Interactions of PPAR-alpha and adenosine receptors in hypoxia-induced angiogenesis

Hypoxia and adenosine are known to upregulate angiogenesis; however, the role of peroxisome proliferator-activated receptor alpha (PPAR alpha) in angiogenesis is controversial. Using transgenic Tg(fli-1:EGFP) zebrafish embryos, interactions of PPAR alpha and adenosine receptors in angiogenesis were evaluated under hypoxic conditions. Epifluorescent microscopy was used to assess angiogenesis by counting the number of intersegmental (ISV) and dorsal longitudinal anastomotic vessel (DLAV) at 28 h post-fertilization (hpf). Hypoxia (6 h) stimulated angiogenesis as the number of ISV and DLAV increased by 18-fold (p < 0.01) and 100 +/- 8% (p < 0.001), respectively, at 28 hpf. Under normoxic and hypoxic conditions, WY-14643 (10 mu M), a PPAR alpha activator, stimulated angiogenesis at 28 hpf, while MK-886 (0.5 mu M), an antagonist of PPAR alpha, attenuated these effects. Compared to normoxic condition, adenosine receptor activation with NECA (10 mu M) promoted angiogenesis more effectively under hypoxic conditions. Involvement of A(2B) receptor was implied in hypoxia-induced angiogenesis as MRS-1706 (10 nM), a selective A(2B) antagonist attenuated NECA (10 mu M)-induced angiogenesis. NECA- or WY-14643-induced angiogenesis was also inhibited by miconazole (0.1 mu M), an inhibitor of epoxygenase dependent production of eicosatrienoic acid (EET) epoxide. Thus, we conclude that: activation of PPAR alpha promoted angiogenesis just as activation of A(2B) receptors through an epoxide dependent mechanism. (C) 2013 Elsevier Inc. All rights reserved.