ESPE Abstracts (2018) 89 FC8.6

The Roles of Steroids in Gonadal Development and Maintenance - Insights from a Zebrafish Model of Androgen and Cortisol Deficiency

James A. Oakesa, Nan Lia, Belinda Wistowa, Karl-Heinz Storbeckb, Vincent T. Cunliffea & Nils Kronea

aUniversity of Sheffield, Sheffield, UK; bStellenbosch University, Stellenbosch, South Africa.

Increasing evidence suggests that the aetiology of disorders of sex development cannot be solely explained by genetic alterations. It is highly likely that environmental factors hitting susceptible genetic backgrounds are partly causative. Zebrafish sex is highly plastic during development, making this species an ideal model for investigation of endocrine disruption and gonadal development and function. However, the hormonal regulation of these processes in zebrafish is poorly understood. Here, we use a model of glucocorticoid and androgen deficiency to explore such processes. In humans, ferredoxin (FDX1) is an electron-providing cofactor required for steroid biosynthesis. The zebrafish homologue of FDX1, fdx1b, plays a crucial additional role in androgen biosynthesis. To investigate the role of steroids in sex development and gonadal differentiation we analysed the phenotype of adult fdx1b mutant zebrafish, which were found to be profoundly cortisol- and androgen-deficient by LC-MS/MS. Downregulation of cortisol responsive genes fkbp5 and pck1 and androgen responsive gene cyp2k22 confirmed systemic steroid deficiencies. Fdx1b mutants exhibit feminised secondary sex characteristics, but may possess either testes or ovaries, and both sexes are sterile. Histological investigation showed abnormal seminiferous tubule structure and disorganisation of fdx1b mutant testes, compared to those of wild-type siblings. To investigate mechanisms behind testicular disruption and sterility we measured expression of genes regulating testicular development or spermatogenesis. We observed downregulation of pro-testis gene sox9a, and igf3, a key factor for spermatogonial proliferation and differentiation, in fdx1b mutant testes. Androgen receptor (ar) expression was upregulated in mutant males, whereas putative AR target gene dmrt1 was expressed at similar levels in mutants and wild-type siblings. These observations suggest that increased ar expression may compensate for the observed androgen deficiency, rescuing dmrt1 expression, or alternatively that in zebrafish, dmrt1 expression is insensitive to reduced androgen levels. Whilst androgens regulate some secondary sex characteristics, they do not promote testis differentiation, as mutants developed distinct ovaries or testes. However, it is clear that androgens have an important role in development, maturation, organisation and function of both male and female gonads, since adult males and females were sterile. Taken together, our observations provide novel insights into the roles of androgens in these processes and for the first time demonstrate a dynamic response to androgen deficiency involving compensatory upregulation of the ar and rescued dmrt1 expression. We anticipate that these insights will support development of model organisms to study the interplay of genetic factors and environment in disorders of sex development.

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