Journal of Steroid Biochemistry & Molecular Biology 79 (2001) 181–185 The ovarian phenotype of the aromatase knockout (ArKO) mouse  Kara L. Britt a , Ann E. Drummond a,∗ , Mitzilee Dyson a , Nigel G. Wreford b , Margaret E.E. Jones a , Evan R. Simpson a , Jock K. Findlay a a Prince Henry’s Institute of Medical Research, P.O. Box 5152, Clayton, Vic. 3168, Australia b Department of Anatomy, Monash University, Clayton, Vic. 3168, Australia Abstract Targeted disruption of exon 9 of the cyp19 gene gives rise to a non-functional aromatase enzyme incapable of converting androgens to oestrogens. The aromatase knockout (ArKO) mouse is, thus, characterised by a dysfunctional pituitary-gonadal axis, which manifests in non-detectable levels of oestrogen in serum. These mice also exhibit elevated levels of circulating gonadotrophins (luteinising hormone (LH) and follicle stimulating hormone (FSH)) and testosterone. The ArKO mouse is infertile due to folliculogenic disruption and a failure to ovulate. The age-dependent ovarian phenotype revealed a block in follicular development at the antral stage and a complete absence of corpora lutea. By 21–23 weeks of age haemorrhagic cystic follicles were present and by 1 year there were abnormal follicles, an absence of secondary and antral follicles and atretic primary follicles. Interstitial tissue remodelling was extensive and exemplified by an increase in collagen deposition and an influx of macrophages, coincident with the loss of follicles. In mice, maintained on a soy-free and, thus, phytoestrogen-free diet, the ovarian phenotype was accelerated and exacerbated. In conclusion, the ovarian phenotype of the ArKO mouse can be attributed to the altered hormonal environment brought about by the absence of aromatase and the failure of androgens to be converted to oestrogens in the presence of elevated gonadotropins. © 2002 Elsevier Science Ltd. All rights reserved. Keywords: Aromatase; Knockout mouse; Ovary; Oestrogen; Gonadotrophin 1. Introduction Aromatase, a member of the cytochrome P450 superfam- ily, is the enzyme responsible for converting androgens (C19 steroids) to oestrogens (C18 steroids). In the ovary, aro- matase is localised to the granulosa cell population where, under the control of follicle stimulating hormone (FSH) [1], it plays a key role in the two cell, two gonadotrophin hy- pothesis; testosterone or androstenedione produced by theca cells in response to luteinising hormone (LH) is transported to the granulosa cells where aromatisation to oestrogen occurs. Oestrogens stimulate the proliferation of granulosa cells and facilitate the LH- and FSH-induced differentiation of these cells [2,3]. The autocrine actions of oestradiol-17 (E 2 ), the principal bioactive oestrogen, are mediated via two nuclear transcription factors, oestrogen receptor (ER)- and -, both of which are present in granulosa cells of mice, rats, primates and humans [4–6]. Despite evidence to support a local role for oestrogen in the development of follicles in the mammalian ovary  Proceedings of the Symposium: ‘Aromatase 2000 and the Third Generation’ (Port Douglas, Australia, 3–7 November 2000). ∗ Corresponding author. Tel.: +61-3-9594-4398; fax: +61-3-9594-6125. E-mail address: ann.drummond@med.monash.edu.au (A.E. Drummond). [7], a specific point of action and an absolute requirement for oestrogen has yet to be demonstrated. Initially, murine models with targeted disruption of the genes for either ER- (ERKO) or ER- (BERKO) [8–10] were generated to address these issues. However, neither of these models is completely free of the capacity to transduce the E 2 signal, given that the alternative estrogen receptor subtype remains [4,11]. Two laboratories [10,12] have now generated double ER knockout (ERKO) mice. The ovarian phenotype of these mice is distinct from the individual ER knockouts, suggesting that both ERs are required for normal ovarian function. An alternative model to ER depletion is to remove oestrogen itself from the ovary. Studies of aromatase de- ficient and thus oestrogen deficient human females report with a non-adrenal form of female pseudohemaphrodism, sexual infantilism, primary amenorrhea, ambiguous external genitalia at birth and polycystic ovaries [13–15]. At puberty these females display no breast development, clitorimegaly, multiple bilateral ovarian cysts and primary amenorrhea, in the face of increased basal levels of gonadotrophins, testos- terone and androstenedione [13–16]. A mouse model lack- ing functional aromatase activity (ArKO), due to targeted disruption of exon 9, a highly conserved and structurally important exon of the cyp19 (aromatase) gene [17] has been developed. The aromatase knockout (ArKO) mouse 0960-0760/02/$ – see front matter © 2002 Elsevier Science Ltd. All rights reserved. PII:S0960-0760(01)00158-3