Plasminogen activators in human breast cancer cell lines: Hormonal regulation and properties

Walter F. Mangel, Diana L. Toledo, Ann M. Nardulli, Georg C.A. Reiner, Mary J. Norman, Benita S. Katzenellenbogen

Research output: Contribution to journalArticle

Abstract

To understand the hormonal regulation of plasminogen activators (PAs) in human breast cancer, we have examined the hormonal regulation and properties of PAs in four human breast cancer cell lines that differ markedly in their estrogen receptor (ER) content: MCF-7 cells contain high levels of ER (approx 7 pmol/mg DNA) and their PA activity was increased 3-4-fold by physiological concentrations of estradiol; T47-D and ZR-75-1 cells contain lower levels of ER (0.9 and 2.1 pmol/mg DNA respectively) and their PA activity was also increased 3-4-fold by estradiol. In contrast, MDA-MB-231 cells, which do not contain ER, showed a high level of PA activity that was not modulated by estradiol. SDS-PAGE followed by zymography indicated that MCF-7 cells secreted tissue-type PA (t-PA), T47-D and ZR-75-1 cells secreted urokinase-type PA (u-PA), and MDA-MB-231 cells secreted both types of PAs. The types of PAs secreted by these cell lines did not change upon treatment with estradiol. Dose-response curves for the stimulation of MCF-7 PA activity by different estrogens showed an excellent correlation between affinities of the estrogens for ER and their potency in stimulating PA activity. With a clonal subline of MCF-7 cells, MCF-L, a soluble inhibitor of both t-PA and u-PA was secreted. Incubation of purified t-PA or u-PA with the serum-free conditioned medium from MCF-L cells resulted in a shift in the mobility of t-PA and u-PA in SDS-polyacrylamide gels to forms increased in molecular mass by about 50,000-70,000. The shifts in molecular mass could be prevented by the presence of the competitive inhibitor p-aminobenzamidine, indicating that the active sites of the PAs were involved in the formation of these complexes. Furthermore, co-cultivation, of RT4-D rat neuroblastoma cells, which exhibit high levels of t-PA activity, with MCF-L cells resulted in a marked decrease in the PA activity of the RT4-D cells. Our results were consistent with the following conclusions: t-PA, u-PA or both were secreted by human breast cancer cells. In the ER-containing cell lines, depending upon the specific cell line, t-PA or u-PA was stimulated by estrogens. The unstimulated levels of PA activity and the magnitude of PA stimulation by estrogens were not closely related to ER content. However, the correlation between stimulation of PA activity by estrogen and the presence of ER in human breast cancer cells suggests that stimulation of PA activity may be a good marker for estrogenic activity, reflecting a functional receptor system capable of mediating estrogen action, and that an assay for stimulation of PA activity would better measure hormone responsiveness and be more predictive of the benefit of endocrine therapy than would be an assay of absolute levels of PA activity.

Original languageEnglish (US)
Pages (from-to)79-88
Number of pages10
JournalJournal of Steroid Biochemistry
Volume30
Issue number1-6
DOIs
StatePublished - 1988

Fingerprint

Plasminogen Activators
Cells
Breast Neoplasms
Cell Line
Estrogen Receptors
Urokinase-Type Plasminogen Activator
Estrogens
Estradiol
MCF-7 Cells
Molecular mass
Assays
Somatostatin-Secreting Cells
DNA
Serum-Free Culture Media
Tissue Plasminogen Activator
Conditioned Culture Medium
Neuroblastoma

ASJC Scopus subject areas

  • Biochemistry
  • Endocrinology

Cite this

Plasminogen activators in human breast cancer cell lines : Hormonal regulation and properties. / Mangel, Walter F.; Toledo, Diana L.; Nardulli, Ann M.; Reiner, Georg C.A.; Norman, Mary J.; Katzenellenbogen, Benita S.

In: Journal of Steroid Biochemistry, Vol. 30, No. 1-6, 1988, p. 79-88.

