title: Adenosine Attenuates Human Coronary Artery Smooth Muscle Cell Proliferation by Inhibiting Multiple Signaling Pathways That Converge on Cyclin D.
abstract: The goal of this study was to determine whether and how adenosine affects the proliferation of human coronary artery smooth muscle cells (HCASMCs). In HCASMCs, 2-chloroadenosine (stable adenosine analogue), but not N(6)-cyclopentyladenosine, CGS21680, or N(6)-(3-iodobenzyl)-adenosine-5'-N-methyluronamide, inhibited HCASMC proliferation (A2B receptor profile). 2-Chloroadenosine increased cAMP, reduced phosphorylation (activation) of ERK and Akt (protein kinases known to increase cyclin D expression and activity, respectively), and reduced levels of cyclin D1 (cyclin that promotes cell-cycle progression in G1). Moreover, 2-chloroadenosine inhibited expression of S-phase kinase-associated protein-2 (Skp2; promotes proteolysis of p27(Kip1)) and upregulated levels of p27(Kip1) (cell-cycle regulator that impairs cyclin D function). 2-Chloroadenosine also inhibited signaling downstream of cyclin D, including hyperphosphorylation of retinoblastoma protein and expression of cyclin A (S phase cyclin). Knockdown of A2B receptors prevented the effects of 2-chloroadenosine on ERK1/2, Akt, Skp2, p27(Kip1), cyclin D1, cyclin A, and proliferation. Likewise, inhibition of adenylyl cyclase and protein kinase A abrogated 2-chloroadenosine's inhibitory effects on Skp2 and stimulatory effects on p27(Kip1) and rescued HCASMCs from 2-chloroadenosine-mediated inhibition. Knockdown of p27(Kip1) also reversed the inhibitory effects of 2-chloroadenosine on HCASMC proliferation. In vivo, peri-arterial (rat carotid artery) 2-chloroadenosine (20 μmol/L for 7 days) downregulated vascular expression of Skp2, upregulated vascular expression of p27(Kip1), and reduced neointima hyperplasia by 71% (P<0.05; neointimal thickness: control, 37 424±18 371 pixels; treated, 10 352±2824 pixels). In conclusion, the adenosine/A2B receptor/cAMP/protein kinase A axis inhibits HCASMC proliferation by blocking multiple signaling pathways (ERK1/2, Akt, and Skp2) that converge at cyclin D, a key G1 cyclin that controls cell-cycle progression.
authors.affiliation: From the Department of Reproductive Endocrinology, University Hospital Zurich, Switzerland (R.K.D., M.R., B.I.); Zurich Center for Integrative Human Physiology (ZIHP), University of Zurich, Switzerland (R.K.D.); Department of Pharmacology and Chemical Biology, University of Pittsburgh School of Medicine (D.G.G., Z.M., E.K.J.); and Preclinical Pharma Research 68/209, F. Hoffmann-La-Roche, Basel, Switzerland (J.F.). Raghvendra.dubey@usz.ch.
authors.forename: Raghvendra K
authors.initials: RK
authors.lastname: Dubey
authors.affiliation: From the Department of Reproductive Endocrinology, University Hospital Zurich, Switzerland (R.K.D., M.R., B.I.); Zurich Center for Integrative Human Physiology (ZIHP), University of Zurich, Switzerland (R.K.D.); Department of Pharmacology and Chemical Biology, University of Pittsburgh School of Medicine (D.G.G., Z.M., E.K.J.); and Preclinical Pharma Research 68/209, F. Hoffmann-La-Roche, Basel, Switzerland (J.F.).
authors.forename: Jürgen
authors.initials: J
authors.lastname: Fingerle
authors.affiliation: From the Department of Reproductive Endocrinology, University Hospital Zurich, Switzerland (R.K.D., M.R., B.I.); Zurich Center for Integrative Human Physiology (ZIHP), University of Zurich, Switzerland (R.K.D.); Department of Pharmacology and Chemical Biology, University of Pittsburgh School of Medicine (D.G.G., Z.M., E.K.J.); and Preclinical Pharma Research 68/209, F. Hoffmann-La-Roche, Basel, Switzerland (J.F.).
