Insulin-Increased L-Arginine Transport Requires A2A Adenosine Receptors Activation in Human Umbilical Vein Endothelium
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{"title"=>"Insulin-increased L-arginine transport requires A2A adenosine receptors activation in human umbilical vein endothelium", "type"=>"journal", "authors"=>[{"first_name"=>"Enrique", "last_name"=>"Guzmán-Gutiérrez", "scopus_author_id"=>"36571659400"}, {"first_name"=>"Francisco", "last_name"=>"Westermeier", "scopus_author_id"=>"35390203600"}, {"first_name"=>"Carlos", "last_name"=>"Salomón", "scopus_author_id"=>"36572383100"}, {"first_name"=>"Marcelo", "last_name"=>"González", "scopus_author_id"=>"7404553454"}, {"first_name"=>"Fabián", "last_name"=>"Pardo", "scopus_author_id"=>"54890231300"}, {"first_name"=>"Andrea", "last_name"=>"Leiva", "scopus_author_id"=>"40661639000"}, {"first_name"=>"Luis", "last_name"=>"Sobrevia", "scopus_author_id"=>"6701652093"}], "year"=>2012, "source"=>"PLoS ONE", "identifiers"=>{"pmid"=>"22844517", "scopus"=>"2-s2.0-84864231326", "pui"=>"365316136", "issn"=>"19326203", "isbn"=>"1932-6203 (Electronic)\\r1932-6203 (Linking)", "doi"=>"10.1371/journal.pone.0041705", "sgr"=>"84864231326"}, "id"=>"9c440699-a459-373d-b844-615c8b664eb3", "abstract"=>"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 A(2A) adenosine receptors (A(2A)AR) in human umbilical vein endothelial cells (HUVECs). Insulin increases hCAT-1 activity and expression in HUVECs, and A(2A)AR stimulation increases insulin sensitivity in subjects with insulin resistance. However, whether A(2A)AR 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°C) in the absence or presence of nitrobenzylthioinosine (NBTI, 10 µmol/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 K(m) 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 A(2A)AR in HUVECs, a mechanism that may be applicable to diseases associated with fetal insulin resistance, such as gestational diabetes.", "link"=>"http://www.mendeley.com/research/insulinincreased-larginine-transport-requires-a2a-adenosine-receptors-activation-human-umbilical-vei-2", "reader_count"=>15, "reader_count_by_academic_status"=>{"Unspecified"=>1, "Researcher"=>2, "Student > Doctoral Student"=>4, "Student > Ph. D. Student"=>2, "Student > Postgraduate"=>1, "Student > Master"=>3, "Student > Bachelor"=>1, "Lecturer > Senior Lecturer"=>1}, "reader_count_by_user_role"=>{"Unspecified"=>1, "Researcher"=>2, "Student > Doctoral Student"=>4, "Student > Ph. D. Student"=>2, "Student > Postgraduate"=>1, "Student > Master"=>3, "Student > Bachelor"=>1, "Lecturer > Senior Lecturer"=>1}, "reader_count_by_subject_area"=>{"Unspecified"=>1, "Biochemistry, Genetics and Molecular Biology"=>2, "Agricultural and Biological Sciences"=>5, "Medicine and Dentistry"=>5, "Chemistry"=>2}, "reader_count_by_subdiscipline"=>{"Medicine and Dentistry"=>{"Medicine and Dentistry"=>5}, "Chemistry"=>{"Chemistry"=>2}, "Agricultural and Biological Sciences"=>{"Agricultural and Biological Sciences"=>5}, "Biochemistry, Genetics and Molecular Biology"=>{"Biochemistry, Genetics and Molecular Biology"=>2}, "Unspecified"=>{"Unspecified"=>1}}, "reader_count_by_country"=>{"Greece"=>1, "Chile"=>1}, "group_count"=>0}

Scopus | Further Information

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Figshare

