组成型小鼠模型 在目标 3 中，我们将利用 IEC 或 ISC 中的诱导 Setd1β 基因敲除来实现这两个目标。 确定生命早期补充 p40 后 TGFβ 产量的持续增加是否是 预防成年小鼠结肠炎所需的药物，我们将使用中和抗体和抑制剂来阻断 TGFβ。 我们还将确定诱导性和自发性结肠炎小鼠的功能是否持续。 TGFβ 产生的增加增强了肠道中 Tregs 的诱导和保护性上皮反应 该目标将在 CD4+ T 中 TGFβ 受体 II 缺失的小鼠中进行测试。 我们提出的研究将共同​​阐明一种新的机制。 揭示 p40 对预防结肠炎的长期影响的后果，并为 开发 p40 早期干预作为预防成人 IBD 的新疗法。