Homozygous deletion of early growth response 1 gene and critical limb ischemia after vascular ligation in mice: Evidence for a central role in vascular homeostasis
ABSTRACT The early growth response 1 gene (Egr1) encodes for an immediate to early response transcription factor that is upregulated by changes in vascular strain and hypoxia and in turn upregulates the downstream expressions of a number of angiogenic growth factors. We therefore hypothesized that early growth response 1 may be a critical early messenger governing revascularization in the setting of acute vascular occlusions.
C57 BL/6 mice deficient in the Egr1 gene (knockout) and their wild-type litter mates underwent ligation and excision of the femoral artery with or without the previous administration of 2.7 x 10(9) particle units of an adenoviral vector coding for the vascular endothelial growth factor gene (VEGF) or Egr1. Distal hind limb perfusion was serially measured in these animals with laser Doppler perfusion imaging.
Wild-type mice (n = 9) had nearly complete restitution of hind limb perfusion by day 35 after ligation. In contrast, all noninjected Egr1 knockout mice (n = 5) had severe ipsilateral limb necrosis develop between 1 and 4 days after ligation (P <.0001). Egr1 knockout mice injected with VEGF vector (n = 4) demonstrated significantly improved perfusion relative to baseline by postligation day 28, which persisted to postligation day 35 (P <.05). Egr1 knockout animals injected with Egr1 vector (n = 7) demonstrated a partial recovery of hind limb perfusion relative to VEGF vector-treated knockout animals at postligation day 4 (P <.01), which persisted to day 35.
These findings suggest that early growth response 1 plays a pivotal role in reperfusion responses to vascular occlusion in mice and possibly other mammals.
SourceAvailable from: Ken-ichiro Sasaki[Show abstract] [Hide abstract]
ABSTRACT: Sprouty proteins (Sproutys) inhibit receptor tyrosine kinase signaling and control various aspects of branching morphogenesis. In this study, we examined the physiological function of Sproutys in angiogenesis, using gene targeting and short-hairpin RNA (shRNA) knockdown strategies. Sprouty2 and Sprouty4 double knockout (KO) (DKO) mice were embryonic-lethal around E12.5 due to cardiovascular defects. The number of peripheral blood vessels, but not that of lymphatic vessels, was increased in Sprouty4 KO mice compared with wild-type (WT) mice. Sprouty4 KO mice were more resistant to hind limb ischemia and soft tissue ischemia than WT mice were, because Sprouty4 deficiency causes accelerated neovascularization. Moreover, suppression of Sprouty2 and Sprouty4 expression in vivo by shRNA targeting accelerated angiogenesis and has a therapeutic effect in a mouse model of hind limb ischemia. These data suggest that Sproutys are physiologically important negative regulators of angiogenesis in vivo and novel therapeutic targets for treating peripheral ischemic diseases.PLoS ONE 02/2009; 4(5):e5467. DOI:10.1371/journal.pone.0005467 · 3.53 Impact Factor
[Show abstract] [Hide abstract]
ABSTRACT: Teratogen exposure activates damage response pathways during organogenesis that determine the fate of the embryo. Cyclophosphamide, an alkylating agent, induces growth reduction defects in organogenesis stage mouse limbs. Here we identify components of the signaling network triggered by in vitro exposure of CD-1 murine limbs to 4-hydroperoxycyclophosphamide (4-OOHCPA), a preactivated analog of cyclophosphamide. The predominant response was downregulation of gene expression; many of the affected genes were transcription factors, transcription regulators, or oncogenes. Pathway analysis of the genes regulated by 4-OOHCPA exposure revealed a novel damage response pathway in limbs comprised of basic transcription factors, Hif1a, Ndn, Hes1 and Myog, transcription activators and repressors, Egr1 and E2f1, intracellular transducers, effectors and modulators, Bmpr1b and Pea15, and oncogenes and tumor suppressors, Hras1, Abl1, Smad1, and Ttf1. Thus, teratogen exposure triggers both developmentally specific signaling pathways and a general damage response. We hypothesize that hypoxia signaling plays a central role in integrating these responses.Reproductive Toxicology 05/2009; 27(2):103-10. DOI:10.1016/j.reprotox.2009.01.014 · 2.77 Impact Factor
[Show abstract] [Hide abstract]
ABSTRACT: Abstract Cardiac gene therapy offers a strategy to treat diffuse coronary artery disease (CAD), a disorder with no therapeutic options. The use of genes to revascularize the ischemic myocardium has been the focus of two decades of preclinical research with a variety of angiogenic mediators, including vascular endothelial growth factor, fibroblast growth factor, hepatocyte growth factor, and others encoded by DNA plasmids or adenovirus vectors. The multifaceted challenge for developing efficient induction of collateral vessels in the ischemic heart requires a choice for route of delivery, dosing level, a relevant animal model, duration of treatment, and assessment of phenotype for efficacy. Overall, studies of gene therapy for ischemia in experimental models are very encouraging, with clear evidence of safety and efficacy, strongly supporting the concept that gene therapy to induce angiogenesis is a viable therapeutic approach for CAD. Clinical studies of cardiac gene therapy with angiogenic factors have added substantially to the evidence for efficacy, but definitive studies have not yet led to commercial approval. This review provides the general concepts for angiogenesis-based therapeutic approaches for diffuse CAD and summarizes the results from key studies in the field with recommendations for refinement to a successful product design and evaluation.Human gene therapy 11/2013; 24(11):948-63. DOI:10.1089/hum.2013.2516 · 3.62 Impact Factor