Estrous cycle modulates ovarian carcinoma growth

Abstract

Purpose: The effects of reproductive hormones on ovarian cancer growth are not well understood. Here, we examined the effects of estrous cycle variation and specific reproductive hormones on ovarian cancer growth.

Experimental design: We investigated the role of reproductive hormones in ovarian cancer growth using both in vivo and in vitro models of tumor growth.

Results: In vivo experiments using the HeyA8 and SKOV3ip1 ovarian cancer models showed that tumor cell inoculation during proestrus significantly increased tumor burden (251-273%) compared with injection during the estrus phase. Treatment of ovariectomized mice with 17beta-estradiol resulted in a 404% to 483% increase in tumor growth compared with controls. Progestins had no significant effect, but did block estrogen-stimulated tumor growth. Tumors collected from mice sacrificed during proestrus showed increased levels of vascular endothelial growth factor (VEGF) and microvessel density compared with mice injected during estrus. HeyA8, SKOV3ip1, and mouse endothelial (MOEC) cells expressed estrogen receptor alpha and beta and progesterone receptor at the protein and mRNA levels, whereas 2774 ovarian cancer cells were estrogen receptor-negative. In vitro assays showed that 17beta-estradiol significantly increased ovarian cancer cell adhesion to collagen in estrogen receptor-positive, but not in estrogen receptor-negative cells. Additionally, 17beta-estradiol increased the migratory potential of MOEC cells, which was abrogated by the mitogen-activated protein kinase (MAPK) inhibitor, PD 09859. Treatment with 17beta-estradiol activated MAPK in MOEC cells, but not in HeyA8 or SKOV3ip1 cells.

Conclusion: Our data suggest that estrogen may promote in vivo ovarian cancer growth, both directly and indirectly, by making the tumor microenvironment more conducive for cancer growth.

Figure 1

Inoculation of tumor cells into the peritoneal cavity of mice during proestrus results in higher tumor weights and nodule formation. A) Characterization of the estrous cycle in nude mice by vaginal cytology. In vivo effects of the estrous cycle on tumor growth was assessed on mice injected i.p. with either HeyA8 (B) or SKOV3ip1 (C) cells during proestrus or estrus. Three to four weeks later, animals were sacrificed and a necropsy was performed. Mice injected during proestrus showed increased tumor burden (251-273%; p < 0.05) and increased tumor nodule formation (178%-196%; p < 0.05).

Figure 2

Effects of reproductive hormones on ovarian cancer growth and angiogenesis. Following recovery from ovariectomy, mice received either 17β-estradiol, progestin, or combination treatment. After three days of treatment, mice were injected with either A) SKOV3ip1 or B) HeyA8 ovarian cancer cells. Four weeks later, mice were sacrificed and tumor burden was assessed. Effect of estrous cycle variation on vessel density (C) and VEGF (D) was assessed in tumors from mice inoculated during proestrus or estrus. All pictures were taken at original magnification, X200.

Figure 3

Hormone receptor expression in tumor and endothelial cell. A) mRNA expression of ER α, ER β and PR in HeyA8, SKOV3ip1 and MOEC cells. B) Western blot analyses show that all three cell lines expressed all three receptors.

Figure 4

Estrogen increases ovarian cancer cell adhesion, endothelial cell migration and activates MAPK in endothelial cells. A) Effect of 17β-estradiol on the adhesive potential of HeyA8, SKOV3ip1 and 2774 cells to collagen I was assessed. B) Effect of 17β-estradiol on MAPK activation in HeyA8, SKOV3ip1 and MOEC cells. C) Effect of 17β-estradiol, PD 09859 or in combination, on migration of MOEC cells was examined with a gelatin-coated membrane.