Research output: Contribution to journalArticle

Mangel, Walter F. ; Toledo, Diana L. ; Nardulli, Ann M. ; Reiner, Georg C.A. ; Norman, Mary J. ; Katzenellenbogen, Benita S. / Plasminogen activators in human breast cancer cell lines : Hormonal regulation and properties. In: Journal of Steroid Biochemistry. 1988 ; Vol. 30, No. 1-6. pp. 79-88.
@article{7784af8cc58743c2b49897616d0a2f56,
title = "Plasminogen activators in human breast cancer cell lines: Hormonal regulation and properties",
abstract = "To understand the hormonal regulation of plasminogen activators (PAs) in human breast cancer, we have examined the hormonal regulation and properties of PAs in four human breast cancer cell lines that differ markedly in their estrogen receptor (ER) content: MCF-7 cells contain high levels of ER (approx 7 pmol/mg DNA) and their PA activity was increased 3-4-fold by physiological concentrations of estradiol; T47-D and ZR-75-1 cells contain lower levels of ER (0.9 and 2.1 pmol/mg DNA respectively) and their PA activity was also increased 3-4-fold by estradiol. In contrast, MDA-MB-231 cells, which do not contain ER, showed a high level of PA activity that was not modulated by estradiol. SDS-PAGE followed by zymography indicated that MCF-7 cells secreted tissue-type PA (t-PA), T47-D and ZR-75-1 cells secreted urokinase-type PA (u-PA), and MDA-MB-231 cells secreted both types of PAs. The types of PAs secreted by these cell lines did not change upon treatment with estradiol. Dose-response curves for the stimulation of MCF-7 PA activity by different estrogens showed an excellent correlation between affinities of the estrogens for ER and their potency in stimulating PA activity. With a clonal subline of MCF-7 cells, MCF-L, a soluble inhibitor of both t-PA and u-PA was secreted. Incubation of purified t-PA or u-PA with the serum-free conditioned medium from MCF-L cells resulted in a shift in the mobility of t-PA and u-PA in SDS-polyacrylamide gels to forms increased in molecular mass by about 50,000-70,000. The shifts in molecular mass could be prevented by the presence of the competitive inhibitor p-aminobenzamidine, indicating that the active sites of the PAs were involved in the formation of these complexes. Furthermore, co-cultivation, of RT4-D rat neuroblastoma cells, which exhibit high levels of t-PA activity, with MCF-L cells resulted in a marked decrease in the PA activity of the RT4-D cells. Our results were consistent with the following conclusions: t-PA, u-PA or both were secreted by human breast cancer cells. In the ER-containing cell lines, depending upon the specific cell line, t-PA or u-PA was stimulated by estrogens. The unstimulated levels of PA activity and the magnitude of PA stimulation by estrogens were not closely related to ER content. However, the correlation between stimulation of PA activity by estrogen and the presence of ER in human breast cancer cells suggests that stimulation of PA activity may be a good marker for estrogenic activity, reflecting a functional receptor system capable of mediating estrogen action, and that an assay for stimulation of PA activity would better measure hormone responsiveness and be more predictive of the benefit of endocrine therapy than would be an assay of absolute levels of PA activity.",
author = "Mangel, {Walter F.} and Toledo, {Diana L.} and Nardulli, {Ann M.} and Reiner, {Georg C.A.} and Norman, {Mary J.} and Katzenellenbogen, {Benita S.}",
year = "1988",
doi = "10.1016/0022-4731(88)90079-9",
language = "English (US)",
volume = "30",
pages = "79--88",
journal = "Journal of Steroid Biochemistry and Molecular Biology",
issn = "0960-0760",
publisher = "Elsevier Limited",
number = "1-6",

}

TY - JOUR

T1 - Plasminogen activators in human breast cancer cell lines

T2 - Hormonal regulation and properties

AU - Mangel, Walter F.

AU - Toledo, Diana L.

AU - Nardulli, Ann M.

AU - Reiner, Georg C.A.

AU - Norman, Mary J.

AU - Katzenellenbogen, Benita S.

