NIOSHTIC-2 Publications Search

Difficulty demonstrating estradiol-mediated Erk1/2 phosphorylation in MCF-7 cells.

Brower SL; Roberts JR; Antonini JM; Miller MR

J Steroid Biochem Mol Biol 2005 Sep; 96(5):375-385

https://doi.org/10.1016/j.jsbmb.2005.05.006

20028684

While some studies report that estradiol (E2) activates extracellular-signal regulated kinase (Erk1/2) in MCF-7 breast cancer cells, others report E2 does not activate this signaling pathway. This study attempted to resolve the conflicting reports by investigating experimental variables that could impact Erk1/2 activation using a high through-put assay that quantitatively assessed Erk1/2 phosphorylation. Variables tested included: cell staging and dosing regimes with and without charcoal-stripped serum, different MCF-7 cell sublines and culture densities and several E2 formulations and solvents. Levels of phosphorylated Erk1/2 were normalized to cellular protein rather than to total Erk1/2 protein because an antibody purported to recognize total Erk1/2 preferentially reacted with non-phosphorylated Erk1/2, potentially exaggerating the apparent level of Erk1/2 activation. Dosing MCF-7 cells with E2 containing small amounts of stripped serum induced Erk1/2 phosphorylation; however, this induction was largely attributed to serum factors. E2 administered in serum-free medium did not significantly alter Erk1/2 phosphorylation under any condition tested; immunocytochemistry corroborated this conclusion. While phosphatase inhibitors generally increased Erk1/2 phosphorylation, they did not impact E2-altered Erk1/2 phosphorylation. It remains important to resolve the basis of conflicting reports regarding E2-induced Erk1/2 activation due to the potential importance of this pathway on breast cancer and other processes.

Breast-cancer; Cancer; Quantitative-analysis; Cell-cultures; Cell-culture-techniques; Solvents

Department of Biochemistry and Molecular Pharmacology, Mary Babb Randolph Cancer Center, P.O. Box 9142, West Virginia University Health Sciences Center, Morgantown, WV 26506-9142, USA

JSBBEZ

20050901

Journal Article

mmiller@hsc.wvu.edu

2005

5

0960-0760

HELD

Work Environment and Workforce: Mixed Exposures

The Journal of Steroid Biochemistry and Molecular Biology

WV