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A novel integrin α5β1 binding domain in module 4 of connective tissue growth factor (CCN2/CTGF) promotes adhesion and migration of activated pancreatic stellate cells
  1. R Gao,
  2. D R Brigstock
  1. Center for Cell and Vascular Biology, Children’s Research Institute, Columbus, Ohio 43205, USA
  1. Correspondence to:
    Dr D R Brigstock
    Children’s Research Institute, Room WA2022, 700 Children’s Drive, Columbus, OH 43205, USA; brigstod{at}ccri.net

Abstract

Background: Connective tissue growth factor (CCN2) is upregulated in pancreatic fibrosis and desmoplastic pancreatic tumours. CCN2 interacts with integrin α5β1 on pancreatic stellate cells (PSC) in which it stimulates fibrogenesis, adhesion, migration, and proliferation.

Aim: To determine the structural domain(s) in CCN2 that interact with integrin α5β1 to regulation PSC functions.

Methods: Primary activated rat PSC were tested for their adherence to isoforms of CCN2 comprising modules 1–4 (CCN21–4), modules 3–4 (CCN23–4), module 3 alone (CCN23), or module 4 alone (CCN24). Adhesion studies were performed in the presence of EDTA, divalent cations, anti-integrin α5β1 antibodies, CCN2 synthetic peptides, or heparin, or after pretreatment of the cells with heparinase, chondroitinase, or sodium chlorate. CCN2 integrin α5β1 binding was analysed in cell free systems. The ability of CCN21–4, CCN23–4, or CCN24 to stimulate PSC migration was evaluated in the presence of anti-integrin α5β1 or heparin.

Results: PSC adhesion was stimulated by CCN21–4, CCN23–4, or CCN24 and supported by Mg2+ but not Ca2+. CCN24 supported PSC adhesion or migration were blocked by anti-integrin α5β1 antibodies or by treatment of cells with heparinase or sodium chlorate. A direct interaction between CCN24 and integrin α5β1 was demonstrated in cell free assays. The sequence GVCTDGR in module 4 mediated the binding between CCN24 and integrin α5β1 as well as CCN24 mediated PSC adhesion and migration.

Conclusions: A GVCTDGR sequence in module 4 of CCN2 is a novel integrin α5β1 binding site that is essential for CCN2 stimulated functions in PSC and which represents a new therapeutic target in PSC mediated fibrogenesis.

  • BSA, bovine serum albumin
  • CCN2, connective tissue growth factor
  • ECM, extracellular matrix
  • FN, fibronectin
  • HSC, hepatic stellate cell
  • HSPG, heparan sulphate proteoglycan
  • MMPs, matrix metalloproteases
  • PSC, pancreatic stellate cell
  • TGF-β, transforming growth factor β
  • pancreatitis
  • pancreatic fibrosis
  • fibrogenesis
  • connective tissue growth factor

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Footnotes

  • Published online first 16 December 2005

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