Angiostatin generation by human pancreatic cancer

Christine A. O'Mahony, Amy Seidel, Daniel Albo, Hong Chang, George P. Tuszynski, David H. Berger

Research output: Contribution to journalArticle

35 Citations (Scopus)

Abstract

Background. Angiostatin, a proteolytic fragment of plasminogen, is a potent inhibitor of angiogenesis. In vitro, angiostatin can be generated by pancreatic elastase proteolysis of plasminogen; however, in vivo, the enzymes responsible for angiostatin production are not known. A recent study demonstrates the involvement of a serine protease in angiostatin generation. In this study we sought to determine if the human pancreatic carcinoma cell line ASPC1 produced enzymatic activity capable of converting plasminogen to angiostatin and to determine if urokinase plasminogen activator (uPA) is involved in this system. Methods. ASPC1 cells were grown to near confluence in 20% FBS-RPMI. Media were changed to serum free and cells cultured for an additional 24 h. The serum free conditioned media (SFCM) was obtained. Angiostatin generation was determined by incubating 20 μg of human plasminogen with 100 μl of SFCM for 0, 3, 8, 12, 24, and 48 h. Plasminogen cleavage was assessed in the presence of the following protease inhibitors: pefabloc, aprotinin, phosphoramidon, leupeptin, and EDTA. The effect of uPA on angiostatin generation was determined by incubating plasminogen with antibody to uPA. Angiostatin generation was determined by Western blot. Results. Incubation of plasminogen with SFCM resulted in the generation of immunoreactive bands at 48 kDa corresponding to human angiostatin. Angiostatin generation by ASPC1 SFCM was time dependent; there was a significant decrease in the plasminogen substrate beginning at 3 h with complete conversion to angiostatin by 48 h. Enzymatic activity leading to angiostatin production was found to be due to a serine protease. Antibody to uPA effectively blocked angiostatin production by ASPC1 SFCM in a dose- dependent manner. Conclusion. Human pancreatic cancer cells express enzymatic activity which leads to the generation of angiostatin. Conversion of plasminogen to angiostatin is due to a serine protease. This serine protease is most likely uPA.

Original languageEnglish (US)
Pages (from-to)55-58
Number of pages4
JournalJournal of Surgical Research
Volume77
Issue number1
DOIs
StatePublished - Jan 1 1998
Externally publishedYes

Fingerprint

Angiostatins
Pancreatic Neoplasms
Plasminogen
Plasminogen Activators
Serum-Free Culture Media
Urokinase-Type Plasminogen Activator
Conditioned Culture Medium
Serine Proteases
Aprotinin
Angiogenesis Inhibitors
Antibodies

Keywords

  • Angiostatin
  • Pancreatic neoplasms
  • Plasminogen
  • Urokinase plasminogen activator

ASJC Scopus subject areas

  • Surgery

Cite this

O'Mahony, C. A., Seidel, A., Albo, D., Chang, H., Tuszynski, G. P., & Berger, D. H. (1998). Angiostatin generation by human pancreatic cancer. Journal of Surgical Research, 77(1), 55-58. https://doi.org/10.1006/jsre.1998.5334

Angiostatin generation by human pancreatic cancer. / O'Mahony, Christine A.; Seidel, Amy; Albo, Daniel; Chang, Hong; Tuszynski, George P.; Berger, David H.

In: Journal of Surgical Research, Vol. 77, No. 1, 01.01.1998, p. 55-58.

Research output: Contribution to journalArticle

O'Mahony, CA, Seidel, A, Albo, D, Chang, H, Tuszynski, GP & Berger, DH 1998, 'Angiostatin generation by human pancreatic cancer', Journal of Surgical Research, vol. 77, no. 1, pp. 55-58. https://doi.org/10.1006/jsre.1998.5334
O'Mahony CA, Seidel A, Albo D, Chang H, Tuszynski GP, Berger DH. Angiostatin generation by human pancreatic cancer. Journal of Surgical Research. 1998 Jan 1;77(1):55-58. https://doi.org/10.1006/jsre.1998.5334
O'Mahony, Christine A. ; Seidel, Amy ; Albo, Daniel ; Chang, Hong ; Tuszynski, George P. ; Berger, David H. / Angiostatin generation by human pancreatic cancer. In: Journal of Surgical Research. 1998 ; Vol. 77, No. 1. pp. 55-58.
@article{f0c968a28831407eb1b8ba123296b2e3,
title = "Angiostatin generation by human pancreatic cancer",
abstract = "Background. Angiostatin, a proteolytic fragment of plasminogen, is a potent inhibitor of angiogenesis. In vitro, angiostatin can be generated by pancreatic elastase proteolysis of plasminogen; however, in vivo, the enzymes responsible for angiostatin production are not known. A recent study demonstrates the involvement of a serine protease in angiostatin generation. In this study we sought to determine if the human pancreatic carcinoma cell line ASPC1 produced enzymatic activity capable of converting plasminogen to angiostatin and to determine if urokinase plasminogen activator (uPA) is involved in this system. Methods. ASPC1 cells were grown to near confluence in 20{\%} FBS-RPMI. Media were changed to serum free and cells cultured for an additional 24 h. The serum free conditioned media (SFCM) was obtained. Angiostatin generation was determined by incubating 20 μg of human plasminogen with 100 μl of SFCM for 0, 3, 8, 12, 24, and 48 h. Plasminogen cleavage was assessed in the presence of the following protease inhibitors: pefabloc, aprotinin, phosphoramidon, leupeptin, and EDTA. The effect of uPA on angiostatin generation was determined by incubating plasminogen with antibody to uPA. Angiostatin generation was determined by Western blot. Results. Incubation of plasminogen with SFCM resulted in the generation of immunoreactive bands at 48 kDa corresponding to human angiostatin. Angiostatin generation by ASPC1 SFCM was time dependent; there was a significant decrease in the plasminogen substrate beginning at 3 h with complete conversion to angiostatin by 48 h. Enzymatic activity leading to angiostatin production was found to be due to a serine protease. Antibody to uPA effectively blocked angiostatin production by ASPC1 SFCM in a dose- dependent manner. Conclusion. Human pancreatic cancer cells express enzymatic activity which leads to the generation of angiostatin. Conversion of plasminogen to angiostatin is due to a serine protease. This serine protease is most likely uPA.",
keywords = "Angiostatin, Pancreatic neoplasms, Plasminogen, Urokinase plasminogen activator",
author = "O'Mahony, {Christine A.} and Amy Seidel and Daniel Albo and Hong Chang and Tuszynski, {George P.} and Berger, {David H.}",
year = "1998",
month = "1",
day = "1",
doi = "10.1006/jsre.1998.5334",
language = "English (US)",
volume = "77",
pages = "55--58",
journal = "Journal of Surgical Research",
issn = "0022-4804",
publisher = "Academic Press Inc.",
number = "1",

