Hypoxia induces cardiac malformations via A1 adenosine receptor activation in chicken embryos

BACKGROUND: The current understanding of the effects of hypoxia on early embryogenesis is limited. Potential mediators of hypoxic effects include adenosine, which increases dramatically during hypoxic conditions and activates A, adenosine receptors (AIARs). METHODS: To examine the influences of hypoxia and adenosine signaling on cardiac development, chicken embryos were studied. Real time RT-PCR assay was used to examine the AIAR gene expression during embryogenesis and after siRNA- mediated knock down. Cell proliferation was determined by counting cell nuclei and PhosphoHistone H3 positive cells. Apoptosis was determined by TUNEL assay. RESULTS: A(1)ARs were found to be expressed in chicken embryos during early embryogenesis. Treatment of Hamburger and Hamilton stage 4 embryos with the AIAR agonist N-6-cyclopentyladenosine caused cardiac bifida and looping defects in 55% of embryos. Hamburger and Hamilton stage 4 embryos exposed to 10% oxygen for 6, 12, 18, and 24 h followed by recovery in room air until stage 11, exhibited cardia bifida and looping defects in 34, 45, 60, and 86% of embryos respectively. Hypoxia-induced abnormalities were reduced when A(1)AR signaling was inhibited by the A(1)AR antagonist 1,3 dipropyl-8-cyclopentylxanthine or by siRNA-targeting A(1)ARs. Hypoxia treatment did not increase apoptosis, but decreased embryonic cell proliferation. CONCLUSIONS: These data indicate that hypoxia adversely influences cardiac malformations during development, in part by A(1)AR signaling.