Inhibition of adenosine kinase induces expression of VEGF mRNA and protein in myocardial myoblasts

Abstract

We tested whether increased endogenous adenosine produced by the adenosine kinase inhibitor GP-515 (Metabasis Therapeutics) can induce vascular endothelial growth factor (VEGF) expression in cultured rat myocardial myoblasts (RMMs). RMMs were cultured for 18 h in the absence (control) and presence of GP-515, adenosine (Ado), adenosine deaminase (ADA), or GP-515 + ADA. GP-515 (0.2-200 microM) caused a dose-related increase in VEGF protein expression (1.99-2.84 ng/mg total cell protein); control VEGF was 1.84 +/- 0.05 ng/mg. GP-515 at 2 and 20 microM also increased VEGF mRNA by 1.67- and 1. 82-fold, respectively. ADA (10 U/ml) decreased baseline VEGF protein levels by 60% and completely blocked GP-515 induction of VEGF. Ado (20 microM) and GP-515 (20 microM) caused a 59 and 39% increase in VEGF protein expression and a 98 and 33% increase in human umbilical vein endothelial cell proliferation, respectively, after 24 h of exposure. GP-515 (20 microM) had no effect on VEGF protein expression during severe hypoxia (1% O(2)) but increased VEGF by an additional 27% during mild hypoxia (10% O(2)). These results indicate that raising endogenous levels of Ado through inhibition of adenosine kinase can increase the expression of VEGF and stimulate endothelial cell proliferation during normoxic and hypoxic conditions.

Adenosine Kinase: Exploitation for Therapeutic Gain

“…Subsequently, those findings were reproduced in a rat model of myocardial infarction (Peart and Gross, 2005). It was shown that the ADK inhibitor GP-515 induced vascular endothelial growth factor expression in cultured rat myocardial myoblasts, an effect that was completely blocked by the addition of ADA, which, when given alone, led to a decrease in baseline vascular endothelial growth factor expression (Gu et al, 2000). Since ADA-in contrast to brain-is highly expressed in the heart (Barankiewicz et al, 1997), ADK inhibitors might not be useful adenosine augmenting agents to promote cardioprotection.…”

Adenosine infusion increases plasma levels of VEGF in humans

“…Adenosine can stimulate endothelial proliferation in vitro (Meininger et al 1988;Ethier & Dobson, 1997;Grant et al 1999; Gu et al 1999) as well as angioge nesis in a variety of in vivo models (Adair et al 1989; Adair et al 1991;Adair, 2005). Although the angiogenic effects of adenosine are poorly understood, it is known that adenosine, mediated by way of adenosine A 2 receptors can upregulate VEGF mRNA and protein expression in many different cell types (Hashimoto et al 1994;Takagi et al 1996;Grant et al 1999;Gu et al 1999;Gu et al 2000;Feoktistov et al 2002;Leibovich et al 2002;Feoktistov et al 2003;Gardener & Olah, 2003). Numerous investigators have postulated that this adenosine-mediated expression of VEGF might have a role in mediating adenosine-induced angiogenesis ; however, it is still not clear whether adenosine can induce VEGF expression in the intact organism.…”

An Emerging Role for Adenosine in Angiogenesis

“…Physiological concentrations of adenosine or adenosine agonists cause a dose-related increase in the proliferation of endothelial cells obtained from bovine aorta, 3 bovine coronary venules, 3 and human umbilical veins, 4,5 but they have no effect on the proliferation of human fibroblasts 4 and rat myocardial myoblasts. 5 Also, raising endogenous levels of adenosine by inhibition of adenosine kinase can stimulate the proliferation of human endothelial cells obtained from umbilical veins. 5 Adenosine stimulates angiogenesis in the body and chorioallantoic membrane of the chick embryo and could mediate a portion of the hypoxia-induced angiogenesis in the same tissues.…”

“…5 Also, raising endogenous levels of adenosine by inhibition of adenosine kinase can stimulate the proliferation of human endothelial cells obtained from umbilical veins. 5 Adenosine stimulates angiogenesis in the body and chorioallantoic membrane of the chick embryo and could mediate a portion of the hypoxia-induced angiogenesis in the same tissues. 6 Early work has also shown that prolonged infusion of adenosine induces angiogenesis in rabbit heart and skeletal muscle.…”

“…7 That adenosine could have a physiological role in the development of the vasculature is supported by the finding that prolonged administration of dipyridamole, an adenosine-reuptake inhibitor, stimulated angiogenesis in both the heart and skeletal muscle in rats; 7 aminophylline, an adenosine-receptor blocker, produced a dose-related decrease in the size of the vasculature in chick embryos. 7 Although the mechanism by which adenosine induces angiogenesis is poorly understood, many studies have shown that the administration of adenosine or adenosine agonists 8 -11 as well as the upregulation of endogenous adenosine 5 can increase the expression of vascular endothelial growth factor (VEGF) in a variety of different cells studied in vitro. VEGF is a key mediator of angiogenesis released from hypoxic tissues in both physiological and pathological conditions 12 and may be subject to negative feedback regulation in the intact animal.…”