Cardiac Stretch-Induced Transcriptomic Changes are Axis-Dependent

The goal of this study was to examine the effect of the major axis of biaxial mechanical stretch on cardiac myocyte gene expression and to identify the signaling pathways and transcription factors regulating these changes. Neonatal cardiac myocytes were cultured on a micropatterned substrate, and the primary stretch axis was applied either parallel or transverse to the myofibril direction. RNA sequencing was conducted to study whole genomic expression changes after acute cardiac myocyte stretch. The results showed a more robust gene response to longitudinal than to transverse stretch. After 30 minutes of stretch, 53 and 168 genes were considered differentially expressed (DE) from transverse and longitudinal stretch, respectively. After 4 hours, the number of DE genes increased to 795 in longitudinal stretch while it decreased to 35 in transverse stretch. Gene ontology term (GO) analysis indicated enrichment of TF activity and protein kinase activity by both stretch axes; whereas longitudinal but not transverse stretch caused expression of genes involved in sarcomere organization and cytoskeletal protein binding. Overall design: Two stretch durations (30 minutes and 4 hours), and both axes of stretch (transverse and longitudinal) were studied for a total of 4 different stretch conditions. A fifth condition of non-stretched cells was used as a control. Three stretchers for each condition served as biological replicates.

本研究旨在探究双轴机械牵拉（biaxial mechanical stretch）的主轴对心肌细胞（cardiac myocyte）基因表达的影响，并鉴定调控此类表达变化的信号通路与转录因子（transcription factors）。研究人员将新生心肌细胞接种于微图案化基底（micropatterned substrate）上，使主要牵拉轴与肌原纤维方向呈平行或垂直状态。随后通过RNA测序（RNA sequencing）分析急性心肌细胞牵拉后全基因组的表达变化。结果表明，相较于垂直牵拉，纵向牵拉所引发的基因应答更为强烈。牵拉30分钟后，垂直牵拉组与纵向牵拉组分别有53个和168个基因被认定为差异表达基因（differentially expressed，DE）；牵拉4小时后，纵向牵拉组的差异表达基因数量增至795个，而垂直牵拉组则下降至35个。基因本体论（Gene Ontology, GO）术语富集分析显示，两种牵拉轴均可富集到转录因子活性与蛋白激酶活性相关的功能条目；仅纵向牵拉可诱导肌节组织与细胞骨架蛋白结合相关基因的表达，垂直牵拉则无此效应。总体实验设计：本研究设置2种牵拉时长（30分钟、4小时）与2种牵拉轴（垂直牵拉、纵向牵拉），共计4种牵拉处理条件；另设未牵拉细胞作为第五组对照。每组设置3个生物学重复（biological replicates），即每个条件采用3套牵拉装置开展平行实验。