  • {"files"=>["https://ndownloader.figshare.com/files/605155"], "description"=>"<p>(a) Luciferase (<i>Luc</i>) reporter constructs containing serial truncations of <i>SLC7A1</i> promoter (−1606 bp (pGL3-hCAT-1<sup>−1606</sup>) or −650 bp (pGL3-hCAT-1<sup>−1606</sup>) from the transcriptional start point) were transfected in primary cultures of HUVECs incubated (8 hours) without (plain bars) or with (dashed bars) 10 μmol/L nitrobenzylthioinosine (NBTI). Cell transfection was done in the absence or presence of 1 nmol/L insulin and/or ZM-241385 (10 nmol/L), along with <i>Renilla</i> reporter plasmid, and assayed for <i>Firefly</i> and <i>Renilla</i> luciferase activity, respectively. Results depict ratio of <i>Firefly</i>/<i>Renilla</i> luciferase activity. Cells were also transfected with the empty pGL3-basic vector or pGL3-control vector (SV40 pGL3) as negative or positive controls, respectively. (b) Fraction of <i>SLC7A1</i> reporter constructs activity inhibited (sensitive) by ZM-241385 in absence (Basal) or presence of insulin or NBTI as in (a). (c) <i>SLC7A1</i> reporter constructs fraction activity non-inhibited (insensitive) by ZM-241385 as in (a). In (a), *<i>P</i><0.05 versus Control, †<i>P</i><0.05 versus corresponding values in the presence of ZM-241385. In (b), *<i>P</i><0.05 versus corresponding Basal, †<i>P</i><0.05 versus values in pGL3-hCAT-1<sup>−1606</sup> in the presence of insulin, ‡<i>P</i><0.05 versus values in pGL3-hCAT-1<sup>−650</sup> in the presence of insulin or insulin + NBTI. Values are mean ± SEM (n = 6).</p>", "links"=>[], "tags"=>["promoter"], "article_id"=>275648, "categories"=>["Physiology", "Molecular Biology", "Biochemistry", "Chemistry", "Biophysics"], "users"=>["Enrique Guzmán-Gutiérrez", "Francisco Westermeier", "Carlos Salomón", "Marcelo González", "Fabián Pardo", "Andrea Leiva", "Luis Sobrevia"], "doi"=>"https://dx.doi.org/10.1371/journal.pone.0041705.g005", "stats"=>{"downloads"=>1, "page_views"=>15, "likes"=>0}, "figshare_url"=>"https://figshare.com/articles/_SLCA71_promoter_activity_in_response_to_insulin_/275648", "title"=>"<i>SLCA71</i> promoter activity in response to insulin.", "pos_in_sequence"=>0, "defined_type"=>1, "published_date"=>"2013-02-20 02:06:33"}
  • {"files"=>["https://ndownloader.figshare.com/files/605322"], "description"=>"<p>Human umbilical vein endothelial cells respond to insulin via activation of insulin receptors (IR) leading to a reduced (–) adenosine uptake via the human equilibrative nucleoside transporters (hENTs). Insulin reduced adenosine uptake leading to extracellular accumulation of this nucleoside, which turns into activation of A<sub>2A</sub> adenosine receptors (A<sub>2A</sub>) at the plasma membrane. Activation of these membrane receptors leads to a mechanism mediated by transcription factors acting as activators (+) between −1606 and −650 bp from the transcription start point of <i>SLC7A1</i> (for hCAT-1) gene promoter. Alternatively, insulin activates transcription factors acting as activators (+) between −650 bp and the transcription start point of <i>SLC7A1</i>. This phenomenon increases (+) by adenosine receptor-activated transcription factors increasing hCAT-1 mRNA expression and protein abundance resulting in stimulation of L-arginine transport by HUVECs in response to insulin.</p>", "links"=>[], "tags"=>["insulin", "requiring", "adenosine", "receptors", "l-arginine", "fetal", "endothelial"], "article_id"=>275808, "categories"=>["Physiology", "Molecular Biology", "Biochemistry", "Chemistry", "Biophysics"], "users"=>["Enrique Guzmán-Gutiérrez", "Francisco Westermeier", "Carlos Salomón", "Marcelo González", "Fabián Pardo", "Andrea Leiva", "Luis Sobrevia"], "doi"=>"https://dx.doi.org/10.1371/journal.pone.0041705.g007", "stats"=>{"downloads"=>0, "page_views"=>1, "likes"=>0}, "figshare_url"=>"https://figshare.com/articles/_Proposed_model_for_insulin_action_requiring_adenosine_receptors_on_L_arginine_transport_in_human_fetal_endothelial_cells_/275808", "title"=>"Proposed model for insulin action requiring adenosine receptors on L-arginine transport in human fetal endothelial cells.", "pos_in_sequence"=>0, "defined_type"=>1, "published_date"=>"2013-02-20 02:07:31"}
