Figure 3.
Microarray analyses of DHT-treated ovaries identified specific changes in gene expression in granulosa and stromal cells, with the notable induction of Vcam1 in the stroma. (A) Microarray analyses were done on total RNA derived from mechanically separated stromal and granulosa cells from ovaries of control mice and mice treated with DHT for 1 wk. A curated list, presented as a heat map, identified ovarian genes regulated selectively by DHT in the ovarian stroma and granulosa fractions. (B) A comparison of the expression levels of ovarian marker genes was carried out on isolated stromal and granulosa compartments to validate the purity (∼90%) of mechanically separated stromal and granulosa compartments. (C) Fshr, a granulosa marker and androgen target, was induced by androgens but only in the granulosa compartment, whereas Vcam1 was potently induced specifically in the stromal compartment. (D) Theca-specific AR deletion (in CYP17A1-iCRE, ARf/f mice) did not prevent the induction of Fshr expression in response to DHT (a granulosa-specific response), but it did inhibit Vcam1 induction in the theca, demonstrating that the signal to upregulate stromal Vcam1 mRNA levels is transmitted through theca-specific AR. (A) Microarrays were carried out on two biological replicates. (B) Microarray validation and (C, D) other results were carried out on N > 3 for all conditions. *P < 0.05 by Student t test.