Identification of a soluble guanylate cyclase in RBCs: preserved activity in patients with coronary artery disease

Abstract

Endothelial dysfunction is associated with decreased NO bioavailability and impaired activation of the NO receptor soluble guanylate cyclase (sGC) in the vasculature and in platelets. Red blood cells (RBCs) are known to produce NO under hypoxic and normoxic conditions; however evidence of expression and/or activity of sGC and downstream signaling pathway including phopshodiesterase (PDE)-5 and protein kinase G (PKG) in RBCs is still controversial. In the present study, we aimed to investigate whether RBCs carry a functional sGC signaling pathway and to address whether this pathway is compromised in coronary artery disease (CAD). Using two independent chromatographic procedures, we here demonstrate that human and murine RBCs carry a catalytically active α1β1-sGC (isoform 1), which converts 32P-GTP into 32P-cGMP, as well as PDE5 and PKG. Specific sGC stimulation by NO+BAY 41-2272 increases intracellular cGMP-levels up to 1000-fold with concomitant activation of the canonical PKG/VASP-signaling pathway. This response to NO is blunted in α1-sGC knockout (KO) RBCs, but fully preserved in α2-sGC KO. In patients with stable CAD and endothelial dysfunction red cell eNOS expression is decreased as compared to aged-matched controls; by contrast, red cell sGC expression/activity and responsiveness to NO are fully preserved, although sGC oxidation is increased in both groups. Collectively, our data demonstrate that an intact sGC/PDE5/PKG-dependent signaling pathway exists in RBCs, which remains fully responsive to NO and sGC stimulators/activators in patients with endothelial dysfunction. Targeting this pathway may be helpful in diseases with NO deficiency in the microcirculation like sickle cell anemia, pulmonary hypertension, and heart failure.

Soluble Guanylate Cyclase Stimulators and Activators

“…Although aspects of its mechanism of action are not completely understood, hydroxyurea may prevent red blood cell sickling by increasing fetal hemoglobin expression via NO release (Cokic et al 2003). Red blood cells appear to have a functional NO/sGC/ cGMP signaling pathway; furthermore, red blood cells from patients with endothelial dysfunction (associated with coronary artery disease) are responsive to NO as well as both sGC stimulators and sGC activators (Cortese-Krott et al 2018). Stimulation of the NO/sGC/cGMP pathway has been shown to decrease vascular inflammation in vivo.…”

PTEN inhibition attenuates endothelial cell apoptosis in coronary heart disease via modulating the AMPK–CREB–Mfn2‐mitophagy signaling pathway

“…human umbilical vein endothelial cells (HUVECs) were cultured in DMEM medium with 10% fetal bovine serum (FBS; W. Han, Shi, et al, ). To induce cell damage, 100 μM ox‐LDL was used to incubate with HUVECs for 24 hr under normal conditions when the HUVECs were approximately 80% confluent (Cortese‐Krott et al, ). The cells incubated in the same concentration of dimethyl sulfoxide (DMSO) were used as a negative control group (Ctrl group).…”

Resveratrol attenuates cerebral ischaemia reperfusion injury via modulating mitochondrial dynamics homeostasis and activating AMPK‐Mfn1 pathway

“…Caspase‐3 and caspase‐9 activities were determined using commercial kits (Beyotime Institute of Biotechnology). The levels of antioxidant factors, including glutathione peroxidase (GPX), superoxide dismutase (SOD) and glutathione (GSH), were measured with ELISA kits purchased from Beyotime Institute of Biotechnology …”