Differential Expression of Estrogen Receptor-α and -β and Androgen Receptor in the Ovaries of Marmosets and Humans

Abstract

Estrogens and androgens are essential for the maturation of the ovarian follicle and normal fertility in the female. We have used antibodies specific for both forms of estrogen receptor (alpha [ERalpha] and beta [ERbeta]) and androgen receptor (AR) to investigate the pattern of receptor expression in ovaries obtained from women and from a New World primate, the Common marmoset (Callthrix jacchus). On Western blots, three antibodies directed against different peptides within human ERbeta all recognized recombinant human (h) ERbeta but did not bind to recombinant hERalpha. The ERbeta protein was extracted from human ovary and prostate and marmoset ovary. In marmoset and human ovaries, ERbeta protein was detected in the nuclei of granulosa cells in all sizes of follicle (both before and after formation of the antrum), and it was also detected in thecal cells, corpora lutea, surface epithelium, and stroma. In contrast, ERalpha protein was not detected in the nuclei of granulosa cells in preantral follicles, was low/absent from stromal and thecal cells, but was expressed in granulosa cells of antral follicles and in the surface epithelium. The pattern of expression of AR protein more closely resembled that of ERbeta than ERalpha. In conclusion, three independent antibodies have demonstrated convincingly that ERbeta is expressed in a wide range of cells in the primate ovary. Granulosa cells in preantral follicles could contain ERbeta:beta dimers. In antral follicles, however, ERalpha is also expressed, and the formation of homo- or heterodimers containing ERalpha may influence the pattern of gene activation within these cells.

Expression of oestrogen receptor beta (ERβ1) protein in human breast cancer biopsies

“…These results were consistent with previously published data (Saunders et al, 2000); no binding of the ERb specific monoclonal to ERa was observed (Figure 2, lower panel, lane a). The anti-hERb monoclonal that was directed against a peptide at the C-terminus of hERb bound to both short (Mosselman et al, 1996) and long (Ogawa et al, 1998a) forms of ERb.…”

“…It was notable that in six of the eight samples proteins migrating with apparent molecular sizes corresponding to both long (*59 Kd) and short (53 Kd) ERb were present. We have found that this antibody recognizes ERb protein extracted from human ovary, prostate (Saunders et al, 2000) endometrium and testis and human cell lines (MCF-7, Ishikawa, unpublished observations). In breast tumour samples that appeared to contain high levels of expression of full length ERb (numbers 1, 3, 4, 7, 8) several lower molecular weight protein species with apparent molecular weights from 32 to 45 Kd were detected.…”

“…A peptide located at the C-terminus of hERb (Mosselman et al, 1996) (CSPAEDSKS-KEGSQNPQSQ) was used to prepare a monoclonal antibody in mice according to standard methods and positive clones were identified by ELISA using recombinant human ERb (P2466, PanVera, Madison, WI, USA) (Saunders et al, 2000). This antibody has been used previously to demonstrate expression of ERb using human ovarian tissue sections (Saunders et al, 2000).…”

“…Recombinant hERa (*66 Kd) was also obtained from Pan Vera. Gel analysis and blotting were carried out as described previously (Saunders et al, 2000). Briefly, proteins were extracted from frozen biopsy specimens by rapid homogenization of tissue in denaturing/loading buffer (50 mM Tris-HCl pH 6.8, 100 mM DTT, 2% SDS, 0.1% bromophenol blue, 10% glycerol, all from Sigma).…”

“…Anti-ERb Immunolocalization was undertaken as described in detail in Saunders et al (2000), Sections were blocked for 30 min in normal rabbit serum (NRS, Diagnostics Scotland, Carluke) diluted 1 : 4 in TBS containing 5% BSA (NRS/TBS/BSA), rinsed briefly in TBS and an avidin biotin block performed using reagents from Vector (Peterborough, UK). Anti-ERb antibody was diluted 1 : 40 in NRS/TBS and incubated on sections overnight at 48C.…”