title: P2Y2 receptor activation by nucleotides released from highly metastatic breast cancer cells increases tumor growth and invasion via crosstalk with endothelial cells.
abstract: BACKGROUND
  Extracellular nucleotides are released and detectable in a high concentration within the tumor microenvironment. G protein-coupled P2Y2 nucleotide receptor (P2Y2R) is activated equipotently by adenosine triphosphate (ATP) and uridine 5'-triphosphate (UTP), which mediate proinflammatory responses such as cell migration and proliferation. However, the role of P2Y2R in the process of cancer metastasis remains unclear. This study aimed to determine the role of P2Y2R in the proliferation, migration and invasion of highly metastatic MDA-MB-231 breast cancer cells through crosstalk with endothelial cells (ECs).
  
  
  METHODS
  ATP release and P2Y2R activity between high metastatic breast cancer cell MDA-MB-231 and low metastatic breast cancer cell MCF-7 were compared. Then, the role of P2Y2R on tumor growth and invasion via crosstalk with ECs was examined in vitro, using MDA-MB-231 cells and ECs transfected with control- or P2Y2R-siRNA, and in vivo, using an animal model injected with control-shRNA- or P2Y2R-shRNA-transfected MDA-MB-231 cells.
  
  
  RESULTS
  We found that this highly metastatic breast cancer cell line released higher levels of ATP and showed a higher P2Y2R activity in comparison to a low metastatic breast cancer cell line, MCF-7. In MDA-MB-231 cells, P2Y2R activation by ATP or UTP increased proliferation at 24 or 72 hours, which was abolished by P2Y2R knock-down. In addition, the adhesion of MDA-MB-231 cells to ECs and cell migration were both significantly increased by ATP or UTP through the expression of intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1) in MDA-MB-231 or ECs but not in cells where P2Y2R was knocked down. Furthermore, ATP- or UTP-mediated activation of P2Y2R induced MDA-MB-231 invasion through ECs, increased matrix metalloproteinase-9 (MMP-9) activity and vascular endothelial growth factor (VEGF) production in MDA-MB-231 and induced the phosphorylation of vascular endothelial (VE)-cadherin in ECs. Tumor growth and metastasis to other tissues were dramatically reduced, and body weight was increased in mice injected with P2Y2R-shRNA-transfected MDA-MB-231 cells compared to mice injected with control shRNA-transfected MDA-MB-231 cells.
  
  
  CONCLUSIONS
  This study suggests that P2Y2R may play an important role in cancer metastasis via modulation of the crosstalk between cancer cells and ECs.
authors.affiliation: Department of Pharmacology, School of Medicine, Institute of Health Sciences, Gyeongsang National University, Jinju, Korea. hahaha-_-0001@hamail.net.
authors.forename: Hana
authors.initials: H
authors.lastname: Jin
authors.affiliation: Department of Pharmacology, School of Medicine, Institute of Health Sciences, Gyeongsang National University, Jinju, Korea. eunsoyg@hanmail.net.
authors.forename: So Young
authors.initials: SY
authors.lastname: Eun
authors.affiliation: Department of Pathology, School of Medicine, Institute of Health Sciences, Gyeongsang National University, Jinju, 660-751, Korea. jongsil25@dreamwiz.com.
authors.forename: Jong Sil
authors.initials: JS
authors.lastname: Lee
authors.affiliation: Department of Pharmacology, School of Medicine, Institute of Health Sciences, Gyeongsang National University, Jinju, Korea. parksw@gnu.ac.kr.
authors.forename: Sang Won
authors.initials: SW
authors.lastname: Park
authors.affiliation: Department of Pharmacology, School of Medicine, Institute of Health Sciences, Gyeongsang National University, Jinju, Korea. jhlee@gnu.ac.kr.
authors.forename: Jae Heun
authors.initials: JH
authors.lastname: Lee
authors.affiliation: Department of Pharmacology, School of Medicine, Institute of Health Sciences, Gyeongsang National University, Jinju, Korea. kcchang@gnu.ac.kr.
authors.forename: Ki Churl
authors.initials: KC
authors.lastname: Chang
authors.affiliation: Department of Pharmacology, School of Medicine, Institute of Health Sciences, Gyeongsang National University, Jinju, Korea. hyejungkim@gnu.ac.kr.
authors.forename: Hye Jung
authors.initials: HJ
authors.lastname: Kim
mesh_terms: D000255:Adenosine Triphosphate / Q000502:physiology*; D000818:Animals; D015703:Antigens, CD / Q000378:metabolism; D001943:Breast Neoplasms / Q000378:metabolism / Q000473:pathology*; D015820:Cadherins / Q000378:metabolism; D002450:Cell Communication; D002465:Cell Movement; D049109:Cell Proliferation; D042783:Endothelial Cells / Q000378:metabolism*; D005260:Female; D006801:Humans; D008175:Lung Neoplasms / Q000378:metabolism / Q000556:secondary*; D061986:MCF-7 Cells; D020780:Matrix Metalloproteinase 9 / Q000378:metabolism; D008819:Mice, Nude; D009368:Neoplasm Transplantation; D010766:Phosphorylation; D011499:Protein Processing, Post-Translational; D058473:Receptors, Purinergic P2Y2 / Q000378:metabolism*; D014544:Uridine Triphosphate / Q000502:physiology; D042461:Vascular Endothelial Growth Factor A / Q000096:biosynthesis; D000099152:Cadherin 5
chemical_list: D000255:Adenosine Triphosphate; D015703:Antigens, CD; D015820:Cadherins; D020780:Matrix Metalloproteinase 9; D058473:Receptors, Purinergic P2Y2; D014544:Uridine Triphosphate; D042461:Vascular Endothelial Growth Factor A; D000099152:Cadherin 5; C467484:VEGFA protein, human; C579270:MMP9 protein, human