Interaction between NOX4 signalling in endothelial colony-forming cells and coronary microvascular function as a key determinant of cardiac remodelling

Abstract

Introduction: Ischaemic heart disease is characterised by restricted blood supply to the myocardium. The progressive death of endothelial cells and cardiomyocytes leads to chronic heart failure (CHF). Highly angiogenic and proliferative umbilical cord blood-endothelial colony-forming cells (CB-ECFCs) express NOX4 as the predominant NADPH isoform for reactive oxygen species (ROS) production, regulating angiogenic capability and the progression of cardiac remodelling.

Methods: Human NOX4 OE or PLAC8 KD CB-ECFCs were cultured under normoxia (21%O2) and hypoxia (1%O2) to assess viability, metabolic/angiogenic function, gene/protein expression, ROS production and protein localisation. NOX4 OE CB-ECFC conditioned media (CM)-treated human coronary microvascular ECs (HCMECs) were incubated under normoxia/hypoxia to assess paracrine effects on barrier function/gene expression. Angiotensin II-induced hypertensive mouse left ventricular (LV) myocardium was injected with mouse NOX4 OE ECFCs to assess cardiac function/structure,gene expression and vasculature.

Results: CB-ECFC angiogenic and metabolic function were suppressed after exposed to hypoxia for 48h, despite maintained cell viability. NOX4 protein expression and ROS production were inhibited, together with downregulated gene expression of NFE2L2, HMOX1, and NOS3. NOX4 OE CB-ECFCs enhanced angiogenic function via activation of eNOS signalling under normoxia, whilst Nrf2 signalling was activated under hypoxia. PLAC8 KD CB-ECFCs exhibited a partially rescued NOX4-mediated H2O2 production and angiogenic function via activation of Nrf2/STAT3-mediated c-Jun/VEGF signalling. NOX4 OE CB-ECFC CM-treated HCMECs exhibited improved migratory ability in both conditions. The predicted activation of heparan sulphate synthesis and inhibition of glycoprotein VI production in NOX4 OE CB-ECFC-treated HCMECs is protective to barrier integrity under hypoxia, whilst barrier function was unchanged. NOX4 OE ECFCs in hypertensive mice increased myocardial vascular density, interstitial collagen deposition and extracellular matrix gene expression (α-SMA and CTGF), with a tendency towards improved LV function.

Conclusions: ECFC NOX4 signalling is critical in regulating angiogenic function, paracrine-mediated HCMEC signalling under hypoxia, and improves vasoreparative capacity on pathological cardiac remodelling.

Date of Award	Dec 2024
Original language	English
Awarding Institution	Queen's University Belfast
Sponsors	Northern Ireland Department for the Economy
Supervisor	Chris Watson (Supervisor), David Grieve (Supervisor) & Karla O'Neill (Supervisor)