Ovarian status modulates endocrine and neuroinflammatory responses to a murine mammary tumor

Abstract

Patients with breast cancer have increased circulating inflammatory markers and mammary tumors increase neuroinflammation in rodent models. Menopausal status is not only important in the context of breast cancer as circulating estrogen influences tumor progression, but also because estrogen is anti-inflammatory and an essential modulator of endocrine function in the brain and body. Here, we manipulated "menopause" status (ovary-intact and ovariectomized) in an estrogen receptor (ER)+ mouse mammary tumor model to determine the extent to which ovarian status modulates: 1) tumor effects on estrogen concentrations and signaling in the brain, 2) tumor effects on estrogen-associated neurobiology and inflammation, and 3) the ability for tumor resection to resolve the effects of a tumor. We hypothesized that reduced circulating estradiol (E₂) after an ovariectomy exacerbates tumor-induced peripheral and central inflammation. Notably, we observed ovarian-dependent modulation on tumor-induced peripheral outcomes, including E₂-dependent processes and, to a lesser degree, circulating inflammatory markers. In the brain, ovariectomy exacerbated neuroinflammatory markers in select brain regions and modulated E₂-related neurobiology due to a tumor and/or resection. Overall, our data suggest that ovarian status has moderate implications for tumor-induced alterations in neuroendocrinology and neuroinflammation and mild effects on peripheral inflammatory outcomes in this murine mammary tumor model.

Keywords: brain; breast cancer; cytokines; estrogen; tumor resection.

Figure 1.

Overall project timeline for tumor-free (Control), tumor-bearing (Tumor), and tumor-resected (Resect) mice. One-half of each experimental cohort was ovariectomized (OVX) before arrival and the other half remained ovary-intact (Intact). This study was completed in four treatment-balanced experiments (n = 30–39/group for entire project).

Figure 2.

Tumors interact with ovarian status to affect circulating estradiol (E₂) and estrogen-dependent reproductive processes. Effects of tumor and tumor resection in Intact and ovariectomized (OVX) mice on circulating E₂ concentrations (n = 19–27/group; A), estrous stages based on cytology from vaginal lavages (Intact only; 7–9 consecutive days; n = 10–22/group; means + SE; B), and uterus mass (n = 15–20/group; C). Data (A and C) are presented as means ± SE. *P ≤ 0.05 relative to ovary-matched Controls; #P ≤ 0.05 relative to Intact counterparts of the same tumor treatment group; &P ≤ 0.05 relative to ovary-matched Tumors. Control, sham control mice; Intact, ovary-intact mice; Resect, tumor-resected mice; Tumor, tumor-bearing mice.

Figure 3.

Tumors or tumor resection modulate ovariectomy-induced alterations in extra-ovarian peripheral estradiol (E₂) concentrations. E₂ concentrations in adrenal gland (n = 10–13/group; A), mammary white adipose (n = 9–12/group; B), and tumor (n = 11–12/group; C) tissues. Data are presented as means ± SE. *P ≤ 0.05 relative to ovary-matched Controls; #P ≤ 0.05 relative to Intact counterparts of the same tumor treatment group; &P ≤ 0.05 relative to ovary-matched Tumors. Control, sham control mice; Intact, ovary-intact mice; Resect, tumor-resected mice; Tumor, tumor-bearing mice.

Figure 4.

Tumors modulate hypothalamic ER and HPO axis-related gene expression in an ovarian status-dependent manner. Hypothalamic gene expression of estrogen receptor 1 (Esr1; A), estrogen receptor 2 (Esr2; B), G protein-coupled estrogen receptor 1 (Gper1; C), gonadotropin-releasing hormone (Gnrh; D), kisspeptin-1 (Kiss1; E), and kisspeptin-1 receptor (Kiss1r; F). n = 7–12/group. Data are presented as means ± SE. *P ≤ 0.05 relative to ovary-matched Controls; #P ≤ 0.05 relative to Intact counterparts of the same tumor treatment group. Control, sham control mice; ER, estrogen receptor; HPO, hypothalamic-pituitary-ovarian; Intact, ovary-intact mice; Resect, tumor-resected mice; Tumor, tumor-bearing mice.

Figure 5.

Ovarian status modestly modulates brain E₂ production and ER expression in a region-specific manner. Aromatase gene (Cyp19a1) expression, E₂ concentrations, and Esr1 and Esr2 gene expression in the hippocampus (A), frontal cortex (B), and hypothalamus (C). n = 7–12/group. Data are presented as means ± SE. *P ≤ 0.05 relative to ovary-matched Controls; #P ≤ 0.05 relative to Intact counterparts of the same tumor treatment group; &P ≤ 0.05 relative to ovary-matched Tumors. Control, sham control mice; ER, estrogen receptor; Esr1, estrogen receptor 1; Esr2, estrogen receptor 2; E₂, estradiol; Intact, ovary-intact mice; Resect, tumor-resected mice; Tumor, tumor-bearing mice.

Figure 6.

Ovarian status modulates select tumor-induced inflammatory markers in circulation. Proinflammatory cytokine interleukin 6 (IL-6; A), chemokines C-X-C motif chemokine ligand 1 (CXCL1; B) and (CXCL10; C), and anti-inflammatory cytokine IL-10 plasma concentrations (D). n = 6–19/group. Data are presented as means ± SE. *P ≤ 0.05 relative to ovary-matched Controls; #P ≤ 0.05 relative to Intact counterparts of the same tumor treatment group; &P ≤ 0.05 relative to ovary-matched Tumors. Control, sham control mice; Intact, ovary-intact mice; Resect, tumor-resected mice; Tumor, tumor-bearing mice.

Figure 7.

Ovarian status modulates the neuroinflammatory response to a tumor. Cytokines Il1b and Il6 gene expression in the hypothalamus (A), hippocampus (B), and frontal cortex (C). n = 7–12/group. Data are presented as means ± SE. *P ≤ 0.05 relative to ovary-matched Controls; #P ≤ 0.05 relative to Intact counterparts of the same tumor treatment group; &P ≤ 0.05 relative to ovary-matched Tumors. Control, sham control mice; Intact, ovary-intact mice; Resect, tumor-resected mice; Tumor, tumor-bearing mice.