Multi-Omics Profiling of Human Endothelial Cells from the Coronary Artery and Internal Thoracic Artery Reveals Molecular but Not Functional Heterogeneity

Abstract

Major adverse cardiovascular events occurring upon coronary artery bypass graft surgery are typically accompanied by endothelial dysfunction. Total arterial revascularisation, which employs both left and right internal thoracic arteries instead of the saphenous vein to create a bypass, is associated with better mid- and long-term outcomes. We suggested that molecular profiles of human coronary artery endothelial cells (HCAECs) and human internal mammary artery endothelial cells (HITAECs) are coherent in terms of transcriptomic and proteomic signatures, which were then investigated by RNA sequencing and ultra-high performance liquid chromatography-mass spectrometry, respectively. Both HCAECs and HITAECs overexpressed molecules responsible for the synthesis of extracellular matrix (ECM) components, basement membrane assembly, cell-ECM adhesion, organisation of intercellular junctions, and secretion of extracellular vesicles. HCAECs were characterised by higher enrichment with molecular signatures of basement membrane construction, collagen biosynthesis and folding, and formation of intercellular junctions, whilst HITAECs were notable for augmented pro-inflammatory signaling, intensive synthesis of proteins and nitrogen compounds, and enhanced ribosome biogenesis. Despite HCAECs and HITAECs showing a certain degree of molecular heterogeneity, no specific markers at the protein level have been identified. Coherence of differentially expressed molecular categories in HCAECs and HITAECs suggests synergistic interactions between these ECs in a bypass surgery scenario.

Proteomic Profiling of Endothelial Cells Exposed to Mitomycin C: Key Proteins and Pathways Underlying Genotoxic Stress-Induced Endothelial Dysfunction

“…It is known that the human coronary artery is most often affected by atherosclerosis, while atherosclerotic lesions of the internal thoracic artery are quite rare [87] due to their hydrodynamic features [88] and the molecular heterogeneity of these vessels [89]. We can suggest that molecular heterogeneity is mainly responsible for the differences in the molecular responses of HCAECs and HITAECs to MMC-induced genotoxic stress.…”

Calciprotein Particles Induce Cellular Compartment-Specific Proteome Alterations in Human Arterial Endothelial Cells

“…The lack of considerable differences in molecular response of HCAEC and HITAEC to CPPs can be explained by their relatively low extent of heterogeneity (250-300 differentially expressed proteins and absence of any specific protein markers) and synergistic interactions between differentially expressed protein groups [63]. These results corroborate our previous findings where HCAEC and HITAEC also demonstrated similar molecular response patterns to CPP-P and CPP-S [16,17], although HCAECs were more susceptible to CPP treatment in some gene expression profiling experiments [14,18].…”

“…Gene expression analysis in CPP-P-and CPP-S-treated HCASMC has been performed by reverse transcription-polymerase chain reaction (RT-qPCR) as in [63]. Briefly, a High-Capacity cDNA Reverse Transcription Kit (4368814, Thermo Fisher Scientific, Waltham, MA, USA) was used for the reverse transcription, and RT-qPCR was carried out employing customised primers (500 nmol/L each, Evrogen, Moscow, Russia, Supplementary Table S1), cDNA (20 ng), and PowerUp SYBR Green Master Mix (A25778, Thermo Fisher Scientific, Waltham, MA, USA) according to the manufacturer's protocol for T m ≥ 60 • C (fast cycling mode).…”

“…Target verification of proteomic profiling results in HCAEC was performed by fluorescent Western blotting as in [63]. Chameleon Duo Pre-Stained Protein Ladder (928-60,000, LI-COR Biosciences, Lincoln, NE, USA) was used as a molecular weight marker.…”

“…Chameleon Duo Pre-Stained Protein Ladder (928-60,000, LI-COR Biosciences, Lincoln, NE, USA) was used as a molecular weight marker. Protein separation and transfer were conducted by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and a dry blotting system (iBlot 2 Gel Transfer Device, Thermo Fisher Scientific, Waltham, MA, USA), using nitrocellulose transfer stacks (IB23001, Thermo Fisher Scientific, Waltham, MA, USA) as previously described [63]. Blocking of nonspecific binding was carried out by incubation of nitrocellulose membranes in protein-free Block'n'Boost!…”