Bik and Bcl-2 regulate Nuclear Factor-kappaB-mediated inflammation

Bik 和 Bcl-2 调节核因子 kappaB 介导的炎症

• 批准号: 9403355
• 负责人: Jane Tully Jones
• 金额: $ 0.14万
• 依托单位: LOVELACE BIOMEDICAL & ENVIRONMENTAL RESEARCH INSTITUTE
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-06-30 至 2018-06-29
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9403355
• 关键词: Address; Affect; Alanine; Alleles; Allergens; Allergic Reaction; Allergic inflammation; Amino Acids; Anti-Inflammatory Agents; Anti-inflammatory; Apoptosis; Apoptotic; Attenuated; BCL-2 Protein; BCL2 gene; BH3 Domain; BH3 peptide; Bacterial Infections; Binding; Cell Culture Techniques; Cell Death; Cell Line; Cells; Chronic; Chronic Bronchitis; Data; Disease; Eosinophilia; Epithelial; Epithelial Cells; Exposure to; Extrinsic asthma; Family; Gene Expression Microarray Analysis; Genetic Transcription; Health; Human; Immunofluorescence Immunologic; In Vitro; Infiltration; Inflammation; Inflammatory; Inflammatory Response; Laboratories; Lasers; Lead; Leucine; Lung; Lung diseases; Lymphocyte; Mediating; Metaplasia; Metaplastic; Microdissection; Modeling; Molecular; Molecular Conformation; Mucous body substance; Mus; Nuclear; Nuclear Translocation; Pathologic; Pathway interactions; Patients; Peptides; Pneumonia; Production; Protein Family; Proteins; Pyroglyphidae; Regulation; Role; Scanning; Signal Transduction; Structure of parenchyma of lung; TNF gene; Testing; Therapeutic; Transgenic Organisms; Translational Research; Wild Type Mouse; airway epithelium; airway inflammation; base; cigarette smoke-induced; cigarette smoking; cytokine; experimental study; expression vector; in vivo; inflammatory lung disease; macrophage; mimetics; mouse model; mutant; novel; overexpression; public health relevance; response; small molecule; transcription factor; translational impact

 DESCRIPTION (provided by applicant): Chronic inflammation and mucous cell metaplasia are common pathologic features of the majority of pulmonary diseases, including allergic asthma, pneumonia, and chronic bronchitis. The number of metaplastic mucous cells is regulated by Bcl-2 family-mediated cell death, specifically Bcl-2 and Bcl-2 interacting killer (Bik), in airway epithelia. However, it is not known whether Bcl-2 and Bik regulate inflammation, or the mechanism by which this regulation occurs. Preliminary results demonstrated that cigarette smoke -induced inflammation is significantly increased in bik-/- compared to bik+/+ mice, shown by increased numbers of macrophages and lymphocytes in the lung. Additionally, transgenic expression of Bik in airway epithelia significantly attenuated allergen-induced eosinophilia in comparison to controls, demonstrating an anti- inflammatory role for Bik. In order to determine of Bik modulates inflammation through inhibition of the pro- inflammatory transcription factor, Nuclear Factor-kappaB (NF-κB), its activation was assessed after adenoviral expression of GFP, wild type Bik, mutant Bik (BH3mutBik, Leucine 61to alanine substitution renders Bik unable to induce cell death), or a peptide containing the mutant BH3 domain of Bik . Interestingly, expression of both Bik and BH3mutBik equally in an airway cell line suppressed NF-κB activation in response to TNF-α stimulation. Inhibition of NF-κB in the presence of BH3mutBik occurred in conjunction with NF-κB's association with Bcl-2, a known binding partner of Bik. Because Bik and Bcl-2 associate via their BH3 domains, the effect on NF-κB activation was examined after blocking of the BH3 domain in Bcl-2 using the BH3 mimetic, ABT-263. ABT-263 reduced activation of NF-κB and enhanced its association to Bcl-2 in comparison to controls. Collectively, these data suggest that binding of the BH3 domain of Bcl-2, either by Bik or a BH3 mimetic, blocks nuclear translocation of NF-κB and inflammation. Specific aims of this proposal will (1) demonstrate the molecular mechanism by which Bik and Bcl-2 regulate NF-κB activation and inflammation and (2) test the role of Bik and Bcl-2 following House Dust Mite (HDM)-induced inflammation in vivo. Expression of Bik and Bcl-2 will be modulated in in vitro airway epithelial models, and Bik/Bcl-2 peptides and truncation mutants will be generated in order to test their impact on NF-κB activation. Additionally, adenoviral constructs of Bik and BH3mutBik, and ABT-263, will be administered in the mouse lung during HDM exposure to evaluate their impact on HDM-induced inflammation. According to the hypothesis, it is anticipated that both overexpression of BH3mutBik and inhibition of Bcl-2 will dampen HDM-induced inflammation in the lung. Based on preliminary data demonstrating that the BH3mutBik dampens NF-κB signaling independent of cell death, a novel role for Bcl-2 proteins will be established. Completion of these aims will not only delineate a novel mechanism for Bcl- 2 proteins and NF-κB signaling, but will also result in highly relevant translational research for inflammatory diseases of the lung.

