Estradiol Has Differential Effects on Acute Colonic Inflammation in the Presence and Absence of Estrogen Receptor β Expression

Abstract

Background: Inflammatory bowel disease (IBD) increases the risk of developing colon cancer. This risk is higher in men compared to women, implicating a role for female hormones in the protection against this disease. Studies from our laboratory demonstrated that estradiol (E₂) protects against inflammation-associated colon tumor formation when administered following chemical carcinogen and induction of chronic colitis.

Aim: This study seeks to better understand the effect of E₂ on acute colitis in the presence and absence of estrogen receptor β (ERβ).

Methods: Inflammation was induced by 2,4,6-trinitrobenzenesulfonic acid in wild-type (WT) and ERβ knockout (ERβKO) mice implanted with a control or E₂-containing pellet and killed 5 days later. Inflammation and injury were scored by a pathologist. Apoptosis and proliferation were assessed by immunohistochemistry. Cytokines were measured by multiplex analysis.

Results: E₂ treatment reduced inflammation in the middle colon in WT mice and the distal colon in ERβKO mice compared to control mice. WT mice had reduced IL-6, IL-12, IL-17, GM-CSF, IFN-γ, MCP-1, MIP-1α, and TNF-α, and ERβKO had reduced IL-6 and IFN-γ expression in response to E₂. Injury scores were lower in E₂-treated ERβKO mice compared to control ERβKO mice. ERβKO mice had increased proliferation in the basal third of crypts in the distal colon and decreased apoptosis in the proximal colon.

Conclusions: These data suggest that E₂ has differential protective effects against acute colitis in the presence or absence of ERβ and provide insight into how E₂ may protect against IBD.

Keywords: Colon cancer; Crohn’s disease; Estradiol; Estrogen receptor beta; Inflammation.

Fig. 1

a Effect of E₂ on TNBS-induced weight loss. Values are mean weight loss per animal in grams ± SEM and b effect of E₂ on colon length. Values are mean colon length ± SEM. n = 12–16 mice per group. Bars without a common letter differ, P ≤ 0.05

Fig. 2

Inflammation was induced in the colon of ovariectomized mice with TNBS. Sectioned tissues were H&E stained and analyzed by a board-certified pathologist. a Inflammation scores and b injury scores. Values are mean score ± SEM. n = 12–16 mice per group. “*” denotes significance between indicated groups, P < 0.05

Fig. 3

Cytokine expression in colon tissue was assessed using a multiplex assay. Values are mean expression ± SEM. Bars without a common letter differ, P < 0.05

Fig. 4

Effect of E₂ on proliferation during acute inflammation. Two hours prior to kill, mice were injected with 5-bromo-2′-deoxyuridine (BrdU). Immunohistochemistry for BrdU was performed on sectioned tissues from the proximal and distal colon. Data are expressed as the percentage of proliferative cells compared to the total number of cells for either the full crypt column or the indicated region of the crypt. Data are representative of 20 well-oriented crypts per animal per proximal or distal colon with n = 12–16 mice per group. a Proliferation in the proximal colon and b proliferation in the distal colon. “*” denotes significance between indicated groups, P < 0.05

Fig. 5

A TUNEL assay was performed on sectioned tissues from the proximal and distal colon. a Apoptosis in the proximal colon and b apoptosis in the distal colon. Data are expressed as the mean percentage of apoptotic cells compared to the total number of cells for either the full crypt column or the indicated region of the crypt ± SEM. Data are representative of 20 well-oriented crypts per animal per proximal or distal colon with n = 12–16 mice per group. “*” denotes significance between indicated groups, P < 0.05