title: 5-fluorouracil causes endothelial cell senescence: potential protective role of glucagon-like peptide 1.
abstract: OBJECTIVE
  5-fluorouracil (5FU) and its prodrug, capecitabine, can damage endothelial cells, whilst endothelial integrity is preserved by glucagon-like peptide 1 (GLP-1). Here, we studied the effect of 5FU on endothelial senescence and whether GLP-1 antagonizes it.
  
  
  METHODS
  EA.hy926 cells were exposed to 5FU or sera from patients taking capecitabine, with or without pre-incubation with GLP-1. Senescence was identified by expression of senescence-associated β-galactosidase and p16INK4aand reduced cell proliferation. Soluble vascular cell adhesion molecule-1 (sVCAM-1), soluble intercellular adhesion molecule-1 (sICAM-1) and CD146 (marker of endothelial injury) were measured by ELISA before and at completion of capecitabine chemotherapy. RT-PCR, western blotting, functional experiments with signalling inhibitors and ERK1/2 silencing were performed to characterize 5FU-induced phenotype and elucidate the pathways underlying 5FU and GLP-1 activity.
  
  
  RESULTS
  Both 5FU and sera from capecitabine-treated patients stimulated endothelial cell senescence. 5FU-elicited senescence occurred via activation of p38 and JNK, and was associated with decreased eNOS and SIRT-1 levels. Furthermore, 5FU up-regulated VCAM1 and TYMP (encodes enzyme activating capecitabine and 5FU), and sVCAM-1 and CD146 concentrations were higher after than before capecitabine chemotherapy. A non-significant trend for higher ICAM1 levels was also observed. GLP-1 counteracted 5FU-initiated senescence and reduced eNOS and SIRT-1 expression, this protection being mediated by GLP-1 receptor, ERK1/2 and, possibly, PKA and PI3K.
  
  
  CONCLUSIONS
  5FU causes endothelial cell senescence and dysfunction, which may contribute to its cardiovascular side effects. 5FU-triggered senescence was prevented by GLP-1, raising the possibility of using GLP-1 analogues and degradation inhibitors to treat 5FU and capecitabine vascular toxicity.
  
  
  BACKGROUND
  This article is part of a themed section on New Insights into Cardiotoxicity Caused by Chemotherapeutic Agents. To view the other articles in this section visit http://onlinelibrary.wiley.com/doi/10.1111/bph.v174.21/issuetoc.
authors.affiliation: Laboratory of Cardiovascular Biology, Department of Internal Medicine, University of Genova, Genova, Italy.
authors.forename: Paola
authors.initials: P
authors.lastname: Altieri
authors.affiliation: Oncology Unit, IRCCS AOU San Martino-IST, Genova, Italy.
authors.forename: Roberto
authors.initials: R
authors.lastname: Murialdo
authors.affiliation: Laboratory of Cardiovascular Biology, Department of Internal Medicine, University of Genova, Genova, Italy.
authors.forename: Chiara
authors.initials: C
authors.lastname: Barisione
authors.affiliation: Laboratory of Cardiovascular Biology, Department of Internal Medicine, University of Genova, Genova, Italy.
authors.forename: Edoardo
authors.initials: E
authors.lastname: Lazzarini
authors.affiliation: Laboratory of Cardiovascular Biology, Department of Internal Medicine, University of Genova, Genova, Italy.
authors.forename: Silvano
authors.initials: S
authors.lastname: Garibaldi
authors.affiliation: Laboratory of Cardiovascular Biology, Department of Internal Medicine, University of Genova, Genova, Italy.
authors.forename: Patrizia
authors.initials: P
authors.lastname: Fabbi
authors.affiliation: Laboratory of Cardiovascular Biology, Department of Internal Medicine, University of Genova, Genova, Italy.
authors.forename: Clarissa
authors.initials: C
authors.lastname: Ruggeri
authors.affiliation: Cardiovascular Disease Unit, IRCCS AOU San Martino-IST, Genova, Italy.
authors.forename: Silvia
authors.initials: S
authors.lastname: Borile
authors.affiliation: First Clinic of Internal Medicine, IRCCS AOU San Martino - IST, Genova, Italy.|Department of Internal Medicine, University of Genova, Genova, Italy.
authors.forename: Federico
authors.identifier: http://orcid.org/0000-0003-2957-4078
authors.initials: F
authors.lastname: Carbone
authors.affiliation: Drug Discovery and Development Department, Italian Institute of Technology (IIT), Genova, Italy.
authors.forename: Andrea
authors.initials: A
authors.lastname: Armirotti
authors.affiliation: Laboratory of Cardiovascular Biology, Department of Internal Medicine, University of Genova, Genova, Italy.|Cardiovascular Disease Unit, IRCCS AOU San Martino-IST, Genova, Italy.
authors.forename: Marco
authors.initials: M
authors.lastname: Canepa
authors.affiliation: Oncology Unit, IRCCS AOU San Martino-IST, Genova, Italy.
authors.forename: Alberto
authors.initials: A
authors.lastname: Ballestrero
authors.affiliation: Laboratory of Cardiovascular Biology, Department of Internal Medicine, University of Genova, Genova, Italy.|Cardiovascular Disease Unit, IRCCS AOU San Martino-IST, Genova, Italy.
authors.forename: Claudio
authors.initials: C
authors.lastname: Brunelli
authors.affiliation: First Clinic of Internal Medicine, IRCCS AOU San Martino - IST, Genova, Italy.|Department of Internal Medicine, University of Genova, Genova, Italy.|Centre of Excellence for Biomedical Research (CEBR), University of Genova, Genova, Italy.
authors.forename: Fabrizio
authors.initials: F
authors.lastname: Montecucco
authors.affiliation: Laboratory of Cardiovascular Biology, Department of Internal Medicine, University of Genova, Genova, Italy.|Cardiovascular Disease Unit, IRCCS AOU San Martino-IST, Genova, Italy.
authors.forename: Pietro
authors.identifier: http://orcid.org/0000-0001-7167-7287
authors.initials: P
authors.lastname: Ameri
authors.affiliation: Laboratory of Cardiovascular Biology, Department of Internal Medicine, University of Genova, Genova, Italy.|Cardiovascular Disease Unit, IRCCS AOU San Martino-IST, Genova, Italy.
authors.forename: Paolo
authors.initials: P
authors.lastname: Spallarossa
mesh_terms: D000368:Aged; D000964:Antimetabolites, Antineoplastic / Q000633:toxicity; D015153:Blotting, Western; D000069287:Capecitabine / Q000008:administration & dosage*; D002460:Cell Line; D016922:Cellular Senescence / Q000187:drug effects*; D042783:Endothelial Cells / Q000187:drug effects* / Q000473:pathology; D004797:Enzyme-Linked Immunosorbent Assay; D005260:Female; D005472:Fluorouracil / Q000633:toxicity*; D052216:Glucagon-Like Peptide 1 / Q000008:administration & dosage* / Q000494:pharmacology; D006801:Humans; D008297:Male; D020133:Reverse Transcriptase Polymerase Chain Reaction
chemical_list: D000964:Antimetabolites, Antineoplastic; D000069287:Capecitabine; D052216:Glucagon-Like Peptide 1; D005472:Fluorouracil