Tissue Culture and Molecular Biology

组织培养和分子生物学

• 批准号: 7492098
• 负责人: EDLUE M TABENGWA
• 金额: $ 16.33万
• 依托单位: UNIVERSITY OF ALABAMA AT BIRMINGHAM
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/7492098
• 关键词: Adult; Alcoholic Beverages; Alcohols; Alteplase; Applications Grants; Biological Assay; Cardiac Myocytes; Cardiovascular system; Consumption; Coronary; Coronary heart disease; Endothelial Cells; Ensure; Epidemiologic Studies; Ethanol; Fibrinolysis; Gene Expression; Goals; Human; In Situ Hybridization; Individual; Logistics; Luciferases; Maintenance; Mammalian Cell; Mitochondria; Molecular; Molecular Biology; Mus; Mutagenesis; Myocardial Infarction; Nuclear; Plasmids; Plasminogen Activator Inhibitor 1; Polymerase Chain Reaction; Preparation; Program Research Project Grants; Regulation; Research; Research Personnel; Research Project Grants; Risk; Run-On Assays; Time; Transfection; Umbilical vein; Vitronectin; cell type; copper zinc superoxide dismutase; cost effective; enolase; expression vector; mortality; non-alcoholic; plasmid DNA; polyphenol; programs; red wine; tissue culture; transcription factor

Numerous epidemiological studies indicate that moderate alcohol and red wine consumption reduces the risk for coronary heart disease and the mortality associated with myocardial infarction. Our preliminary studies have demonstrated that the cardioprotective effects of alcoholic beverages result from both alcohol and non-alcoholic components (red wine polyphenols). However, the exact molecular mechanisms of how alcohol and polyphenols result in cardioprotection have not been fully elucidated. The overall programmatic theme of this Program Project Grant application is to further identify/define the molecular regulatory mechanisms that underlie this low ethanol- or polyphenol-induced cardioprotection. The overall goal of Tissue Culture and Plasmid Preparation Core Unit (Core B) is to better ensure high quality research, consistency, facilitate the optimal functioning and collaborative interaction between individual Program Project Grant research projects and investigators. Core B will provide Program Project investigators with cultured mammalian cell types and relevant plasmid DNA for their individual research projects in a highly cost effective, reproducible and efficient manner. These core unit support functions will alleviate Program Project Grant investigators of the logistic responsibilities, labor-intensive culture/maintenance of mammalian cell types and expansion and purification of the relevant plasmid DNA required by the each project. Core Unit B will routinely provide cultured human umbilical vein endothelial cells (HUVECs), human aortic endothelial cells (HAECs), human coronary endothelial cells and cultured mouse endothelial cells to Projects 1 ("Ethanol and Polyphenolic Induced Endothelial Fibrinolysis", Booyse, years 1- 5), 2 ("Regulation of PAI-1 Gene Expression by Ethanol and Polyphenols", Grenett, years 1-5) and 3; ("Cardiovascular Protection by Ethanol and Polyphenols", Parks, years 1-5); human aortic endothelial cells (HAECs) to project 3 (years 1-5) and adult mouse heart muscle cells to Project 4 ("Mitochondria and Cardioprotection by Ethanol and Polyphenols", Darley-Usmar, years 1-5). Core B will also routinely expand and purify relevant plasmid DNA to provide Program Project Grant projects (Projects 1 [years 1-5]; Project 2 [years 1-5]; Project 3 [years 1-5] and Project 4 [years 1-2]) for in-situ hybridizations (t-PA, u-PA, PAI-1, u-PAR, Ann-II, ocenolase eNOS, iNOS, CuZnSOD, MnSOD); Real-Time PCR (PAs, PARs, PAL1, vitronectin, eNOS, iNOS, MnSOD, CuZnSOD); nuclear run-on assays (u-PAR, Ann II, _ enolase, e-NOS, iNOS, MnSOD, CuZnSOD); luciferase constructs for transient transfection assays (t-PA, u-PA, PAI-1); luciferase mutagenesis constructs (PAI-1) and expression vectors for transcription factors.

许多流行病学研究表明，适量饮酒和红酒可以降低患上以下疾病的风险 冠心病和心肌梗死相关死亡率。我们的初步研究表明 表明酒精饮料的心脏保护作用来自酒精和非酒精 红酒多酚（Red Wine Polyphenols）然而，酒精和多酚 对心脏的保护作用尚未完全阐明。本方案项目的总体方案主题 拨款申请是为了进一步确定/定义这种低乙醇-或 多酚诱导的心脏保护作用。组织培养与质粒制备核心单元（Core B）是为了更好地确保高质量的研究，一致性，促进最佳运作和协作 个别计划项目补助金研究项目和研究人员之间的互动。核心B将提供 计划项目研究人员培养的哺乳动物细胞类型和相关的质粒DNA，为他们的个人 以高成本效益、可复制和高效的方式开展研究项目。这些核心单元支持功能 将减轻计划项目赠款调查人员的后勤责任，劳动密集型 哺乳动物细胞类型的培养/维持以及相关质粒DNA的扩增和纯化 每个项目都需要。核心单元B将常规提供培养的人脐静脉内皮细胞 人血管内皮细胞（HUVECs）、人主动脉内皮细胞（HAECs）、人冠状动脉内皮细胞和培养的小鼠血管内皮细胞（HUVECs）。 内皮细胞与项目1（“乙醇和多酚诱导的内皮纤维蛋白溶解”，Booyse，第1年- 5）、2（“Regulation of派-1 Gene Expression by Ethanol and Polyphenols”，Grenett，years 1-5）和3; （“Cardiovascular Protection by Ethanol and Polyphenols”，Parks，1-5年）;人主动脉内皮细胞 （HAECs）为项目3（1-5年），成年小鼠心肌细胞为项目4（“线粒体和 乙醇和多酚类化合物的生物保护”，Darley-Usmar，1-5年）。核心B也将常规地扩展 并纯化相关质粒DNA，以提供计划项目资助项目（项目1 [1-5年];项目2 [第1-5年];项目3 [第1-5年]和项目4 [第1-2年]）进行原位杂交（t-PA、u-PA、派-1、u-PAR， Ann-II，辛烯醇酶eNOS，iNOS，CuZnSOD，MnSOD）;实时PCR（PA，PAR，PAL 1，玻连蛋白，eNOS， iNOS，MnSOD，CuZnSOD）;核连续测定（u-PAR，AnnII，α-烯醇化酶，e-NOS，iNOS，MnSOD， CuZnSOD）;用于瞬时转染测定的荧光素酶构建体（t-PA、u-PA、派-1）;荧光素酶诱变 构建体（派-1）和转录因子的表达载体。