PY - 1988

Y1 - 1988

N2 - To understand the hormonal regulation of plasminogen activators (PAs) in human breast cancer, we have examined the hormonal regulation and properties of PAs in four human breast cancer cell lines that differ markedly in their estrogen receptor (ER) content: MCF-7 cells contain high levels of ER (approx 7 pmol/mg DNA) and their PA activity was increased 3-4-fold by physiological concentrations of estradiol; T47-D and ZR-75-1 cells contain lower levels of ER (0.9 and 2.1 pmol/mg DNA respectively) and their PA activity was also increased 3-4-fold by estradiol. In contrast, MDA-MB-231 cells, which do not contain ER, showed a high level of PA activity that was not modulated by estradiol. SDS-PAGE followed by zymography indicated that MCF-7 cells secreted tissue-type PA (t-PA), T47-D and ZR-75-1 cells secreted urokinase-type PA (u-PA), and MDA-MB-231 cells secreted both types of PAs. The types of PAs secreted by these cell lines did not change upon treatment with estradiol. Dose-response curves for the stimulation of MCF-7 PA activity by different estrogens showed an excellent correlation between affinities of the estrogens for ER and their potency in stimulating PA activity. With a clonal subline of MCF-7 cells, MCF-L, a soluble inhibitor of both t-PA and u-PA was secreted. Incubation of purified t-PA or u-PA with the serum-free conditioned medium from MCF-L cells resulted in a shift in the mobility of t-PA and u-PA in SDS-polyacrylamide gels to forms increased in molecular mass by about 50,000-70,000. The shifts in molecular mass could be prevented by the presence of the competitive inhibitor p-aminobenzamidine, indicating that the active sites of the PAs were involved in the formation of these complexes. Furthermore, co-cultivation, of RT4-D rat neuroblastoma cells, which exhibit high levels of t-PA activity, with MCF-L cells resulted in a marked decrease in the PA activity of the RT4-D cells. Our results were consistent with the following conclusions: t-PA, u-PA or both were secreted by human breast cancer cells. In the ER-containing cell lines, depending upon the specific cell line, t-PA or u-PA was stimulated by estrogens. The unstimulated levels of PA activity and the magnitude of PA stimulation by estrogens were not closely related to ER content. However, the correlation between stimulation of PA activity by estrogen and the presence of ER in human breast cancer cells suggests that stimulation of PA activity may be a good marker for estrogenic activity, reflecting a functional receptor system capable of mediating estrogen action, and that an assay for stimulation of PA activity would better measure hormone responsiveness and be more predictive of the benefit of endocrine therapy than would be an assay of absolute levels of PA activity.

AB - To understand the hormonal regulation of plasminogen activators (PAs) in human breast cancer, we have examined the hormonal regulation and properties of PAs in four human breast cancer cell lines that differ markedly in their estrogen receptor (ER) content: MCF-7 cells contain high levels of ER (approx 7 pmol/mg DNA) and their PA activity was increased 3-4-fold by physiological concentrations of estradiol; T47-D and ZR-75-1 cells contain lower levels of ER (0.9 and 2.1 pmol/mg DNA respectively) and their PA activity was also increased 3-4-fold by estradiol. In contrast, MDA-MB-231 cells, which do not contain ER, showed a high level of PA activity that was not modulated by estradiol. SDS-PAGE followed by zymography indicated that MCF-7 cells secreted tissue-type PA (t-PA), T47-D and ZR-75-1 cells secreted urokinase-type PA (u-PA), and MDA-MB-231 cells secreted both types of PAs. The types of PAs secreted by these cell lines did not change upon treatment with estradiol. Dose-response curves for the stimulation of MCF-7 PA activity by different estrogens showed an excellent correlation between affinities of the estrogens for ER and their potency in stimulating PA activity. With a clonal subline of MCF-7 cells, MCF-L, a soluble inhibitor of both t-PA and u-PA was secreted. Incubation of purified t-PA or u-PA with the serum-free conditioned medium from MCF-L cells resulted in a shift in the mobility of t-PA and u-PA in SDS-polyacrylamide gels to forms increased in molecular mass by about 50,000-70,000. The shifts in molecular mass could be prevented by the presence of the competitive inhibitor p-aminobenzamidine, indicating that the active sites of the PAs were involved in the formation of these complexes. Furthermore, co-cultivation, of RT4-D rat neuroblastoma cells, which exhibit high levels of t-PA activity, with MCF-L cells resulted in a marked decrease in the PA activity of the RT4-D cells. Our results were consistent with the following conclusions: t-PA, u-PA or both were secreted by human breast cancer cells. In the ER-containing cell lines, depending upon the specific cell line, t-PA or u-PA was stimulated by estrogens. The unstimulated levels of PA activity and the magnitude of PA stimulation by estrogens were not closely related to ER content. However, the correlation between stimulation of PA activity by estrogen and the presence of ER in human breast cancer cells suggests that stimulation of PA activity may be a good marker for estrogenic activity, reflecting a functional receptor system capable of mediating estrogen action, and that an assay for stimulation of PA activity would better measure hormone responsiveness and be more predictive of the benefit of endocrine therapy than would be an assay of absolute levels of PA activity.

UR - http://www.scopus.com/inward/record.url?scp=0023801178&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=0023801178&partnerID=8YFLogxK

U2 - 10.1016/0022-4731(88)90079-9

DO - 10.1016/0022-4731(88)90079-9

M3 - Article

C2 - 3386280

AN - SCOPUS:0023801178

VL - 30

SP - 79

EP - 88

JO - Journal of Steroid Biochemistry and Molecular Biology

JF - Journal of Steroid Biochemistry and Molecular Biology

SN - 0960-0760

IS - 1-6

ER -