%0 Journal Article
%T Insulin-Increased L-Arginine Transport Requires A2A Adenosine Receptors Activation in Human Umbilical Vein Endothelium
%A Enrique Guzm¨¢n-Guti¨¦rrez
%A Francisco Westermeier
%A Carlos Salom¨®n
%A Marcelo Gonz¨¢lez
%A Fabi¨¢n Pardo
%A Andrea Leiva
%A Luis Sobrevia
%J PLOS ONE
%D 2012
%I Public Library of Science (PLoS)
%R 10.1371/journal.pone.0041705
%X Adenosine causes vasodilation of human placenta vasculature by increasing the transport of arginine via cationic amino acid transporters 1 (hCAT-1). This process involves the activation of A2A adenosine receptors (A2AAR) in human umbilical vein endothelial cells (HUVECs). Insulin increases hCAT-1 activity and expression in HUVECs, and A2AAR stimulation increases insulin sensitivity in subjects with insulin resistance. However, whether A2AAR plays a role in insulin-mediated increase in L-arginine transport in HUVECs is unknown. To determine this, we first assayed the kinetics of saturable L-arginine transport (1 minute, 37¡ãC) in the absence or presence of nitrobenzylthioinosine (NBTI, 10 ¦Ìmol/L, adenosine transport inhibitor) and/or adenosine receptors agonist/antagonists. We also determined hCAT-1 protein and mRNA expression levels (Western blots and quantitative PCR), and SLC7A1 (for hCAT-1) reporter promoter activity. Insulin and NBTI increased the extracellular adenosine concentration, the maximal velocity for L-arginine transport without altering the apparent Km for L-arginine transport, hCAT-1 protein and mRNA expression levels, and SLC7A1 transcriptional activity. An A2AAR antagonist ZM-241385 blocked these effects. ZM241385 inhibited SLC7A1 reporter transcriptional activity to the same extent in cells transfected with pGL3-hCAT-1£¿1606 or pGL3-hCAT-1£¿650 constructs in the presence of NBTI + insulin. However, SLC7A1 reporter activity was increased by NBTI only in cells transfected with pGL3-hCAT-1£¿1606, and the ZM-241385 sensitive fraction of the NBTI response was similar in the absence or in the presence of insulin. Thus, insulin modulation of hCAT-1 expression and activity requires functional A2AAR in HUVECs, a mechanism that may be applicable to diseases associated with fetal insulin resistance, such as gestational diabetes.
%U http://www.plosone.org/article/info%3Adoi%2F10.1371%2Fjournal.pone.0041705