authors.forename: Delbert G
authors.initials: DG
authors.lastname: Gillespie
authors.affiliation: From the Department of Reproductive Endocrinology, University Hospital Zurich, Switzerland (R.K.D., M.R., B.I.); Zurich Center for Integrative Human Physiology (ZIHP), University of Zurich, Switzerland (R.K.D.); Department of Pharmacology and Chemical Biology, University of Pittsburgh School of Medicine (D.G.G., Z.M., E.K.J.); and Preclinical Pharma Research 68/209, F. Hoffmann-La-Roche, Basel, Switzerland (J.F.).
authors.forename: Zaichuan
authors.initials: Z
authors.lastname: Mi
authors.affiliation: From the Department of Reproductive Endocrinology, University Hospital Zurich, Switzerland (R.K.D., M.R., B.I.); Zurich Center for Integrative Human Physiology (ZIHP), University of Zurich, Switzerland (R.K.D.); Department of Pharmacology and Chemical Biology, University of Pittsburgh School of Medicine (D.G.G., Z.M., E.K.J.); and Preclinical Pharma Research 68/209, F. Hoffmann-La-Roche, Basel, Switzerland (J.F.).
authors.forename: Marinella
authors.initials: M
authors.lastname: Rosselli
authors.affiliation: From the Department of Reproductive Endocrinology, University Hospital Zurich, Switzerland (R.K.D., M.R., B.I.); Zurich Center for Integrative Human Physiology (ZIHP), University of Zurich, Switzerland (R.K.D.); Department of Pharmacology and Chemical Biology, University of Pittsburgh School of Medicine (D.G.G., Z.M., E.K.J.); and Preclinical Pharma Research 68/209, F. Hoffmann-La-Roche, Basel, Switzerland (J.F.).
authors.forename: Bruno
authors.initials: B
authors.lastname: Imthurn
authors.affiliation: From the Department of Reproductive Endocrinology, University Hospital Zurich, Switzerland (R.K.D., M.R., B.I.); Zurich Center for Integrative Human Physiology (ZIHP), University of Zurich, Switzerland (R.K.D.); Department of Pharmacology and Chemical Biology, University of Pittsburgh School of Medicine (D.G.G., Z.M., E.K.J.); and Preclinical Pharma Research 68/209, F. Hoffmann-La-Roche, Basel, Switzerland (J.F.).
authors.forename: Edwin K
authors.initials: EK
authors.lastname: Jackson
mesh_terms: D015762:2-Chloroadenosine / Q000494:pharmacology; D000241:Adenosine / Q000494:pharmacology*; D000818:Animals; D015153:Blotting, Western; D049109:Cell Proliferation / Q000187:drug effects* / Q000235:genetics; D002478:Cells, Cultured; D003331:Coronary Vessels / Q000166:cytology; D000242:Cyclic AMP / Q000378:metabolism; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism; D056741:Cyclin D / Q000235:genetics / Q000378:metabolism*; D050760:Cyclin-Dependent Kinase Inhibitor p27 / Q000235:genetics / Q000378:metabolism; D048049:Extracellular Signal-Regulated MAP Kinases / Q000378:metabolism; D015870:Gene Expression / Q000187:drug effects; D006801:Humans; D008297:Male; D032389:Myocytes, Smooth Muscle / Q000187:drug effects* / Q000378:metabolism; D034622:RNA Interference; D011921:Rats, Inbred WKY; D043682:Receptor, Adenosine A1 / Q000235:genetics / Q000378:metabolism; D043704:Receptor, Adenosine A2B / Q000235:genetics / Q000378:metabolism; D044786:S-Phase Kinase-Associated Proteins / Q000235:genetics / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects* / Q000235:genetics
chemical_list: D056741:Cyclin D; D043682:Receptor, Adenosine A1; D043704:Receptor, Adenosine A2B; D044786:S-Phase Kinase-Associated Proteins; D015762:2-Chloroadenosine; D050760:Cyclin-Dependent Kinase Inhibitor p27; D000242:Cyclic AMP; D017868:Cyclic AMP-Dependent Protein Kinases; D048049:Extracellular Signal-Regulated MAP Kinases; D000241:Adenosine
keywords: A2B receptor; Skp2; adenosine; cyclin D1; p27Kip1; vascular smooth muscle cells