Transforming growth factor-beta 1 inhibits generation of angiostatin by human pancreatic cancer cells

Surgery. 1998 Aug;124(2):388-93.

Abstract

Background: Angiostatin, a proteolytic fragment of plasminogen, is a potent inhibitor of angiogenesis. We have previously shown that the human pancreatic cancer cell line ASPC-1 produces enzymatic activity capable of generating angiostatin. In this study we sought to determine whether angiostatin production by ASPC-1 cells was regulated by the growth factor transforming growth factor-beta 1 (TGF-beta 1), a key mediator of tumor angiogenesis.

Methods: ASPC-1 cells were grown to 70% to 80% confluence in 20% fetal calf serum-RPMI. Medium was changed to serum free. TGF-beta 1 was added at concentrations of 0, 1, 5, and 10 ng/mL with or without plasminogen activator inhibitor type-1 (PAI-1) at concentrations of 0, 5, 10, 50, and 100 micrograms/mL. Cells were then cultured for an additional 24 hours. The serum-free conditioned medium was obtained. Angiostatin generation was determined by incubating 20 micrograms of plasminogen with 100 microL of serum-free conditioned medium for 0, 1, 2, 3, 6, 12, and 24 hours. Samples were run on 12% sodium dodecyl sulfate-polyacrylamide gel electrophoresis and transferred. The membrane was probed with a monoclonal antibody to the kringle 1-3 fragment of plasminogen and developed using enhanced chemiluminescence.

Results: TGF-beta 1 and PAI-1 inhibited the conversion of plasminogen into angiostatin in a time- and dose-dependent manner. Antibody to PAI-1 completely blocks TGF-beta 1 mediated angiostatin inhibition.

Conclusions: TGF-beta 1 inhibits the generation of the antiangiogenic molecule angiostatin by human pancreatic cancer cells in a time- and dose-dependent manner. This effect is mediated through modulation of the plasminogen/plasmin system.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Adenocarcinoma
  • Angiostatins
  • Antibodies / pharmacology
  • Antineoplastic Agents / analysis
  • Antineoplastic Agents / metabolism*
  • Blotting, Western
  • Dose-Response Relationship, Drug
  • Humans
  • Neovascularization, Pathologic / enzymology
  • Neovascularization, Pathologic / physiopathology
  • Pancreatic Neoplasms*
  • Peptide Fragments / analysis
  • Peptide Fragments / biosynthesis*
  • Peptide Fragments / metabolism
  • Plasminogen / analysis
  • Plasminogen / biosynthesis*
  • Plasminogen / metabolism
  • Plasminogen Activator Inhibitor 1 / immunology
  • Plasminogen Activator Inhibitor 1 / pharmacology
  • Serine Proteinase Inhibitors / immunology
  • Serine Proteinase Inhibitors / pharmacology
  • Transforming Growth Factor beta / pharmacology*
  • Tumor Cells, Cultured / drug effects
  • Tumor Cells, Cultured / metabolism

Substances

  • Antibodies
  • Antineoplastic Agents
  • Peptide Fragments
  • Plasminogen Activator Inhibitor 1
  • Serine Proteinase Inhibitors
  • Transforming Growth Factor beta
  • Angiostatins
  • Plasminogen