Transcriptional regulation of goblet cell metaplasia

杯状细胞化生的转录调控

• 批准号: 8744367
• 负责人: Vladimir Kalinichenko
• 金额: $ 39万
• 依托单位: CINCINNATI CHILDRENS HOSP MED CTR
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-08-05 至 2018-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8744367
• 关键词: Allergens; Applications Grants; Asthma; Binding; Biological Assay; Boxing; Cell Differentiation process; Chronic; Chronic Obstructive Airway Disease; Clara cell; Clinical; Cystic Fibrosis; Data; Disease; Doxycycline; Embryonic Development; Epithelial; Epithelial Cells; Family; Gene Expression; Gene Targeting; Genes; Genetic; Genetic Transcription; Goals; Goblet Cells; Human; Immunohistochemistry; In Vitro; Interleukin-13; Laboratories; Lasers; Lung Inflammation; Malignant Neoplasms; Mediating; Mediator of activation protein; Metaplasia; Modeling; Molecular; Morbidity - disease rate; Mucous body substance; Mus; Pathogenesis; Pathway interactions; Patients; Peptides; Phenotype; Play; Production; Promoter Regions; Proteins; Pyroglyphidae; Regulation; Regulatory Element; Reporter; Reverse Transcriptase Polymerase Chain Reaction; Role; Signal Pathway; Signal Transduction; Testing; Therapeutic; Time; Transcriptional Regulation; Transgenes; Transgenic Mice; Up-Regulation; Western Blotting; airway epithelium; airway hyperresponsiveness; chromatin immunoprecipitation; efficacy testing; forkhead protein; gain of function; in vivo; inhibitor/antagonist; loss of function; lung injury; methacholine; mortality; mouse model; new therapeutic target; novel; prevent; promoter; public health relevance; research study; response; small hairpin RNA; small molecule

DESCRIPTION (provided by applicant): Goblet cell metaplasia, associated with increased mucus production, is a key pathogenic feature of chronic airway disorders, such as asthma, COPD and cystic fibrosis, all of which contribute to significant morbidity and mortality worldwide. Pharmacological targeting of goblet cell metaplasia represents a significant clinical challenge. Therefore, identification of new molecular mechanisms in airway epithelial differentiation will provide novel therapeutic targets for treatment of chronic airway disorders. Our grant proposal focuses on novel molecular mechanisms in goblet cell differentiation that are regulated by FoxM1, a transcription factor from the Forkhead box (FOX) family. While FoxM1 plays an important role in embryonic development and pathogenesis of various cancers, the role of FoxM1 in chronic airway diseases is unknown. In our preliminary data, FoxM1 was induced in airway epithelial cells of mice with asthma-like diseases caused by OVA, IL-13 and house dust mite extract (HDM). FoxM1 was also induced in airway epithelial cells of human patients with asthma and COPD. Genetic deletion of the Foxm1 gene (CCSP-Cre) or pharmacological inhibition of the FoxM1 protein (ARF peptide) in HDM-challenged airway epithelium effectively diminished goblet cell metaplasia, reduced lung inflammation and decreased airway hyper-responsiveness to methacholine. While these data suggest that FoxM1 plays a key role in asthma pathogenesis, molecular mechanisms regulated by FoxM1 remain uncharacterized. We propose to test the hypothesis that FoxM1 acts downstream of the IL-13/Stat6 pathway to induce expression of goblet cell-specific genes in airway epithelial cells. In Aim I, we will use transgenic mice with FoxM1 "gain-of-function" and "loss-of-function" in airway Clara cells to identify downstream FoxM1 target genes critical for allergen-mediated differentiation of Clara cells into goblet cells. Furthermore, we provide preliminary data demonstrating that IL-13 induces FoxM1 expression in cultured human airway epithelial cells and airway epithelium of transgenic mice. Knockdown of FoxM1 in vitro inhibited differentiation of airway epithelial cells toward goblet cell phenotype in response to IL-13 stimulation. In Aim II, we will determine if FoxM1 is required for IL-13/Stat6 signaling to induce goblet cell differentiation in vivo. The IL-13/Stat6 signaling pathway will be activated using intratracheal administration of IL-13 and a Doxycycline-inducible IL-13 transgene. FoxM1 inhibition will be achieved by a genetic approach (IL-13/ CCSP- Cre/ Foxm1-/- mice) and a pharmacological approach (ARF peptide and novel small molecule FoxM1 inhibitors). These experiments will determine whether inactivation of FoxM1 will prevent or decrease IL- 13/Stat6 signaling in airway epithelial cells in vivo. Altogether, these studies will identify molecular mechanisms regulated by FoxM1 in airway epithelial cells and determine the therapeutic benefit of FoxM1 inhibitors in mouse asthma models.

描述（由申请方提供）：杯状细胞化生与粘液生成增加相关，是慢性气道疾病（如哮喘、COPD和囊性纤维化）的关键致病特征，所有这些疾病均导致全球范围内的显著发病率和死亡率。 杯状细胞化生的药理学靶向是一个重大的临床挑战。因此，气道上皮分化的新分子机制的鉴定将为慢性气道疾病的治疗提供新的治疗靶点。我们的资助计划集中在杯状细胞分化的新分子机制上，这些机制由FoxM 1调节，FoxM 1是叉头盒（FOX）家族的一种转录因子。虽然FoxM 1在胚胎发育和各种癌症的发病机制中起着重要作用，但FoxM 1在慢性气道疾病中的作用尚不清楚。在我们的初步数据中，FoxM 1在由OVA、IL-13和屋尘螨提取物（HDM）引起的哮喘样疾病小鼠的气道上皮细胞中诱导。FoxM 1在哮喘和COPD患者的气道上皮细胞中也被诱导。在HDM激发的气道上皮中，Foxm 1基因（CCSP-Cre）的遗传缺失或FoxM 1蛋白（ARF肽）的药理学抑制有效地减少了杯状细胞化生，减少了肺部炎症，降低了气道对乙酰甲胆碱的高反应性。虽然这些数据表明FoxM 1在哮喘发病机制中起着关键作用，但FoxM 1调控的分子机制仍不清楚。我们建议测试的假设，FoxM 1的行为下游的IL-13/Stat 6途径，诱导表达杯状细胞特异性基因在气道上皮细胞。在目标I中，我们将使用在气道Clara细胞中具有FoxM 1“功能获得”和“功能丧失”的转基因小鼠来鉴定对于变应原介导的Clara细胞分化为杯状细胞至关重要的下游FoxM 1靶基因。此外，我们提供的初步数据表明，IL-13诱导FoxM 1表达在培养的人气道上皮细胞和转基因小鼠的气道上皮。FoxM 1在体外的敲除抑制气道上皮细胞对IL-13刺激的反应向杯状细胞表型的分化。在目的II中，我们将确定FoxM 1是否是IL-13/Stat 6信号传导所必需的，以诱导杯状细胞在体内分化。IL-13/Stat 6信号传导途径将使用IL-13和多西环素诱导型IL-13转基因的腹膜内施用来激活。将通过遗传方法（IL-13/CCSP-Cre/Foxm 1-/-小鼠）和药理学方法（ARF肽和新型小分子FoxM 1抑制剂）实现FoxM 1抑制。这些实验将确定FoxM 1的失活是否会阻止或减少体内气道上皮细胞中的IL- 13/Stat 6信号传导。总之，这些研究将确定气道上皮细胞中FoxM 1调节的分子机制，并确定FoxM 1抑制剂在小鼠哮喘模型中的治疗益处。