title: Platelet-derived growth factor-D and Rho GTPases regulate recruitment of cancer-associated fibroblasts in cholangiocarcinoma.
abstract: Cholangiocarcinoma (CCA) is characterized by an abundant stromal reaction. Cancer-associated fibroblasts (CAFs) are pivotal in tumor growth and invasiveness and represent a potential therapeutic target. To understand the mechanisms leading to CAF recruitment in CCA, we studied (1) expression of epithelial-mesenchymal transition (EMT) in surgical CCA specimens and CCA cells, (2) lineage tracking of an enhanced green fluorescent protein (EGFP)-expressing human male CCA cell line (EGI-1) after xenotransplantation into severe-combined-immunodeficient mice, (3) expression of platelet-derived growth factors (PDGFs) and their receptors in vivo and in vitro, (4) secretion of PDGFs by CCA cells, (5) the role of PDGF-D in fibroblast recruitment in vitro, and (6) downstream effectors of PDGF-D signaling. CCA cells expressed several EMT biomarkers, but not alpha smooth muscle actin (α-SMA). Xenotransplanted CCA masses were surrounded and infiltrated by α-SMA-expressing CAFs, which were negative for EGFP and the human Y-probe, but positive for the murine Y-probe. CCA cells were strongly immunoreactive for PDGF-A and -D, whereas CAFs expressed PDGF receptor (PDGFR)β. PDGF-D, a PDGFRβ agonist, was exclusively secreted by cultured CCA cells. Fibroblast migration was potently induced by PDGF-D and CCA conditioned medium and was significantly inhibited by PDGFRβ blockade with Imatinib and by silencing PDGF-D expression in CCA cells. In fibroblasts, PDGF-D activated the Rac1 and Cdc42 Rho GTPases and c-Jun N-terminal kinase (JNK). Selective inhibition of Rho GTPases (particularly Rac1) and of JNK strongly reduced PDGF-D-induced fibroblast migration.
  
  
  CONCLUSIONS
  CCA cells express several mesenchymal markers, but do not transdifferentiate into CAFs. Instead, CCA cells recruit CAFs by secreting PDGF-D, which stimulates fibroblast migration through PDGFRβ and Rho GTPase and JNK activation. Targeting tumor or stroma interactions with inhibitors of the PDGF-D pathway may offer a novel therapeutic approach.
authors.affiliation: Department of Surgery, Oncology and Gastroenterology, University of Padova, Italy.
authors.forename: Massimiliano
authors.initials: M
authors.lastname: Cadamuro
authors.forename: Giorgia
authors.initials: G
authors.lastname: Nardo
authors.forename: Stefano
authors.initials: S
authors.lastname: Indraccolo
authors.forename: Luigi
authors.initials: L
authors.lastname: Dall'olmo
authors.forename: Luisa
authors.initials: L
authors.lastname: Sambado
authors.forename: Lidia
authors.initials: L
authors.lastname: Moserle
authors.forename: Irene
authors.initials: I
authors.lastname: Franceschet
authors.forename: Michele
authors.initials: M
authors.lastname: Colledan
authors.forename: Marco
authors.initials: M
authors.lastname: Massani
authors.forename: Tommaso
authors.initials: T
authors.lastname: Stecca
authors.forename: Nicolò
authors.initials: N
authors.lastname: Bassi
authors.forename: Stuart
authors.initials: S
authors.lastname: Morton
authors.forename: Carlo
authors.initials: C
authors.lastname: Spirli
authors.forename: Romina
authors.initials: R
authors.lastname: Fiorotto
authors.forename: Luca
authors.initials: L
authors.lastname: Fabris
authors.forename: Mario
authors.initials: M
authors.lastname: Strazzabosco
mesh_terms: D000818:Animals; D000970:Antineoplastic Agents / Q000494:pharmacology; D001549:Benzamides / Q000494:pharmacology; D001650:Bile Duct Neoplasms / Q000473:pathology / Q000503:physiopathology*; D001653:Bile Ducts, Intrahepatic; D045744:Cell Line, Tumor; D002465:Cell Movement / Q000187:drug effects / Q000502:physiology*; D049109:Cell Proliferation / Q000187:drug effects; D002478:Cells, Cultured; D018281:Cholangiocarcinoma / Q000473:pathology / Q000503:physiopathology*; D058750:Epithelial-Mesenchymal Transition / Q000502:physiology; D005347:Fibroblasts / Q000473:pathology*; D064593:Heterografts; D006801:Humans; D000068877:Imatinib Mesylate; D066298:In Vitro Techniques; D008222:Lymphokines / Q000502:physiology*; D008297:Male; D051379:Mice; D016513:Mice, SCID; D010879:Piperazines / Q000494:pharmacology; D010982:Platelet-Derived Growth Factor / Q000502:physiology*; D011743:Pyrimidines / Q000494:pharmacology; D015398:Signal Transduction / Q000502:physiology; D020741:rho GTP-Binding Proteins / Q000502:physiology*
chemical_list: D000970:Antineoplastic Agents; D001549:Benzamides; D008222:Lymphokines; C427493:PDGFD protein, human; D010879:Piperazines; D010982:Platelet-Derived Growth Factor; D011743:Pyrimidines; C075278:platelet-derived growth factor A; D000068877:Imatinib Mesylate; D020741:rho GTP-Binding Proteins