Tissue compartment-specific estrogen receptor-alpha participation in the mouse uterine epithelial secretory response

17 beta-Estradiol (E-2) acts through the estrogen receptor (ER) to regulate uterine epithelial cell growth, proliferation, differentiation, and secret ory protein production. We have previously shown that E-2-induced uterine e pithelial proliferation is mediated indirectly by ER alpha-positive stroma; epithelial ER alpha is neither necessary nor sufficient for E-2-induced ut erine epithelial mitogenesis. in the present study, we addressed the questi on of whether production of uterine epithelial secretory proteins and their messenger RNAs (mRNAs) requires ER alpha in stroma, epithelium, or both by analyzing tissue recombinations composed of uterine tissue from adult ER a lpha knockout (ko) and neonatal BALB/c (wt) mice. Stroma (S) and epithelium (E) were separated by trypsinization, and four types of uterine tissue rec ombinants were prepared: wt-S + wt-E, wt-S + ko-E, ko-S + wt-E, and ko-S ko-E. These tissue recombinants were grown as subrenal capsule grafts in in tact female nude mice for 4 weeks, at which time the hosts were ovariectomi zed. To assess the production of secretory proteins and their mRNAs, 1 week after ovariectomy the hosts were given three daily injections of oil or E- 2 (100 ng), and then 24 h later the grafts were recovered and used for eith er ER or lactoferrin (LF) immunohistochemistry. To assess steady state mRNA levels by Northern blotting, hosts received one injection of oil or E-2 24 h before harvest. ER immunohistochemistry was used to monitor the complete ness of tissue separation. In wt-S + wt-E tissue recombinants from E-2-trea ted hosts, the epithelium stained intensely for LF (an abundant E-2-depende nt uterine secretory protein), whereas similar tissue recombinants from oil -treated hosts showed minimal immunostaining. Conversely, LF immunostaining was minimal in wt-S + ko-E grafts from both oil- and E-2-treated hosts. LF staining was also minimal in ko-S + ko-E and ko-S + wt-E tissue recombinan ts regardless of hormone treatment. For Northern analyses, the epithelial c ontent of the tissue recombinants was monitored using the reference epithel ial transcript, E-cadherin. While all tissue recombinant groups expressed E -cadherin mRNA, wt-S + wt-E tissue recombinants from E-2-treated hosts prod uced a strong, single 2.6-kb band of LF mRNA. LF transcripts were minimal o r absent in all other tissue recombinant types. Northern blotting results i dentical to those seen for LF were also observed for the uterine secretory protein complement component C3. Our data demonstrate that both stromal and epithelial ER alpha are required for the production of LF protein and of L F or C3 mRNAs in response to E-2. Thus, in contrast to E-2-induced epitheli al mitogenesis, which requires only stromal ER alpha, both epithelial and s tromal ER alpha are necessary for the production of E-2-dependent epithelia l secretory proteins.