Stress hormone-mediated invasion of ovarian cancer cells

Abstract

Purpose: There is growing evidence that stress and other behavioral factors may affect cancer progression and patient survival. The underlying mechanisms for this association are poorly understood. The purpose of this study is to determine the effects of stress-associated hormones norepinephrine, epinephrine, and cortisol on the invasive potential of ovarian cancer cells.

Experimental design: The ovarian cancer cells EG, SKOV3, and 222 were exposed to increasing levels of either norepinephrine, epinephrine, or cortisol, and the in vitro invasive potential was determined using the membrane invasion culture system. Additionally, the effects of these stress hormones on matrix metalloproteinase-2 (MMP-2) and MMP-9 were determined by ELISA. The effects of the beta-adrenergic agonist isoproterenol on in vivo tumor growth were determined using nude mice.

Results: Stress levels of norepinephrine increased the in vitro invasiveness of ovarian cancer cells by 89% to 198%. Epinephrine also induced significant increases in invasion in all three cell lines ranging from 64% to 76%. Cortisol did not significantly affect invasiveness of the EG and 222 cell lines but increased invasion in the SKOV3 cell line (P = 0.01). We have previously shown that ovarian cancer cells express beta-adrenergic receptors. The beta-adrenergic antagonist propanolol (1 mumol/L) completely blocked the norepinephrine-induced increase in invasiveness. Norepinephrine also increased tumor cell expression of MMP-2 (P = 0.02 for both SKOV3 and EG cells) and MMP-9 (P = 0.01 and 0.04, respectively), and pharmacologic blockade of MMPs abrogated the effects of norepinephrine on tumor cell invasive potential. Isoproterenol treatment resulted in a significant increase in tumor volume and infiltration in the SKOV3ip1 in vivo model, which was blocked by propranolol.

Conclusions: These findings provide direct experimental evidence that stress hormones can enhance the invasive potential of ovarian cancer cells. These effects are most likely mediated by stimulation of MMPs.

Fig. 1

Invasion profile of ovarian cancer cell lines (222, EG, and SKOV3) in the presence or absence of norepinephrine (NE; A), epinephrine (Epi; B), or cortisol (Cort; C). Bars, SE.

Fig. 2

Effect of β-blocker (propranolol) on ovarian cancer invasion in the presence or absence of norepinephrine. Bars, SE.

Fig. 3

MMP-9 (A and B) and MMP-2 (C and D) concentrations in conditioned medium from SKOV3 (A and C) and EG(B and D) cell lines after treatment with norepinephrine. Bars, SE.

Fig. 4

MMP-9 (A and B) and MMP-2 (C and D) concentrations in conditioned medium from SKOV3 (A and C) and EG (B and D) cell lines after treatment with epinephrine. Bars, SE.

Fig. 5

The in vitro invasive potential of ovarian cancer cell lines (EG and SKOV3) through a defined basement membrane matrix over 24 hours was normalized to a value of 100%, and the invasive potential cells treated with 1 µmol/L norepinephrine and/or 5 µg/mL CMT-3 during the assay compared with the control value of 100%. Bars, SE.

Fig. 6

Effect of β-adrenergic agonist isoproterenol on in vivo growth of SKOV3ip1 cells injected s.c. in nude mice. H&E sections (original magnification, ×40) were obtained from (A) normal mouse skin without tumor and from tumor-bearing mice treated daily i.p. for 7 days with (B) PBS, (C) isoproterenol (10 mg/kg), or (D) isoproterenol (10 mg/kg) plus propranolol (2 mg/kg). The tumor (T) is seen infiltrating deeply through the muscle layer after daily isoproterenol treatment (C), but remained encapsulated in the other two groups.