 描述（由适用提供）：慢性炎症和粘液细胞化生是大多数肺部疾病的常见病理特征，包括过敏性哮喘，肺炎和慢性支气管炎。在气道上皮中，Bcl-2家族介导的细胞死亡（特异性BCL-2和BCL-2相互作用杀手（BIK））调节化生粘膜细胞的数量。但是，尚不清楚Bcl-2和BIK是否调节炎症，或该调节发生的机制。初步结果表明，与Bik+/+小鼠相比，BIK - / - 中烟雾诱导的感染显着增加，这显示出肺中巨噬细胞和淋巴细胞数量的增加。此外，与对照相比，BIK在气道上皮中的转基因表达显着减弱了过敏原诱导的嗜酸性粒细胞，这表明BIK具有抗炎作用。 In order to determine Bik modulations injection through inhibition of the pro-inflammatory transcription factor, Nuclear Factor-kappaB (NF-κB), its activation was assessed after adenoviral expression of GFP, wild type Bik, mutant Bik (BH3mutBik, Leucine 61to alanine substitution renders Bik unable to induce cell death), or a peptide containing the mutant BH3 domain of Bik .有趣的是，在气道细胞系中，Bik和BH3Mutbik的表达同样地抑制了NF-κB激活，响应TNF-α刺激。在BH3Mutbik存在下NF-κB的抑制作用与NF-κB与BCL-2的关联（BCL-2）的关联（BCL-2）是BCL-2（BIK的已知结合伙伴）。由于BIK和BCL-2通过其BH3结构域关联，因此使用BH3 Mimetic，ABT-263在BCL-2中阻断BH3域后对NF-κB激活的影响进行了检查。与对照组相比，ABT-263降低了NF-κB的激活，并增强了其与Bcl-2的关联。总的来说，这些数据表明，BCC-2的BH3结构域的结合，无论是BIK还是BH3 MIMETIC，都会阻止NF-κB和炎症的核易位。该提案的具体目的（1）证明了BIK和BCL-2调节NF-κB激活和炎症的分子机制，以及（2）测试BIK和BCL-2在体内诱导的house dust mite（HDM）诱导的炎症之后的作用。 BIK和BCL-2的表达将在体外气道上皮模型中进行调节，并且将生成Bik/Bcl-2 Pepperides和截断突变体，以测试其对NF-κB激活的影响。此外，在HDM暴露期间，将在小鼠肺中施用BIK和BH3Mutbik和ABT-263的腺病毒构建体，以评估其对HDM诱导的炎症的影响。根据该假设，预计BH3Mutbik的过度表达和Bcl-2的抑制作用都会使HDM诱导的肺部炎症。基于初步数据，表明BH3Mutbik DAMBS NF-κB信号独立于细胞死亡，将确定Bcl-2蛋白的新作用。这些目标的完成不仅会描绘出Bcl-2蛋白和NF-κB信号传导的新型机制，而且还将导致对肺部炎症性疾病的高度相关的转化研究。