In situ detection of stalled cleavage complexes for studies in aging

用于衰老研究的失速裂解复合物的原位检测

• 批准号: 10303806
• 负责人: VLADIMIR V DIDENKO
• 金额: $ 22.5万
• 依托单位: BAYLOR COLLEGE OF MEDICINE
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-09-30 至 2023-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10303806
• 关键词: Aging; Alzheimer' s Disease; Apoptosis; Benchmarking; Biochemical; Biological Assay; Biomedical Research; Catalysis; Cell Aging; Cell Cycle; Cell Nucleus; Cell Survival; Cells; Cellular Metabolic Process; Characteristics; Complex; DNA; DNA Sequence Alteration; DNA Topoisomerases; DNA strand break; Data; Degenerative Disorder; Detection; Development; Disease; Ensure; Enzymes; Eukaryota; Fluorescence Microscopy; Generations; Goals; Heart Diseases; Image; In Situ; In Vitro; Individual; Label; Ligation; Malignant Neoplasms; Mammalian Cell; Masks; Methods; Modeling; Molecular; Molecular Analysis; Natural Products; Normal Cell; Pathologic; Pathology; Pattern; Performance; Poison; Process; Production; Proteins; Reproducibility; Research; Role; Sampling; Signal Transduction; Source; Specificity; Spottings; Stroke; Suspensions; Technology; Testing; Time; Tissues; Topoisomerase; Topoisomerase II; age related; base; drug development; experimental study; in vivo; innovation; molecular marker; new technology; normal aging; novel; novel strategies; prevent; tool

In situ detection of stalled cleavage complexes for studies in aging Abstract This project will introduce and validate the first quantitative in situ approach detecting stalled covalent cleavage complexes (SCCs), which spontaneously originate in mammalian cells during their normal aging. These covalent topoisomerase-DNA strand break intermediates are continually produced and slowly accumulate in aging cells. Their formation is accelerated by age-related degenerative diseases, such as Alzheimer’s. SCCs are generated in abortive catalysis and were recently revealed to be a critical part of the aging process in eukaryotes. In spite of the essential role of SCCs in the normal aging process and in the pathologies associated with aging, at present time, there are no specific approaches detecting and quantitating these DNA alterations in situ with the individual complex sensitivity. The existing bulk biochemical approaches are not applicable to the in situ formats, such as fixed cells and tissue sections. In this project, we will overcome this limitation by developing a new and enabling technology for molecular analysis of aging. The innovative in situ method will be based on novel labeling principles, and will simultaneously co-detect several characteristic features of SCCs. This study will close a significant technological gap and will introduce the first assay for quantitative assessment of SCCs in situ. The project will create a new tool for molecular research in aging and age-related disorders, with broad and general utility in biomedicine at large. The project will reach these Specific Aims: 1. To introduce the first quantitative in situ approach detecting covalent stalled cleavage complexes (SCCs), which spontaneously originate in mammalian cells during their normal aging. To test and validate the labeling principles of the new technology by using in situ models with regulated production of SCCs. To ensure the quantitative ability of the new approach and its high specificity for SCCs. 2. To test and validate the new method in the cell and tissue section models with biochemical and in vivo generation of SCCs. To assess and confirm the quantitative ability of the new approach and its high specificity for SCCs. To ensure robust and reliable assay performance in various samples.

原位检测停滞的切割复合物，用于衰老的研究 抽象的 该项目将介绍并验证第一个定量的原位方法，以确定停滞的共价裂解 复合物（SCC），它们在正常衰老过程中自发起源于哺乳动物。 共价拓扑异构酶-DNA链中间堆积在中间积聚 老化的细胞形成。 SCC在催化性催化中生成 在真核生物中，尽管SCC在正常的衰老过程中起着重要作用 与衰老相关，目前尚无特定方法检测和定量DNA 现有的双重敏感性的原位变化。 适用于原位格式，例如固定细胞和组织切片。 在这个项目中，我们通过开发新的和有利的分子来发展限制 衰老的分析。 同时共同检测SCC的几个特征。 这项研究将缩小大量的技术差距，并将引入第一个定量测定法 SCC的原位评估将创建一个新的工具分子研究 疾病，在生物医学中具有广泛和一般的效用。 该项目将达到这些特定目标： 1。引入第一种定量原位方法，以确定共价裂解复合物（SCC），请 在正常衰老期间自发起源于哺乳动物。 新技术的原理通过使用定期生产的原位模型来确保您 新方法的定量能力及其对SCC的高特异性。 2。在使用生化和体内测试和验证细胞和组织模型中的新方法 生成SCC，以评估和确认新方法的定量能力 用于SCC，以确保各种样品中的稳健和可靠的测定性能。