|dc.identifier.citation||Wallace, J., Foy, D., Yousuf, H., Warnock, M. & Gow, I. (2012-03) The Effect of 17-_ Oestradiol, Resveratrol, and Genistein on Na+/H+ Exchange in Breast Cancer Cells Lines, Proceedings of the Nutrition Society, vol. 71.||
|dc.description.abstract||The tumour environment is more acidic than normal (pH 6.8 vs 7.3)(1), and this may aid tumour progression or affect the uptake of drugs.
The extracellular pH may be partly-regulated by cellular sodium/hydrogen exchangers (NHEs), and NHE activation may in turn be
regulated by hormones such as oestradiol(2). Some breast cancers possess receptors for oestradiol, and stimulation or blockade of these
receptors may modulate the cell's ability to regulate pH. We studied this by investigating the effect of oestradiol on the ability of breast
cancer cell lines to regulate pH by NHE. We used MCF-7 (expresses oestrogen receptors a and b) and MDA-MB-231 (expresses only
oestrogen receptor b) cell lines. Given the interaction of phytoestrogens with the oestrogen receptor(3), we also looked at the effect of
resveratrol (RSV) or genistein (Gen).
MCF-7 and MDA-MB-231 cells were used either as suspensions, or on coverslips, in the absence of added oestradiol. Cells were loaded
with H+ -sensitive fluorescent probe BCECF, then washed to remove excess dye. Suspended cells were allowed to adhere to the glass
bottom of the 35mm perfusion chamber, coverslips with cells were added to the chamber directly. Cells were perfused (1 ml/min) at 37C
with HEPES-buffed Tyrode (pH 7.4) to which drugs were added as required. Acidification was induced by the NH4Cl pre-pulse technique,
recovery was achieved by returning to normal Tyrode. Cells were illuminated by alternating wavelengths of 440 and 503 nm light (2 Hz
sample rate), and fluorescent light (>535 nm) was captured by a CCD camera. The fluorescence ratios are directly proportional to the H+
concentration. Drugs were added directly to the Tyrode, and since RSV and Gen were dissolved in DMSO, DMSO (0.01% final) was
added to the pre-perfusing Tyrode in those experiments, and used as a control in the absence of drugs.
Results are mean ratio valuesSEM or slopes of rate of change of ratio/minSEM for three separate experimental days, except for the
DMSO controls where n = 6. Differences were assessed by Student's paired or unpaired t tests. A brief (4 min) exposure of breast cancer
cells to NH4Cl (20mM) caused a transient alkalisation (decrease in 440/503 ratio, i.e. decrease in [H + ]i) followed by an acidification and
recovery after ammonium removal. In MDA cells, addition of 17b-oestradiol (E2; 1 pM final concentration) had no effect on the rate of
recovery from acidification (Control: - 3.44E-041.34E-04; MDA: - 2.26E-040.69E-04). In MCF-7 cells, perfusing with Tyrode
containing 1 pM E2 significantly increased the rate of recovery from acidification (Control: - 1.61E-040.22E-04; MCF-7: - 3.66E-
040.18E-04; p<0.05). Inclusion of DMSO alone in the medium perfusing MCF-7 cells significantly increased the basal ratio compared
with cells in Tyrode alone (Control: 1.240.18; DMSO: 1.710.09; p<0.05), and also increased the rate of recovery (Control: - 1.61E-
040.22E-04; DMSO: - 3.97E-040.52E-04; p<0.02). Inclusion of RSV (1 mM) in the perfusate with MCF-7 cells also caused an
increase in the rate of recovery relative to the DMSO-only control (DMSO: - 3.97E-040.52E-04; RSV: - 6.84E-040.56E-04;
p<0.02). Gen (1 mM) had no significant effects in MCF-7 cells, though all three recovery rates were numerically higher than the DMSO
In summary, we have shown that E2 and RSV at physiological levels can increase the rate of recovery from acidification in a breast
cancer cell line expressing oestrogen receptors a and b. Given the potential importance of cytosolic and extracellular pH regulation in
both the ability of cytotoxic drugs to cross the membrane or the cell to proliferate, the physiological effects of foodborne phytohormones
on pathways regulating cell pH clearly requires further investigation.
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3. Hwang CS, Kwak HS, Lim HJ, Lee SH, Kang YS, Choe TB, Hur HG & Han KO (2006) J Steroid Biochem Mol Biol 101: 246-253.||