RNA sequencing on uninjured and injured cardiac regions of B6 and C3H strains. Mus musculus

Purpose: To analyze gene expression in calcific (C3H) and non-calcific (B6) hearts Methods: B6 and C3H mice were induced cardiac injury by rapid freeze thaw of cardiac tissue (cryo-injury). A left thoracotomy was performed at the level of 2nd intercostal space and the exposed beating heart was frozen for 10 seconds by gently pressing a pre-cooled steel rod of 1mm diameter in dry ice. Freezing of cardiac tissue was confirmed by the rapid discoloration of the tissue. Seven days after injury, injured and uninjured regions from same heart were used for RNA sequence. Results: In contrast to only 70 odd genes that were differentially upregulated following injury in non-calcified mouse hearts (B6) about 960 genes were upregulated in C3H hearts following injury induced calcification. Out of the 960 differentially upregulated genes, only 35 were found to be common or upregulated in both C3H and B6 hearts after injury illustrating of the overlapping but dramatically different magnitude of the injury response. Families of genes regulating diverse aspects of an injury response including inflammation, extracellular matrix proteins, cell proliferation and collagen production were differentially expressed between the calcific and non-calcific hearts after injury. The mean expression of osteogenic genes (osteogenic signature) was significantly higher in injured C3H hearts compared to uninjured C3H hearts. The osteogenic signature was not higher in injured B6 hearts compared to control uninjured B6 hearts. Conclusions: Calcific hearts compared to non-calcific hearts responded to injury with a dramatically different transcriptional program. Overall design: Gene expression were measured in uninjured and injured regions from each heart. N=3 animals for B6 uninjured and injured hearts; n=3 for C3H uninjured hearts and n=2 for C3H injured heart.