The Role of Estrogens in Insulin Secretion. Implications for Aromatase Inhibitor Treatment

Introduction The main hormonal treatment of estrogen sensitive breast cancer includes the use of selective estrogen receptor modulators, aromatase inhibitors and GnRH-Analogs. It has been observed that administering aromatase inhibitor to breast cancer patients not only impairs their glucose metabolism but it can even cause frank diabetes mellitus. Moreover, it has been hypothesized that aromatase inhibitors may have an impact on glucose metabolism by their effect on estrogen levels. Therefore, we designed an experiment in order to assess the effect of estrogens on insulin secretion from rat beta pancreatic insulinoma INS-1 cells. Methods Experiments were conducted on an INS-1 cell line, a rodent beta cell line derived from a rat insulinoma induced by X-ray irradiation, which displays high insulin content, production of both proinsulin I and II and responsiveness to glucose and hormones. INS-1 cells were routinely cultured in 75cm2 flasks (T75) containing 10ml of appropriate culture media and then were transferred into 6-well plates and treated with 17β-E2. Proliferation, as well as insulin expression in the 17β-E2 treated cells in mRNA and protein level was then investigated. Cell cultures were treated with 12.5mM, 25mM, 50mM, 100mM and 200mM estradiol. After 24 hours, RNA as well as protein extraction was carried out. Gene expression was examined in mRNA and protein level, by real time quantitative reverse transcriptase PCR and by western blotting, respectively. Extraction of total RNA was achieved with the use of NucleoZOL (Macherey-Nagel, Duren, Germany). Protein concentration of samples was measured by using the bicinchoninic acid (BCA) assay and bovine serum albumin (BSA) as the standard (Thermo Scientific TM Pierce TM BCA Protein Assay Kit, USA). Results Real time PCR showed a dose-dependent up regulation of insulin gene expression by estradiol. The findings were consistent with the results from western blot, although the estradiol dose-dependent increase in gene expression was not so clear. Finally, both protein and mRNA expression of insulin increased in a dose dependent manner, with mRNA reaching a plateau at 100nM estradiol treatment. Conclusion The experimental data suggest that there might be a direct effect of estrogen on beta cells and insulin secretion.