Single-cell sequencing to investigate metabolic stress in the pathology of organ injury following cardiac surgery

Organ injury is a major cause of health complications and in-hospital mortality following cardiac surgery. Mechanisms of biological ageing promote the development of long-term (chronic) conditions which can increase patient susceptibility to these adverse outcomes. To characterise these underlying mechanisms at single-cell resolution, this project sought to profile transcriptomic and chromatin accessibility profiles of atrial cardiac cells from multimorbidity and non-multimorbidity patients recruited to the Ob-Card trial (NCT02908009) using single-cell RNA sequencing (scRNA-seq) and single-nuclei assay for transposase-accessible chromatin sequencing (snATAC-seq). Optimised methods for single-nuclei RNA sequencing (snRNA-seq) provided a better representation of cardiomyocytes and data reproducibility than methods for scRNA-seq using mouse and human heart tissue. Peripheral blood leucocytes from patients and healthy volunteers were used to optimise methods for scRNA-seq and chromatin immunoprecipitation (ChIP) quantitative polymerase chain reaction (qPCR). Methods for snRNA-seq were used for atrial tissue from high (n=9) and low (n=12) multimorbidity patients. Cardiomyocytes (47.2%), fibroblasts (10.2%), various subpopulations of endothelial cells, myeloid cells, lymphoid cells, and other cardiac cell types were identified. Differential gene expression and pathway enrichment analysis found a significant upregulation of translational, ribosomal, protein homeostatic, and apoptotic pathways in cardiac cell types of high multimorbidity patients, with a significant downregulation of cardiac development and contraction processes. Other cell type-specific biological ageing mechanisms, including epigenetic dysregulation, DNA damage/oxidative stress, and immune/inflammatory responses were identified. This was supported by transcription factor (TF) regulatory network and cell-type interaction analyses between immune cells and non-immune cells. snATAC-seq of snap-frozen patient atrial tissue could not be used to relate findings to changes in chromatin structure. These results provide an exploratory approach in characterising biological ageing mechanisms in cardiac cell types of high multimorbidity patients at single-cell resolution, which can help to develop effective risk stratification and therapeutic strategies to avert organ injury and poor outcomes following cardiac surgery.