Crosstalk Between Oxidative Stress and Endoplasmic Reticulum (ER) Stress in Endothelial Dysfunction and Aberrant Angiogenesis Associated With Diabetes: A Focus on the Protective Roles of Heme Oxygenase (HO)-1

Abstract

Type-2 diabetes prevalence is continuing to rise worldwide due to physical inactivity and obesity epidemic. Diabetes and fluctuations of blood sugar are related to multiple micro- and macrovascular complications, that are attributed to oxidative stress, endoplasmic reticulum (ER) activation and inflammatory processes, which lead to endothelial dysfunction characterized, among other features, by reduced availability of nitric oxide (NO) and aberrant angiogenic capacity. Several enzymatic anti-oxidant and anti-inflammatory agents have been found to play protective roles against oxidative stress and its downstream signaling pathways. Of particular interest, heme oxygenase (HO) isoforms, specifically HO-1, have attracted much attention as major cytoprotective players in conditions associated with inflammation and oxidative stress. HO operates as a key rate-limiting enzyme in the process of degradation of the iron-containing molecule, heme, yielding the following byproducts: carbon monoxide (CO), iron, and biliverdin. Because HO-1 induction was linked to pro-oxidant states, it has been regarded as a marker of oxidative stress; however, accumulating evidence has established multiple cytoprotective roles of the enzyme in metabolic and cardiovascular disorders. The cytoprotective effects of HO-1 depend on several cellular mechanisms including the generation of bilirubin, an anti-oxidant molecule, from the degradation of heme; the induction of ferritin, a strong chelator of free iron; and the release of CO, that displays multiple anti-inflammatory and anti-apoptotic actions. The current review article describes the major molecular mechanisms contributing to endothelial dysfunction and altered angiogenesis in diabetes with a special focus on the interplay between oxidative stress and ER stress response. The review summarizes the key cytoprotective roles of HO-1 against hyperglycemia-induced endothelial dysfunction and aberrant angiogenesis and discusses the major underlying cellular mechanisms associated with its protective effects.

Keywords: ER stress; angiogenesis; diabetes; endothelial dysfunction; heme oxygenase-1 (Ho-1); hyperglycemia; oxidative stress.

FIGURE 1

Intracellular hyperglycemia results in increased production of mitochondrial ROS. ROS cause DNA damage releasing PARP-1 which inhibits G3PDH. This results in the accumulation of upstream glycolytic intermediates. Other pathways are activated as a result of the accumulation of glycolytic intermediates upstream of G3P which have detrimental effects on the cell; these pathways are; (1) Polyol pathway which depletes the cell of its antioxidant GSH system due to scavenging of NADPH, thereby augmenting ROS production, (2) Methylglyoxal which is a precursor to AGEs/RAGEs which activate pro-inflammatory and pro-oxidant molecules, (3) DAG/PKC which impairs insulin signaling and eNOS activity, activates pro-inflammatory and pro-oxidant molecules, (4) Hexosamine pathway with the resultant activation of pro-inflammatory molecules, inhibition of eNOS at the Akt activation site and activation of TGF-β with increase in ROS. All these pathways increase ROS in a positive feedback loop. AGEs, advanced glycation end products; DAG, diacylglycerol; eNOS, endothelial NO synthase; G3PDH, Glyceraldehyde 3-phosphate dehydrogenase; GSH, Glutathione; ICAM-1, intracellular adhesion molecule-I; IL-6, interleukin 6; NAD, nicotinamide adenine dinucleotide; NADP, NAD phosphate; NF-κB, nuclear factor kappa B; NO, nitric oxide; PARP-1, poly ADP ribose polymerase 1; PKC, protein kinase C; RAGE, AGEs receptor; ROS, reactive oxygen species; TGF-β, transforming growth factor β; TNF-α, tumor necrosis factor α; VCAM-1, vascular cell adhesion molecule 1. The symbol means inhibition.

FIGURE 2

Pentose phosphate pathway (PPP) is inhibited by hyperglycemia and ROS. Hyperglycemia-induced ROS cause the inhibition of G6PDH which is the rate limiting enzyme for this pathway. This results in reduced NADPH production, a cofactor fundamental in the GSH/peroxidase antioxidant system, with the consequent buildup of more ROS causing a vicious circle of oxidative stress induction. G6PDH, glucose-6-phosphate dehydrogenase; GSH, glutathione; NADPH, reduced NAD phosphate. The symbol means inhibition.

