1.1 Mechanisms behind increased ROS production in ECs
At moderate concentrations, ROS play an important role in maintaining
the proliferation and survival of ECs, but excessive ROS have
detrimental effects on the vascular system (Forstermann et al. ,
2017). In patients with DM, hyperglycaemia accelerates protein glycation
and forms advanced glycation end products (AGEs) (Kay et al. ,
2016). These AGEs bind with cell surface receptors for AGEs (RAGE) to
activate downstream signaling pathways, such as extracellular
signal-regulated kinase (ERK), and subsequently increase ROS production
(Yuan et al. , 2019). Excessive ROS also trigger nuclear poly
ADP-ribose polymerase, which inhibits activity of
glyceraldehydes-3-phosphate dehydrogenase (GADPH) and activates the
polyol pathway of glucose (Giri et al. , 2018). Activated polyol
pathway produces ROS via depleting nicotinamide adenine dinucleotide
phosphate and glutathione, as well increasing the oxidation of
nicotinamide adenine dinucleotide hydrogen during the conversion of
sorbitol to fructose. Inhibition of GADPH also accelerates the
generation of diacylglycerol, subsequently activating protein kinase C
(PKC) and stimulating nicotinamide adenine dinucleotide phosphate
oxidases (NOXs) to generate additional ROS (Yuan et al. , 2019).
Mitochondria are the central regulators for aerobic energy generation,
and ROS are the essential by-product during this process. Diabetes and
hyperglycemia disrupt the mitochondrial respiratory chain and alter
mitochondrial ultrastructure (e.g. mitochondrial fission and fusion),
thereby increasing the ROS production within mitochondria (Forresteret al. , 2018; Brownlee, 2001). Hyperglycemia upregulates both
abundance and activity of the sodium-hydrogen exchanger (NHE) within ECs
(Klug et al. , 2021). Activated NHE promotes the influx of sodium
influx and enhances intracellular calcium (Ca2+) via
triggering sodium-calcium exchanger (Baartscheer et al. , 2017).
Increased intracellular sodium (Na+) triggers the
sodium-calcium exchanger (NCX) and enhances calcium
(Ca2+) influx into the cytosol. The increased
cytosolic Ca2+ then stimulates the PKC-NOXs pathway,
which further increases ROS production (Rastogi et al. , 2016).
Recently, Uthman et al have directly proven the causal link between NHE
activity and oxidative stress in ECs: tumor necrosis factor-α (TNF-α)
enhanced NHE activity and intracellular sodium (Na+),
as well ROS production, and the increased ROS generation was mitigated
by cariporide, a potent inhibitor for NHE. The crucial role of
NHE/Na+-axis in inflammatory related oxidative stress
was further supported by the fact that sodium pump inhibitor ouabain
increased intracellular Na+ and ROS production in
human ECs (Uthman et al. , 2022).
Excessive ROS increase vascular tone and undermine cardiac inotropic
function, contributing to cardiomyopathy (Ritchie et al. , 2020).
Oxidative stress causes endothelial nitric oxide synthase (eNOS)
uncoupling and impairs NO production, the key vasodilator. ROS produced
by NOXs also oxidize the sarcoendoplasmic reticulum calcium transport
ATPase and limit the sensitivity of SMC to NO (Griendling et al. ,
2021). Besides, ROS induce vascular stiffness via upregulating the
expression of vasoactive factors like vascular endothelial growth factor
(VEGF) and extracellular proteins like matrix metalloproteinases
(Griendling et al. , 2021). Vascular remodeling elevates blood
pressure and increases the intensity of cyclic stretch caused by
vasoconstriction-dilation circles (Ohishi, 2018). Enhanced stretch might
exacerbate oxidative stress via upregulating expression of NOXs in ECs,
further increasing ROS production within in ECs (Li et al. ,
2021). Besides, oscillatory shear stress created by disturbed blood flow
also induces oxidative stress in ECs via activating NOXs (Siu et
al. , 2016).
Depletion of nitric oxide (NO) is a crucial mediator for ROS related
cardiac dysfunction (Shah et al. , 2021). EC-derived NO triggers
soluble guanylate cyclase (cGMP) and protein kinase G (PKG) of adjacent
CMs and leads to the phosphorylation of troponin I and reduction of
myofilament Ca2+ sensitivity, thus enhancing
myocardial relaxation in both isolated CMs and whole hearts (Królet al. , 2021; Feil et al. ). Activation of the NO/cGMP/PKG
signaling pathway also maintains phosphorylation of titin within CMs and
prevents the development of cardiac hypertrophy (Shah et al. ,
2021). Moreover, oxidative stress leads to increased secretion of
pro-inflammatory cytokines and chemokines from ECs, as well as
upregulated expression of adhesion molecules and enhanced
monocyte-endothelial attachment (Yuan et al. , 2019). Intensified
ROS production activates src family kinase (SFK) to phosphorylate
vascular endothelial-cadherin (VE-cadherin), leading to VE-cadherin
internalisation and adherens junction disruption. Activated SFK also
promotes the transformation from G actin to F actin to generate stress
fibres under the cellular membrane, increasing the intracellular tension
(Zhang et al. , 2017). ROS are also involved in the endothelial
dysfunction induced by stretch and oscillatory shear stress through
intracellular cascades, such as mitogen-activated protein kinase p38
(p38 MAPK), ERK, c-Jun N-terminal kinase (JNK) and nuclear factor kappa
B (NF-κB) (Lehoux, 2006). The pivotal role of EC-derived ROS in
development of cardiovascular disease is summarised in Figure 1.