Single Cell Spatial Transcriptomics Redefines the Borderzone induced by Myocardial Infarction and Mechanical Injury

The ischemic borderzone (BZ) is a geographically complex and biologically enigmatic interface separating poorly perfused infarct zones (IZ) from comparatively healthy remote zones (RZ).  BZ cellular and molecular mechanisms are not well understood because efforts to dissect it inevitably include RZ and IZ in uncontrolled proportions. Here, we use single-cell/nuclei RNA-sequencing, spatial transcriptomics, and multiplexed RNA fluorescence in situ hybridization (mFISH) to identify BZ cardiomyocytes (CMs) subsets. BZ1 (Nppa+Xirp2­-) forms a hundreds-of-microns-thick transitional layer adjacent to RZ, while BZ2 (Nppa+Xirp2­+) forms a tens-of-microns-thick layer that the IZ edge. BZ2 CMs have reduced CM cell contact; colocalize with matricellular-protein-expressing myofibroblasts; and upregulate focal adhesion-, sarcomere-, and cytoskeletal-genes.  Surprisingly, the transcriptional BZ emerges within an hour of injury and is inducible by non-ischemic fine-needle-trauma. We suggest that mechanical instability and “loss of neighbor” at the BZ edge are the dominant inducers of the BZ transcriptional response.