  • {"files"=>["https://ndownloader.figshare.com/files/604925"], "description"=>"<p>L-Arginine transport kinetics (3 μCi/mL L-[<sup>3</sup>H] arginine, 1 minute, 37°C) in HUVECs cultured (8 hours) without (–NBTI; a, c) or with (+NBTI; b, d) 10 μmol/L nitrobenzylthioinosine (NBTI). (a, b) Saturable L-arginine transport in absence (control) or presence of 1 nmol/L insulin and/or ZM-241385 (10 nmol/L) or CGS-21680 (30 nmol/L) as indicated. (c, d) Eadie-Hofstee plot of the L-arginine transport data in (a) and (b), respectively. Values are mean ± SEM (n = 6–18).</p>", "links"=>[], "tags"=>["diabetes and endocrinology", "molecular biology", "physiology", "biophysics", "Biochemistry"], "article_id"=>275412, "categories"=>["Physiology", "Molecular Biology", "Biochemistry", "Chemistry", "Biophysics"], "users"=>["Enrique Guzmán-Gutiérrez", "Francisco Westermeier", "Carlos Salomón", "Marcelo González", "Fabián Pardo", "Andrea Leiva", "Luis Sobrevia"], "doi"=>"https://dx.doi.org/10.1371/journal.pone.0041705.g002", "stats"=>{"downloads"=>0, "page_views"=>4, "likes"=>0}, "figshare_url"=>"https://figshare.com/articles/_L_Arginine_transport_kinetics_/275412", "title"=>"L-Arginine transport kinetics.", "pos_in_sequence"=>0, "defined_type"=>1, "published_date"=>"2013-02-20 02:05:12"}
  • {"files"=>["https://ndownloader.figshare.com/files/605085"], "description"=>"<p>(a) Western blots (representative of other 6 experiments) for hCAT-2A/B in HUVECs incubated (8 hours) without (–NBTI) or with (+NBTI) 10 μmol/L nitrobenzylthioinosine (NBTI), in the absence (–) or presence (+) of 1 nmol/L insulin, CGS-21680 (30 nmol/L) and/or ZM-241385 (10 nmol/L) (ß-actin is internal control). <i>Lower panel</i>: hCAT-2A/B/ß-actin ratio densitometries from data in absence (white bars) or presence (black bars) of NBTI normalized to 1 in cells in the absence of NBTI, insulin, CGS-21680 or ZM-241385 (control). Values are mean ± SEM (n = 6).</p>", "links"=>[], "tags"=>["abundance", "insulin", "adenosine", "receptor", "agonists"], "article_id"=>275572, "categories"=>["Physiology", "Molecular Biology", "Biochemistry", "Chemistry", "Biophysics"], "users"=>["Enrique Guzmán-Gutiérrez", "Francisco Westermeier", "Carlos Salomón", "Marcelo González", "Fabián Pardo", "Andrea Leiva", "Luis Sobrevia"], "doi"=>"https://dx.doi.org/10.1371/journal.pone.0041705.g004", "stats"=>{"downloads"=>2, "page_views"=>7, "likes"=>0}, "figshare_url"=>"https://figshare.com/articles/_hCAT_2A_B_protein_abundance_in_response_to_insulin_and_adenosine_receptor_agonists_and_antagonists_/275572", "title"=>"hCAT-2A/B protein abundance in response to insulin and adenosine receptor agonists and antagonists.", "pos_in_sequence"=>0, "defined_type"=>1, "published_date"=>"2013-02-20 02:06:10"}
  • {"files"=>["https://ndownloader.figshare.com/files/605399"], "description"=>"<p>HUVECs were coincubated (8 hours) in the absence (−<i>NBTI</i>) or presence (+<i>NBTI</i>) of 10 µmol/L nitrobenzylthioinosine (<i>NBTI</i>), without (Control) or with insulin (1 nmol/L), ZM-241385 (10 nmol/L) and/or CGS-21680 (30 nmol/L) (see <a href=\"http://www.plosone.org/article/info:doi/10.1371/journal.pone.0041705#s2\" target=\"_blank\">Methods</a>). Maximal velocity (<i>V</i><sub>max</sub>) and apparent Michaelis-Menten constant (<i>K</i><sub>m</sub>) of saturable transport were calculated assuming a single Michaelis-Menten hyperbola. *<i>P</i><0.05 versus Control in the absence of NBTI. Values are mean ± SEM (n = 19).