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1 Davis Heart and Lung Research Institute, The Ohio State University Medical Center, Columbus, Ohio, United States
2 Department of Surgery, Ohio State University Medical Center, Columbus, Ohio, United States; Davis Heart and Lung Research Institute, The Ohio State University Medical Center, Columbus, Ohio, United States
* To whom correspondence should be addressed. E-mail: chandan.sen{at}osumc.edu.
Myocardial infarction caused by ischemia-reperfusion in the coronary vasculature is a focal event characterized by an infarct-core, bordering peri-infarct zone and remote non-infarct zone. Recently we have reported the first technique, based on laser microdissection pressure catapulting (LMPC), enabling the dissection of infarction-induced biological responses in multi-cellular regions of the heart. Molecular mechanisms in play at the peri-infarct zone are central to myocardial healing. At the infarct site, myocytes are more sensitive to insult than robust fibroblasts. Understanding of cell-specific responses in the said zones is therefore critical. In this work, we describe the first technique to collect the myocardial tissue with a single-cell resolution. The infarcted myocardium was identified using a truncated hematoxylin-eosin (tH&E) stain. Cell elements from the infarct, peri-infarct and non-infarct zones with micron-level surgical precision were collected in a chaotropic RNA lysis solution. Isolated RNA was analyzed for quality employing microfluidics technology and reverse transcribed to generate cDNA. Purity of the collected specimen was established by real-time PCR analyses of cell-specific genes. Previously we have reported that the oxygen-sensitive induction of p21/Cip1/Waf1/Sdi1 in cardiac fibroblasts in the peri-infarct zone plays a vital role in myocardial remodeling. Using the novel LMPC-technique developed herein we confirmed that finding and report for the first time that the induction of p21 in the peri-infarct zone is not limited to fibroblasts but is also evident in myocytes. This work presents the first account of an analytical technique that applies the LMPC technology to study myocardial remodeling with a cell-type specific resolution.
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