Mitogenic stimulation of human breast cancer cells in a growth factor‐defined medium: Synergistic action of insulin and estrogen

Abstract

The cooperative action of 17 beta-estradiol (E2) and polypeptide growth factors in stimulating proliferation of human breast cancer cells in vitro was investigated. To prevent background estrogenic stimulation, only phenol red-free media were used. When cultured in media supplemented with steroid-stripped serum in which all polypeptide growth factor activity had been chemically inactivated, MCF7 cells were unable to proliferate and became virtually quiescent. In the additional presence of insulin, epidermal growth factor (EGF), and E2, however, cells proliferated as rapidly as did cells cultured in media supplemented with fetal calf serum. Analysis by DNA flow cytometry showed that in the absence of external growth factors, MCF7 cells became arrested predominantly in the G1/G0 phase of the cell cycle. Upon addition of insulin in combination with EGF and E2, however, cells reentered the cell cycle with a high degree of synchrony. When added alone, E2 induced only slight mitogenic effects under these growth factor-defined conditions. In contrast, this steroid induced optimal proliferation in conventional steroid-stripped serum, which in itself contained considerable mitogenic activity. Insulin (at 10 micrograms/ml) was the most potent stimulator of MCF7 cell proliferation under growth factor-defined conditions, resulting in a more than sixfold increase in cell number after 96 hours. Other growth factors such as platelet-derived growth factor (PDGF), transforming growth factor beta (TGF beta), and EGF had little effect by themselves and only slightly influenced insulin-induced proliferation. At suboptimal concentrations of insulin (10-100 ng/ml), however, strong synergism was observed between E2 and insulin in inducing MCF7 proliferation. Using the CG5 cell line, a highly E2-sensitive MCF7 variant, synergism with E2 was already observed at 1 ng/ml insulin. It is concluded that MCF7 cells require insulin (or insulin-like growth factors) for proliferation. At suboptimal insulin concentrations, E2 acts synergistically with insulin, possibly by inducing autocrine production of polypeptide growth factors by these cells.

Insulin/Insulin-like Growth Factor-I and Estrogen Cooperate to Stimulate Cyclin E-Cdk2 Activation and Cell Cycle Progression in MCF-7 Breast Cancer Cells through Differential Regulation of Cyclin E and p21WAF1/Cip1

“…1A, stimulation of arrested cells with insulin (10 g/ml) or IGF-I (1 nM) alone resulted in a minor increase in S-phase cells to ϳ10 -12% at 26 h. Insulin dose-response experiments in serum-free medium without ICI 182780 indicated a maximal response between 100 ng/ml and 10 g/ml insulin (data not shown), and a similar maximal response was achieved with 1 nM IGF-I. A supraphysiological concentration (10 g/ml) of insulin, known to activate both insulin and IGF-I receptor signaling in breast epithelial cells (44,45), was used in all subsequent experiments, in agreement with the design of earlier studies demonstrating synergistic effects on breast epithelial cell proliferation (15,16,46). In marked contrast, estradiol (100 nM) alone stimulated a significantly greater S-phase entry of 43%.…”

“…In breast epithelial cells, insulin (13) and IGF-I (14) are potent mitogens and synergize with estrogen to stimulate cell proliferation (15,16). Several potential mechanisms have been proposed to account for these effects.…”

Glucose-lowering agents and cancer mortality rates in type 2 diabetes: assessing effects of time-varying exposure

“…The role of insulin in the relationship between type 2 diabetes and cancer mortality rates is supported by a biologically plausible mechanism. Insulin is a growth hormone and is known to have arthrogenic and mitogenic properties [34][35][36]. Specifically, it is suggested that hyperinsulinaemia and elevated levels of IGF-1 promote tumour cell growth in patients with type 2 diabetes [7,8].…”

Anthropometric measures, plasma adiponectin, and breast cancer risk

“…Insulin has been shown to stimulate the proliferation of breast cancer cells by binding and signaling through the insulin and insulinlike growth factor I (IGF-I) receptors (Milazzo et al 1992, Lai et al 2001. In addition, insulin may synergize with the mitogenic effects of estrogen (van der Burg et al 1988) and may also upregulate the expression of vascular endothelial growth factor (VEGF), a potent angiogenic agent that is secreted by breast cancer cells (Bachelder et al 2002). Moreover, adiponectin has been inversely associated with estrogen levels (Gavrila et al 2003).…”