</p>", "links"=>[], "tags"=>["parameters", "l-arginine"], "article_id"=>275889, "categories"=>["Physiology", "Molecular Biology", "Biochemistry", "Chemistry", "Biophysics"], "users"=>["Enrique Guzmán-Gutiérrez", "Francisco Westermeier", "Carlos Salomón", "Marcelo González", "Fabián Pardo", "Andrea Leiva", "Luis Sobrevia"], "doi"=>"https://dx.doi.org/10.1371/journal.pone.0041705.t002", "stats"=>{"downloads"=>0, "page_views"=>6, "likes"=>0}, "figshare_url"=>"https://figshare.com/articles/_Kinetic_parameters_of_L_arginine_transport_/275889", "title"=>"Kinetic parameters of L-arginine transport.", "pos_in_sequence"=>0, "defined_type"=>3, "published_date"=>"2013-02-20 02:07:57"}
  • {"files"=>["https://ndownloader.figshare.com/files/604855"], "description"=>"<p>(a) Overall (300 µmol/L) L-arginine transport in HUVECs cultured (8 hours) in absence (white bars) or presence (black bars) of 1 nmol/L insulin, without (–NBTI) or with (+NBTI) 10 µmol/L nitrobenzylthioinosine (NBTI). Cells were co-incubated in culture medium without (–) or with (+) CGS-21680 (30 nmol/L) or ZM-21385 (10 nmol/L). (b) L-Arginine transport in the absence or presence of NECA (10 nmol/L) or DPCPX (10 nmol/L) in cells as in (a). (c) Adenosine concentration in the culture medium in cells as in (a). *<i>P</i><0.05 versus cells in the absence of NBTI, CGS-21680 and ZM-241385 (control). Values are mean ± SEM (n = 6–18).</p>", "links"=>[], "tags"=>["extracellular", "adenosine"], "article_id"=>275343, "categories"=>["Physiology", "Molecular Biology", "Biochemistry", "Chemistry", "Biophysics"], "users"=>["Enrique Guzmán-Gutiérrez", "Francisco Westermeier", "Carlos Salomón", "Marcelo González", "Fabián Pardo", "Andrea Leiva", "Luis Sobrevia"], "doi"=>"https://dx.doi.org/10.1371/journal.pone.0041705.g001", "stats"=>{"downloads"=>1, "page_views"=>5, "likes"=>0}, "figshare_url"=>"https://figshare.com/articles/_L_Arginine_transport_and_extracellular_adenosine_concentration_/275343", "title"=>"L-Arginine transport and extracellular adenosine concentration.", "pos_in_sequence"=>0, "defined_type"=>1, "published_date"=>"2013-02-20 02:04:46"}
  • {"files"=>["https://ndownloader.figshare.com/files/605425"], "description"=>"<p>Data are mean ± SEM (range). OGTT, oral glucose tolerance test; BMI, body mass index.</p>", "links"=>[], "tags"=>["characteristics", "patients"], "article_id"=>275920, "categories"=>["Physiology", "Molecular Biology", "Biochemistry", "Chemistry", "Biophysics"], "users"=>["Enrique Guzmán-Gutiérrez", "Francisco Westermeier", "Carlos Salomón", "Marcelo González", "Fabián Pardo", "Andrea Leiva", "Luis Sobrevia"], "doi"=>"https://dx.doi.org/10.1371/journal.pone.0041705.t001", "stats"=>{"downloads"=>0, "page_views"=>3, "likes"=>0}, "figshare_url"=>"https://figshare.com/articles/_Clinical_characteristics_of_patients_and_newborns_/275920", "title"=>"Clinical characteristics of patients and newborns.", "pos_in_sequence"=>0, "defined_type"=>3, "published_date"=>"2013-02-20 02:08:09"}