FIGURE 3

Polyol and triose phosphate pathways activation caused by hyperglycemia-induced ROS. Hyperglycemia results in an excess of ROS production which inhibits G3PDH with the consequent accumulation of glycolytic intermediates upstream to G3P leading to increase flux into other pathways, namely polyol and triose phosphate. In polyol pathway, NAD reducing equivalent is consumed by sorbitol dehydrogenase reaction leading to reductions in NAD levels inside the cell which further confounds the inhibition of G3PDH. DHAP, dihydroxyacetone phosphate; G3PDH, glyceraldehyde 3-phosphate dehydrogenase; NAD, nicotinamide adenine dinucleotide; TPI, triose phosphate isomerase. The symbo means inhibition.

FIGURE 4

UPR response activation. ER membrane-bound proteins, PERK, ATF-6 and IRE-1α, are activated when unfolded proteins accumulate within the ER lumen and GRP78/BiP dissociates from them. Downstream effects of PERK involve the phosphorylation of eIF-2α resulting in the attenuation of protein translation with the preferential increase in ATF-4 expression. ATF-6 (p90) translocates to the Golgi body where it undergoes proteolytic cleavage. Active cleaved ATF-6 (p50) translocates then to the nucleus and stimulates the expression of molecular chaperones to aid in the folding process. Finally, active IRE-1α through its endoribonuclease activity undergoes intron splicing of XBP-1 mRNA to synthesize the active spliced XBP-1s which binds to the promoter regions of several genes involved in the synthesis of chemical chaperones and ERAD proteins to restore normal protein homeostasis as part of the UPR rescue response. ATF-6, activating transcription factor 6; BiP, binding immunoglobulin protein; eIF-2α, Eukaryotic translation initiation factor 2α; ERAD, endoplasmic-reticulum-associated protein degradation; GRP78, 78-kDa glucose regulated protein; IRE-1α, inositol-requiring enzyme 1α; PERK, PKR-like ER kinase; XBP-1, X-box binding protein 1.

FIGURE 5

Schematic representation showing ER protein folding as a source of ROS. During ER stress response, there is accumulation of misfolded proteins within the ER where non-native disulfide bonds form resulting in the depletion of glutathione (GSH) that is trying to correct the aberrant bonds. IRE-1α, inositol-requiring enzyme 1 α; PDI, protein disulfide isomerase.

FIGURE 6

Hyperglycemia-induced ROS generation and activation of ER stress response. Hyperglycemia-generated ROS can promote the accumulation of unfolded proteins within the ER contributing to ER stress response through different mechanisms. ER dysfunction causes GSH depletion, Ca²⁺ leak from the ER lumen into the cytosol and into the mitochondria, and ATP depletion that can impair its structural and functional integrity and stimulate mitochondrial ROS production. ROS, in turn, can alter protein folding, thereby inducing unfolded protein accumulation within the ER lumen. ATF-6, activating transcription factor 6; BiP, binding immunoglobulin protein; CHOP, C/EBP homologous protein; ERAD, endoplasmic-reticulum-associated protein degradation; GSH, Glutathione; GSSG, oxidized Glutathione; IRE-1α, inositol-requiring enzyme 1α; JNK, c-Jun N-terminal kinases; NF-κB, nuclear factor kappa B; PERK, PKR-like ER kinase; ROS, reactive oxygen species.

FIGURE 7

Mitochondrial electron transport chain in hyperglycemia. Under hyperglycemic conditions, higher levels of glucose-derived pyruvate increase the flux of NADH and FADH₂ into the electron transport chain and consequently increase the proton gradient across the mitochondrial membrane. The transfer of free electrons to complex III is blocked, leading the electrons to be donated to coenzyme Q10, which then transfers electrons to molecular oxygen, thereby generating O₂^-. CoQ10, coenzyme Q10; e^-, electrons; O₂^-, superoxide; GSH, glutathione; GSSG, oxidized glutathione; H₂O₂, hydrogen peroxide; Pi, inorganic phosphate; SOD, O₂^- dismutase.

FIGURE 8

Mechanisms of aberrant angiogenesis induced in hyperglycemia. Increased ROS production in hyperglycemic environment both in vitro and in vivo is believed to cause insult on the angiogenic process. Theories proposed include suppression of VEGF-A signaling, suppression of EPC mobilization, proliferation and differentiation, reduction in NO bioavailability and suppression of Keap1/NrF-2 pathway with resultant suppression in HO-1 expression. EPC, Endothelial progenitor cell; Keap1, Kelch-like ECH-associated protein 1; NrF-2, nuclear factor erythroid 2–related factor 2; VEGF-A, Vascular endothelial growth factor A. The symbol means inhibition.