Effects of isoliquiritigenin on ovarian antral follicle growth and steroidogenesis

Abstract

Isoliquiritigenin is a botanical estrogen used as a dietary supplement. Previous studies show that other botanical estrogens affect ovarian estradiol synthesis, but isoliquiritigenin's effects on the ovary are unknown. Thus, this study tested the hypothesis that isoliquiritigenin inhibits ovarian antral follicle growth and steroidogenesis. Antral follicles from CD-1 mice were cultured with vehicle control (dimethyl sulfoxide; DMSO) or isoliquiritigenin (0.6μM, 6 μM, 36 μM, and 100 μM) for 48-96h. During culture, follicle diameters were measured daily to assess follicle growth. After culture, media were collected for hormone assays and follicles were collected for gene expression analysis of steroidogenic enzymes. Isoliquiritigenin inhibited antral follicle growth and altered estradiol, testosterone, and progesterone levels. Additionally, isoliquiritigenin altered the mRNA levels of cytochrome P450 steroid 17-α-hydroxylase 1, aromatase, 17β-hydroxysteroid dehydrogenase 1, and steroidogenic acute regulatory protein. These data indicate that exposure to isoliquiritigenin inhibits growth and disrupts steroid production in antral follicles.

Keywords: Antral follicle; Isoliquiritigenin; Licorice root; Ovary; Steroidogenesis.

Fig. 1

Effect of isoliquiritigenin on follicle growth overtime. Antral follicles were isolated from adult CD-1 mice and cultured with vehicle (DMSO) or isoliquiritigenin (ISL) for total culture periods of 48, 72, and 96 h. Follicle diameters were measured every 24 h along perpendicular axes and percent change in growth was determined over each total culture period. The graph represents the means ± SEM of percent change in follicle growth from 4 to 11 separate experiments. Asterisks (*) represent a significant difference between vehicle control and isoliquiritigenin (p ≤ 0.05).

Fig. 2

Effect of isoliquiritigenin on follicle estradiol production. Antral follicles were isolated from adult CD-1 mice and cultured with vehicle (DMSO) or isoliquiritigenin (ISL) for 48, 72, and 96 h. Subsequently, media were pooled within experiment by treatment group and subjected to enzyme-linked immunosorbent assays for estradiol. The graph represents the means ± SEM from 3 to 4 separate experiments. Asterisks (*) represent a significant difference between vehicle control and isoliquiritigenin (p ≤ 0.05).

Fig. 3

Effect of isoliquiritigenin on follicle testosterone production. Antral follicles were isolated from adult CD-1 mice and cultured with vehicle (DMSO) or isoliquiritigenin (ISL) for 48, 72, and 96 h. Subsequently, media were pooled within experiment by treatment group and subjected to enzyme-linked immunosorbent assays for testosterone. The graph represents the means ± SEM from 3 to 4 separate experiments. Asterisks (*) represent a significant difference between vehicle control and isoliquiritigenin (p ≤ 0.05).

Fig. 4

Effect of isoliquiritigenin on follicle progesterone production. Antral follicles were isolated from adult CD-1 mice and cultured with vehicle (DMSO) or isoliquiritigenin (ISL) for 48, 72, and 96 h. Subsequently, media were pooled within experiment by treatment group and subjected to enzyme-linked immunosorbent assays for progesterone. The graph represents the means ± SEM from 3 to 4 separate experiments. Asterisks (*) represent a significant difference between vehicle control and isoliquiritigenin (p ≤ 0.05).

Fig. 5

Effect of isoliquiritigenin on follicle mRNA level of regulators of steroidogenesis. Antral follicles were isolated from adult CD-1 mice and cultured with vehicle (DMSO) orisoliquiritigenin (ISL) for 48 and 96 h. Subsequently, follicles were pooled within experiment by treatment group and subjected to quantitative polymerase chain reaction for Cyp17a1 (panel A), Cyp19a1 (panel B), Hsd17b1 (panel C), and Star (panel D). Values were normalized to Actb. The graph represents the means ± SEM from 3 to 4 separate experiments. Asterisk (*) represents a significant difference between vehicle control and isoliquiritigenin (p ≤ 0.05).

Fig. 6

Effect of isoliquiritigenin on follicle steroidogenesis. This schematic of the estradiol biosynthesis pathway shows how estradiol is produced from cholesterol through the action of steroidogenic enzymes in the ovarian antral follicle. The effects of different concentrations of isoliquiritigenin (ISL) on the levels of progesterone, testosterone, and estradiol, as well as the transport protein STAR and the enzymes CYP17A1, CYP19A1, and HSD17B1 are summarized.