title: β Common receptor integrates the erythropoietin signaling in activation of endothelial nitric oxide synthase.
abstract: Erythropoietin (EPO), the key hormone for erythropoiesis, also increases nitric oxide (NO) bioavailability in endothelial cells (ECs), yet the definitive mechanisms are not fully understood. Increasing evidence has demonstrated that β common receptor (βCR) plays a crucial role in EPO-mediated non-hematopoietic effects. We investigated the role of βCR in EPO-induced endothelial NO synthase (eNOS) activation in bovine aortic ECs (BAECs) and the molecular mechanisms involved. Results of confocal microscopy and immunoprecipitation analyses revealed that βCR was colocalized and interacted with EPO receptor (EPOR) in ECs. Inhibition of βCR or EPOR by neutralizing antibodies or small interfering RNA abolished the EPO-induced NO production. Additionally, blockage of βCR abrogated the EPO-induced increase in the phosphorylation of eNOS, Akt, Src, or Janus kinase 2 (JAK2). Immunoprecipitation analysis revealed that treatment with EPO increased the interaction between βCR and eNOS, which was suppressed by inhibition of Src, JAK2, or Akt signaling with specific pharmacological inhibitors. Furthermore, EPO-induced EC proliferation, migration, and tube formation were blocked by pretreatment with βCR antibody and Src, JAK2, or PI3K/Akt inhibitors. Moreover, in vivo experiments showed that EPO increased the level of phosphorylated eNOS, Src, JAK2, and Akt, as well as βCR-eNOS association in aortas and promoted the angiogenesis in Matrigel plug, which was diminished by βCR or EPOR neutralizing antibodies. Our findings suggest that βCR may play an integrative role in the EPO signaling-mediated activation of eNOS in ECs.
authors.affiliation: Department of Physiology, National Yang-Ming University, Taipei, Taiwan.
authors.forename: Kuo-Hui
authors.initials: KH
authors.lastname: Su
authors.forename: Song-Kun
authors.initials: SK
authors.lastname: Shyue
authors.forename: Yu Ru
authors.initials: YR
authors.lastname: Kou
authors.forename: Li-Chieh
authors.initials: LC
authors.lastname: Ching
authors.forename: An-Na
authors.initials: AN
authors.lastname: Chiang
authors.forename: Yuan-Bin
authors.initials: YB
authors.lastname: Yu
authors.forename: Chien-Yu
authors.initials: CY
authors.lastname: Chen
authors.forename: Ching-Chian
authors.initials: CC
authors.lastname: Pan
authors.forename: Tzong-Shyuan
authors.initials: TS
authors.lastname: Lee
mesh_terms: D000818:Animals; D057134:Antibodies, Neutralizing / Q000494:pharmacology; D002417:Cattle; D002465:Cell Movement; D049109:Cell Proliferation; D002478:Cells, Cultured; D053648:Cytokine Receptor Common beta Subunit / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D042783:Endothelial Cells / Q000187:drug effects / Q000201:enzymology*; D004789:Enzyme Activation; D004921:Erythropoietin / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D047468:Immunoprecipitation; D053614:Janus Kinase 2 / Q000037:antagonists & inhibitors / Q000378:metabolism; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D008822:Mice, Transgenic; D018613:Microscopy, Confocal; D018919:Neovascularization, Physiologic; D009569:Nitric Oxide / Q000378:metabolism; D052250:Nitric Oxide Synthase Type III / Q000378:metabolism*; D058539:Phosphatidylinositol 3-Kinase / Q000378:metabolism; D000081082:Phosphoinositide-3 Kinase Inhibitors; D010766:Phosphorylation; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D051057:Proto-Oncogene Proteins c-akt / Q000037:antagonists & inhibitors / Q000378:metabolism; D034622:RNA Interference; D017467:Receptors, Erythropoietin / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D016185:Receptors, Interleukin-3 / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D011994:Recombinant Proteins; D015398:Signal Transduction / Q000187:drug effects; D013997:Time Factors; D014162:Transfection; D019061:src-Family Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism
chemical_list: D057134:Antibodies, Neutralizing; D053648:Cytokine Receptor Common beta Subunit; D000081082:Phosphoinositide-3 Kinase Inhibitors; D047428:Protein Kinase Inhibitors; D017467:Receptors, Erythropoietin; D016185:Receptors, Interleukin-3; D011994:Recombinant Proteins; D004921:Erythropoietin; C072185:Csf2rb2 protein, mouse; D009569:Nitric Oxide; D052250:Nitric Oxide Synthase Type III; C496315:Nos3 protein, mouse; D058539:Phosphatidylinositol 3-Kinase; C507926:Jak2 protein, mouse; D053614:Janus Kinase 2; D019061:src-Family Kinases; D051057:Proto-Oncogene Proteins c-akt