Abstract
Primary human endothelial cells represent an essential tool to model endothelial dysfunction and to screen interventions for its treatment. Here, we developed a protocol for the synchronous isolation of primary human saphenous vein endothelial cells (HSaVEC), human internal thoracic artery endothelial cells (HITAEC), and human microvascular endothelial cells (HMVEC) from SV and ITA utilized as conduits during coronary artery bypass graft surgery and from subcutaneous adipose tissue excised while providing an access to the heart. Treatment by collagenase type IV and magnetic separation with anti-CD31-antibody-coated beads ensured relatively high efficiency of the isolation (≈60% for HSaVEC, ≈50% for HITAEC, and ≈20% for HMVEC) and high purity (≥99%) of isolated ECs within ≈2 weeks (HSaVEC), ≈2-3 weeks (HITAEC), and ≈3-4 weeks (HMVEC). A colorimetric assay of cell viability and proliferation, as well as real-time bioimpedance monitoring using the xCELLigence instrument, demonstrated high proliferative activity in HSaVEC, HITAEC, and HMVEC, whilst the in vitro tube formation assay indicated their angiogenic potential. The isolation of HSaVEC, HITAEC, and HMVEC from patients undergoing coronary artery bypass graft surgery is a promising option to investigate endothelial heterogeneity, to interrogate endothelial responses to various stresses, and to pinpoint the optimal approaches for restoring endothelial homeostasis, thereby reproducing them within the bedside-to-bench-to-bedside concept.
Keywords:
angiogenesis; coronary artery bypass graft surgery; coronary artery disease; endothelial cell isolation; endothelial cell proliferation; immunophenotyping; internal thoracic artery; microvessels; saphenous vein; subcutaneous adipose tissue.