Endothelial-Specific Expression of Mitochondrial Thioredoxin Promotes Ischemia-Mediated Arteriogenesis and Angiogenesis

Abstract

Objective-Thioredoxin-2 (Trx2), a major antioxidant protein in mitochondria, enhances nitric oxide bioavailability and inhibits ASK1-dependent apoptosis in endothelial cells (ECs). However, the in vivo role of Trx2 in angiogenesis has not been defined. Here we used EC-specific transgenesis of Trx2 (Trx2-TG) in mice to determine the in vivo function of Trx2 in arteriogenesis and angiogenesis. Methods and Results-In a femoral artery ligation model, Trx2-TG mice had enhanced capacity in limb perfusion recovery and ischemic reserve capacity compared to the nontransgenic littermates. Ischemia-initiated arteriogenesis in the upper limb was augmented in Trx2-TG mice. Trx2-TG mice also showed significantly enhanced capillary formation and maturation in the lower limb. In nontransgenic limb, ischemia specifically induced a downregulation of Trx2 protein, leading to increased oxidative stress, ASK1 activation, and EC apoptosis. In contrast, Trx2-TG maintained a constitutive level of Trx2, reducing the ischemia-induced deleterious responses. We then defined the mechanism by which Trx2 increases angiogenesis using ECs isolated from Trx2-TG mice. Trx2-TG ECs showed increased NO and NO-dependent migration. In addition, these cells were more resistant to oxidative stress-induced activation of ASK1 signaling and apoptosis. Moreover, Trx2-augmented EC survival is NO-independent. To define the relative contributions of Trx2-increased NO and Trx2-reduced ASK1 apoptotic activity to angiogenesis in vivo, we examined Trx2 effects on ischemia-induced angiogenesis in eNOS-deficient mice. The eNOS deletion caused severe impairment in the functional flow recovery in response to ischemia. Trx2 expression in eNOS-KO mice still dramatically inhibited ischemia-induced ASK1 and EC apoptosis, leading to an enhanced functional flow recovery. Conclusion-These in vivo and in vitro data support that Trx2 maintains EC function by two parallel pathwaysscavenging ROS to increase NO bioavailability and inhibiting ASK1 activity to enhance EC survival, facilitating ischemia-mediated arteriogenesis and angiogenesis. Key Words: thioredoxin Ⅲ angiogenesis Ⅲ ischemia Ⅲ apoptosis A ngiogenesis, a process of new blood vessel formation, contributes to various physiological processes and pathological settings. 1 While excessive angiogenesis links atherosclerosis, cancer, and diabetic retinopathy, defects in angiogenesis directly contribute to myocardial infarction and peripheral arterial disease. Recent reports suggest that reactive oxygen species (ROS) can positively or negatively regulate angiogenesis. Although physiological levels of ROS are required for angiogenesis, excess amount of ROS generated during inflammation and ischemic response may inhibit reparative vascular remodeling by inducing endothelial dysfunction and apoptosis. ROS-producing systems in vascular endothelial cells (ECs) are numerous including various NADPH oxidases, xanthine oxidase, the uncoupling of NO synthase as well as mitochondria. [2][3][4][5] The NADPH oxidases have been considered...

Endothelial Dysfunction, and A Prothrombotic, Proinflammatory Phenotype Is Caused by Loss of Mitochondrial Thioredoxin Reductase in Endothelium

“…Thus, EC-specific transgenesis of the mitochondria-located antioxidant Txn2 resulted in significantly enhanced recovery after hindlimb ischemia, despite limiting the usual increase of ROS in response to ischemia. 19 In addition, this EC-specific Txn2 overexpression also improved EC function as a whole and reduced atherosclerotic lesions in apolipoprotein E-deficient mice. 28 In our study, the loss of Txnrd2 in ECs negatively affected endothelial function and attenuated vascular remodeling processes.…”

“…18 It has been shown that Txn2 is downregulated in ECs during an ischemic insult and that overexpression of the enzyme can limit the usually observed, subsequent increase in ROS levels, leukocyte infiltration, and apoptosis. 19 An ubiquitous overexpression of Txn2 can also limit angiotensin II-induced ROS increases, both by controlling ROS emission from the mitochondria and by regulating cytosolic dihydronicotinamide adenine dinucleotide phosphate oxidase subunit expression. 20 Nothing, however, is known on the role of the upstream enzyme, mitochondrial Txnrd2 in the vasculature or specifically in ECs, despite its vital importance, as demonstrated in other organs.…”

Thioredoxin-Interacting Protein Expression Is Required for VEGF-Mediated Angiogenic Signal in Endothelial Cells

“…Other studies showed that enhanced TRX can increase VEGF expression (17,19,31,50), which can account for the observation of similar VEGF levels in TKO and WT animals. These findings clearly suggest that impaired VEGF angiogenic response in this model is likely due to its impaired signaling rather than levels of VEGF.…”

“…Shifting redox state to more GSSG reflects a state of oxidative stress, while shifting to more GSH reflects a state of reductive stress. A great body of evidence supports the emerging role of the TRX system in modulating VEGF and angiogenesis (17,31,33,50). The TRX system is a ubiquitous thiol-reducing system that consists of TRX, NADPH, and homodimeric selenoprotein TRX reductase (36).…”

Deletion of thioredoxin-interacting protein in mice impairs mitochondrial function but protects the myocardium from ischemia-reperfusion injury

“…Trx2 prevents ischemiainduced ROS production in a femoral artery ligation model (37). Overexpression of Trx2-dependent peroxidase (peroxiredoxin-3) improves survival after myocardial infarction with an attenuation of mitochondrial oxidative stress (38).…”