}

TY - JOUR

T1 - Angiostatin generation by human pancreatic cancer

AU - O'Mahony, Christine A.

AU - Seidel, Amy

AU - Albo, Daniel

AU - Chang, Hong

AU - Tuszynski, George P.

AU - Berger, David H.

PY - 1998/1/1

Y1 - 1998/1/1

N2 - Background. Angiostatin, a proteolytic fragment of plasminogen, is a potent inhibitor of angiogenesis. In vitro, angiostatin can be generated by pancreatic elastase proteolysis of plasminogen; however, in vivo, the enzymes responsible for angiostatin production are not known. A recent study demonstrates the involvement of a serine protease in angiostatin generation. In this study we sought to determine if the human pancreatic carcinoma cell line ASPC1 produced enzymatic activity capable of converting plasminogen to angiostatin and to determine if urokinase plasminogen activator (uPA) is involved in this system. Methods. ASPC1 cells were grown to near confluence in 20% FBS-RPMI. Media were changed to serum free and cells cultured for an additional 24 h. The serum free conditioned media (SFCM) was obtained. Angiostatin generation was determined by incubating 20 μg of human plasminogen with 100 μl of SFCM for 0, 3, 8, 12, 24, and 48 h. Plasminogen cleavage was assessed in the presence of the following protease inhibitors: pefabloc, aprotinin, phosphoramidon, leupeptin, and EDTA. The effect of uPA on angiostatin generation was determined by incubating plasminogen with antibody to uPA. Angiostatin generation was determined by Western blot. Results. Incubation of plasminogen with SFCM resulted in the generation of immunoreactive bands at 48 kDa corresponding to human angiostatin. Angiostatin generation by ASPC1 SFCM was time dependent; there was a significant decrease in the plasminogen substrate beginning at 3 h with complete conversion to angiostatin by 48 h. Enzymatic activity leading to angiostatin production was found to be due to a serine protease. Antibody to uPA effectively blocked angiostatin production by ASPC1 SFCM in a dose- dependent manner. Conclusion. Human pancreatic cancer cells express enzymatic activity which leads to the generation of angiostatin. Conversion of plasminogen to angiostatin is due to a serine protease. This serine protease is most likely uPA.

AB - Background. Angiostatin, a proteolytic fragment of plasminogen, is a potent inhibitor of angiogenesis. In vitro, angiostatin can be generated by pancreatic elastase proteolysis of plasminogen; however, in vivo, the enzymes responsible for angiostatin production are not known. A recent study demonstrates the involvement of a serine protease in angiostatin generation. In this study we sought to determine if the human pancreatic carcinoma cell line ASPC1 produced enzymatic activity capable of converting plasminogen to angiostatin and to determine if urokinase plasminogen activator (uPA) is involved in this system. Methods. ASPC1 cells were grown to near confluence in 20% FBS-RPMI. Media were changed to serum free and cells cultured for an additional 24 h. The serum free conditioned media (SFCM) was obtained. Angiostatin generation was determined by incubating 20 μg of human plasminogen with 100 μl of SFCM for 0, 3, 8, 12, 24, and 48 h. Plasminogen cleavage was assessed in the presence of the following protease inhibitors: pefabloc, aprotinin, phosphoramidon, leupeptin, and EDTA. The effect of uPA on angiostatin generation was determined by incubating plasminogen with antibody to uPA. Angiostatin generation was determined by Western blot. Results. Incubation of plasminogen with SFCM resulted in the generation of immunoreactive bands at 48 kDa corresponding to human angiostatin. Angiostatin generation by ASPC1 SFCM was time dependent; there was a significant decrease in the plasminogen substrate beginning at 3 h with complete conversion to angiostatin by 48 h. Enzymatic activity leading to angiostatin production was found to be due to a serine protease. Antibody to uPA effectively blocked angiostatin production by ASPC1 SFCM in a dose- dependent manner. Conclusion. Human pancreatic cancer cells express enzymatic activity which leads to the generation of angiostatin. Conversion of plasminogen to angiostatin is due to a serine protease. This serine protease is most likely uPA.

KW - Angiostatin

KW - Pancreatic neoplasms

KW - Plasminogen

KW - Urokinase plasminogen activator

UR - http://www.scopus.com/inward/record.url?scp=0031880835&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=0031880835&partnerID=8YFLogxK

U2 - 10.1006/jsre.1998.5334

DO - 10.1006/jsre.1998.5334

M3 - Article

C2 - 9698533

AN - SCOPUS:0031880835

VL - 77

SP - 55

EP - 58

JO - Journal of Surgical Research

JF - Journal of Surgical Research

SN - 0022-4804

IS - 1

ER -