Effect of Prenatal and Neonatal Anti‐Androgen Flutamide Treatment on Aquaporin 5 Expression in the Adult Porcine Ovary

Abstract

The growth of ovarian follicles is accompanied by fluid-filled antrum formation. Water movement within the follicular wall is predominantly transcellular via membranous water channels named aquaporins (AQPs). Androgens are important regulators of mammalian folliculogenesis, and their prenatal and/or neonatal deficiency affects female fertility in adulthood. Therefore, this study was performed to determine whether gestational or neonatal exposure to the anti-androgen flutamide influences androgen-dependent AQP5 expression in pre-antral and large antral follicles of adult pigs. Flutamide was injected into pregnant gilts between days 80 and 88 of gestation and into female piglets between days 2 and 10 post-natally. The ovaries were collected from flutamide-treated and non-treated (control) sexually mature pigs. In pre-antral follicles, AQP5 mRNA and protein levels were both downregulated following maternal (p < 0.01 and p < 0.01, respectively) and neonatal (p < 0.01 and p < 0.01, respectively) flutamide exposure. Likewise, the expression of mRNA (p < 0.01 and p < 0.001, respectively) and protein (p < 0.05 and p < 0.01, respectively) for AQP5 were diminished in large antral follicles in both groups. Immunohistochemistry showed decreased intensity of AQP5 immunoreaction in pre-antral (p < 0.01) and large antral (p < 0.001) follicles following flutamide treatment. Moreover, radioimmunological analysis revealed that changes observed in AQP5 expression corresponded with diminished follicular androgens production after both maternal (p < 0.05 and p < 0.05, respectively) and neonatal (p < 0.05 and p < 0.01, respectively) flutamide administration. Therefore, AQP5 appears to be a potential regulator of follicular fluid accumulation, under androgen control, and may be a key factor in antral follicle growth.

Androgen Stimulates Growth of Mouse Preantral Follicles In Vitro: Interaction With Follicle-Stimulating Hormone and With Growth Factors of the TGFβ Superfamily

“…Aquaporins facilitate water transport and are expressed in GCs (40), and testosterone stimulated the swelling of porcine GCs in hypotonic culture medium, an effect that was reversed by flutamide, suggesting that aquaporins are sensitive to androgen (41). A recent study by the same group confirmed that follicles from pigs that were exposed to flutamide in utero expressed less Aqp5 messenger RNA and protein, demonstrating that androgen could potentially stimulate aquaporin expression leading to antrum formation (42). …”

Expression of aquaporin 4 in the chicken ovary in relation to follicle development

“…Recently, Grzesiak et al. () revealed that maternal and neonatal exposure to anti‐androgen flutamide leads to decreased AQP5 expression in ovarian follicles of adult pigs. Moreover, presence of the oestrogen‐response element was confirmed in the promoter region of the AQP2 (Zou et al., ) and AQP5 (Kobayashi, Takahashi, Miyagawa, Watanabe, & Iguchi, ) genes, and androgen‐response element in the promoter of AQP5 gene (Moehren, Denayer, Podvinec, Verrijdt, & Claessens, ).…”

“…According to Grzesiak et al. (), supernatants containing 20 μg of total protein were mixed with loading buffer (62.5 m m Tris–HCl pH 6.8, 2% SDS, 25% glycerol, 0.01% bromophenol blue, 5% β‐mercaptoethanol) and warmed at 99.9°C for 3 min. After denaturation, samples were loaded into 12% SDS‐polyacrylamide gel, and proteins were separated by electrophoresis under reducing conditions.…”

The expression profile of <i>AQP1</i>, <i>AQP5</i> and <i>AQP9</i> in granulosa and theca cells of porcine ovarian follicles during oestrous cycle and early pregnancy

“…Whereas, the presence of AQP5 mRNA in porcine ovarian follicles was reported recently by Grzesiak et al (2016). In that study diminished AQP5 mRNA and protein expressions within pre-antral and large antral follicles of adult pigs were presented.…”