Sequence Variants Impacting Cardiomyocyte S-phase Activity in Inbred Mice Following Injury

序列变异影响近交小鼠损伤后心肌细胞 S 期活性

基本信息

批准号：
10550204
负责人：
LOREN J FIELD
金额：
$ 50.44万
依托单位：
INDIANA UNIV-PURDUE UNIV AT INDIANAPOLIS
依托单位国家：
美国
项目类别：
财政年份：
2021
资助国家：
美国
起止时间：
2021-02-15 至 2025-01-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10550204
关键词：
Abbreviations Adult Alleles Anatomy Animals Apoptosis Backcrossings Bromodeoxyuridine Candidate Disease Gene Cardiac Cardiac Myocytes Cell Cycle Cell Cycle Progression Chromosome 3 Congenic Mice Distal Echocardiography Exhibits Genes Genetic Genetically Modified Animals Heart Heart Injuries Heterozygote Hypertrophy Image Analysis Inbred DBA Mice Inbred Mouse Inbred Strains Mice Infarction Infusion procedures Injury Location Measurement Molecular Analysis Monitor Mouse Strains Mus Myocardial Myocardial Infarction Natural regeneration Nucleotides Pattern Phenotype Reporter S phase Structure System Systems Analysis Testing Time Transgenes Transgenic Organisms Validation Variant Ventricular Remodeling autosome chromosomal location coronary fibrosis exome sequencing experimental study genetic testing genetic variant genome editing heart function hemodynamics improved insight interest myocardial injury offspring protein structure regenerative growth therapeutic development therapeutic target trait transcriptome sequencing

项目摘要

Numerous studies have shown that induction of cardiomyocyte cell cycle activity can have a profound beneficial impact on cardiac structure and function following myocardial infarction. It has also been shown that genetic background can impact the intrinsic rate of cardiomyocyte cell cycle activity in mice. We have observed that mice in a DBA/2J genetic background (abbreviated DBA) have very low levels of cardiomyocyte cell cycle activity following myocardial infarction. However, when crossed with C57Bl6/NCR mice (abbreviated NCR), the resulting (DBA x NCR)-F1 animals mice exhibit a marked increase in cardiomyocyte S-phase activity following infarction, indicating the presence of an autosomal gene (or genes) in the NCR background which acts in a dominant manner to facilitate cell cycle re-entry. Analysis of backcross mice established that this gene (or genes) resides in a region of interest (ROI) located on the distal end of chromosome 3. The experiments proposed in Aim 1 will test hypothesis that a single gene within the ROI is responsible for elevated cardiomyocyte S-phase activity post-infarction. Candidate genes within this region will be identified based on expression patterns observed in infarcted DBA vs. NCR hearts as well as by the presence of sequence variants predicted to impact protein structure and/or activity. The candidates will be systematically tested by generating genetically modified animals, subjecting them to myocardial infarction, and then monitoring the level of cardiomyocyte S-phase activity; an induction of cardiomyocyte cell cycle activity would confirm that the candidate gene being tested is responsible for the trait. The experiments proposed in Aim 2 will test the hypothesis that the elevated cell cycle activity encoded by the NCR ROI alleles has a positive impact on the diminished cardiac function and adverse myocardial remodeling which is encountered post-infarction. Congenic mice in a DBA genetic background which retain heterozygosity on the distal end of chromosome 3 and thus carry the NCR allele (or alleles) which is a major contributor to cardiomyocyte S-phase induction will be generated. The mice will then be subjected to myocardial infarction and longitudinal functional analysis. Terminal analyses will include comprehensive hemodynamic measurements as well as assessment of adverse remodeling (cardiomyocyte apoptosis, hypertrophy and myocardial fibrosis); relative improvements in cardiac function and structure would indicate a beneficial effect from the NCR-encoded cell cycle activity following myocardial injury. Ultimately, the identification and validation of genes underlying intrinsic differences in cardiomyocyte cell cycle rates observed in different strains of mice could suggest potential therapeutic targets with which to enhance regenerative growth in injured hearts.

大量研究表明，诱导心肌细胞细胞周期活性可以产生深远的影响。对心肌梗塞后的心脏结构和功能产生有益的影响。还表明遗传背景可以影响小鼠心肌细胞周期活动的内在速率。我们观察到 DBA/2J 遗传背景（缩写为 DBA）的小鼠心肌细胞周期水平非常低心肌梗塞后的活动。然而，当与 C57Bl6/NCR 小鼠（缩写为 NCR）杂交时，由此产生的 (DBA x NCR)-F1 动物小鼠表现出心肌细胞 S 期活性显着增加梗塞，表明 NCR 背景中存在常染色体基因（或多个基因），该基因在促进细胞周期重新进入的主导方式。对回交小鼠的分析表明该基因（或多个基因）位于 3 号染色体远端的感兴趣区域 (ROI)。目标 1 将检验 ROI 内的单个基因导致心肌细胞 S 期升高的假设梗塞后的活动。将根据表达模式识别该区域内的候选基因在梗死的 DBA 与 NCR 心脏中观察到的结果以及预测会影响的序列变异的存在蛋白质结构和/或活性。候选人将通过生成转基因药物进行系统测试对动物进行心肌梗塞，然后监测心肌细胞S期的水平活动;心肌细胞细胞周期活性的诱导将证实正在测试的候选基因是对特质负责。目标 2 中提出的实验将检验以下假设：细胞周期延长 NCR ROI 等位基因编码的活性对心脏功能减弱和不良反应具有积极影响梗塞后发生的心肌重塑。具有 DBA 遗传背景的同类小鼠 3 号染色体远端保留杂合性，因此携带 NCR 等位基因（或多个等位基因），该等位基因是心肌细胞S期诱导的主要贡献者将产生。然后小鼠将受到心肌梗塞和纵向功能分析。终端分析将包括全面的血流动力学测量以及不良重塑评估（心肌细胞凋亡、肥厚和心肌纤维化）；心脏功能和结构的相对改善表明心肌损伤后 NCR 编码的细胞周期活动产生有益作用。最终，鉴定并验证不同细胞中观察到的心肌细胞细胞周期率内在差异的基因小鼠品系可能会提出潜在的治疗靶标，以增强受伤小鼠的再生生长心。