  • {"files"=>["https://ndownloader.figshare.com/files/604994"], "description"=>"<p>(a) Western blots (representative of other 16 experiments) for hCAT-1 in HUVECs incubated (8 hours) without (–NBTI) or with (+NBTI) 10 μmol/L nitrobenzylthioinosine (NBTI), in the absence (–) or presence (+) of 1 nmol/L insulin and/or ZM-241385 (10 nmol/L) (ß-actin is internal control). <i>Lower panel</i>: hCAT-1/ß-actin ratio densitometries from data in absence (white bars) or presence (black bars) of NBTI, normalized to 1 in cells in the absence of NBTI, insulin and ZM-241385 (control). (b) Western blots for hCAT-1 in the absence or presence of insulin, CGS-21680 and/or ZM-241385 as in (a). <i>Lower panel</i>: hCAT-1/ß-actin ratio densitometries from data in absence (white bars) or presence (black bars) of NBTI normalized to 1 in cells in absence of NBTI, insulin, CGS-21680 and ZM-241385 (control). (c) hCAT-1 mRNA expression relative to 28S rRNA (internal reference) as in (a) and (b). *<i>P</i><0.05 versus values in control. Values are mean ± SEM (n = 16).</p>", "links"=>[], "tags"=>["abundance", "insulin", "adenosine", "receptor", "agonists"], "article_id"=>275480, "categories"=>["Physiology", "Molecular Biology", "Biochemistry", "Chemistry", "Biophysics"], "users"=>["Enrique Guzmán-Gutiérrez", "Francisco Westermeier", "Carlos Salomón", "Marcelo González", "Fabián Pardo", "Andrea Leiva", "Luis Sobrevia"], "doi"=>"https://dx.doi.org/10.1371/journal.pone.0041705.g003", "stats"=>{"downloads"=>0, "page_views"=>6, "likes"=>0}, "figshare_url"=>"https://figshare.com/articles/_hCAT_1_protein_abundance_in_response_to_insulin_and_adenosine_receptor_agonists_and_antagonists_/275480", "title"=>"hCAT-1 protein abundance in response to insulin and adenosine receptor agonists and antagonists.", "pos_in_sequence"=>0, "defined_type"=>1, "published_date"=>"2013-02-20 02:05:37"}
  • {"files"=>["https://ndownloader.figshare.com/files/605234"], "description"=>"<p>(a) Endothelium-intact (circles) or endothelium-denuded (squares) human umbilical vein rings were exposed (8 hours) to increasing concentrations of insulin in absence (○,□) or presence (•,▪) of ZM-241385 (10 nmol/L). (b) Endothelium-intact human umbilical vein rings incubated in absence (–, control) or presence (+) of 10 μmol/L nitrobenzylthioinosine (NBTI) and/or ZM-241384 as in (a). Reactivity was measured in vessel rings co-incubated without (white bars) or with (black bars) 1 nmol/L insulin. Relative responses are given as a percentage fraction of the initial vessel response to KCl (see <a href=\"http://www.plosone.org/article/info:doi/10.1371/journal.pone.0041705#s2\" target=\"_blank\">Methods</a>). (c) Umbilical vein rings with (+ Endothelium) or without (– Endothelium) were exposed for 2 minutes or 8 hours 30 nmol/L CGS-21680 (white bars) in the absence or presence of 10 nmol/L ZM-241385 (black bars) or 10 nmol/L DPCPX (grey bars). Relative responses are as in (a). In (a), *<i>P</i><0.05 versus corresponding values in endothelium-intact vessels. In (b), *<i>P</i><0.05 versus control. In (c), *<i>P</i><0.05 versus all other values except in DPCPX for 2 minutes in endothelium intact vessels. Values are mean ± SEM (n = 7–9).</p>", "links"=>[], "tags"=>["adenosine", "receptors", "insulin", "umbilical", "vein", "rings"], "article_id"=>275717, "categories"=>["Physiology", "Molecular Biology", "Biochemistry", "Chemistry", "Biophysics"], "users"=>["Enrique Guzmán-Gutiérrez", "Francisco Westermeier", "Carlos Salomón", "Marcelo González", "Fabián Pardo", "Andrea Leiva", "Luis Sobrevia"], "doi"=>"https://dx.doi.org/10.1371/journal.pone.0041705.g006", "stats"=>{"downloads"=>1, "page_views"=>5, "likes"=>0}, "figshare_url"=>"https://figshare.com/articles/_Involvement_of_adenosine_receptors_on_insulin_effect_in_human_umbilical_vein_rings_reactivity_/275717", "title"=>"Involvement of adenosine receptors on insulin effect in human umbilical vein rings reactivity.", "pos_in_sequence"=>0, "defined_type"=>1, "published_date"=>"2013-02-20 02:06:59"}

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