Tissue interactions and estrogenic response during human female fetal reproductive tract development

Abstract

The role of tissue interactions was explored to determine whether epithelial differentiation within the developing human reproductive tract is induced and specified by mesenchyme in tissue recombinants composed of mouse vaginal mesenchyme + human uterine tubal epithelium (mVgM+hTubE). The tissue recombinants were grown in DES-treated ovariectomized athymic mice. After 2-4 weeks of in vivo growth, several vaginal specific features were expressed in the human tubal epithelium. The mesenchyme-induced effects included morphological change as well as expression of several immunohistochemical markers. Although the mesenchyme-induced shift in vaginal differentiation in the human tubal epithelium was not complete, the partial induction of vaginal markers in human tubal epithelium verifies the importance of mesenchymal-epithelial interactions in development of the human female reproductive tract. In a separate experiment, DES-induction of uterine epithelial progesterone receptor (PGR) and estrogen receptor 1 (ESR1) was explored in tissue recombinants composed of wild-type or Esr1KO mouse uterine mesenchyme + human fetal uterine epithelium (wt UtM+hUtE and Esr1KO UtM+hUtE). The rationale of this experiment was to determine whether DES-induction of PGR and ESR1 is mediated directly via epithelial ESR1 or indirectly (paracrine mechanism) via mesenchymal ESR1. DES-induction of uterine epithelial ESR1 and PGR in Esr1KO UtM+hUtE tissue recombinants (devoid of mesenchymal ESR1) formally eliminates the paracrine mechanism and demonstrates that DES induction of human uterine epithelial ESR1 and PGR is directly mediated via epithelial ESR1.

Keywords: Diethylstilbestrol; Estrogen receptor; Estrogenic response; Human female fetal reproductive tract; Mesenchymal-epithelial interactions; Progesterone receptor.

Figure 1

Section of a tissue recombinant composed of neonatal mouse vaginal mesenchyme plus 13 week human fetal uterine tube epithelium (mVgM+hTubE) stained with Hoechst dye 33258 to verify the species origin of the mesenchyme and epithelium. Mouse nuclei contain many bright chromatin bodies, whereas human nuclei lack such intra-nuclear bodies (Cunha and Vanderslice, 1984). The red-circled pycnotic nucleus in the lower magnification images can be seen at higher magnification.

Figure 2

Tissue recombinants composed of neonatal mouse vaginal mesenchyme plus 13 week human fetal uterine tube epithelium (mVgM+hTubE) grown for 4 weeks in DES-treated hosts and immunostained for various vaginal epithelial markers as indicated. Human uterine tube (A, D, G, J) and vagina (B, E, H, K) at 16 to 18 weeks of gestation serve as controls. Note induction of KRT6, TP63 and RUNX1 and down regulation of AR in epithelium of the mVgM+hTubE recombinants, indicative of an effect of mouse vaginal mesenchyme on expression of differentiation markers in human tubal epithelium. (+) and (−) indicate epithelial marker expression.

Figure 3

Sections of developing human fetal uterine corpus immunostained for ESR1 at the ages indicated (A-D). At all stages (8 to 18 weeks) ESR1 is undetectable in the uterine epithelium. At 18 weeks, the stroma is ESR1-positive. (E & F) are sections of grafts of 10 week (E) and 13 week (F) human fetal uterine corpus grown for 4 weeks in DES-treated ovariectomized hosts and immunostained for ESR1. Note induction of epithelial ESR1.

Figure 4

Tissue recombinants composed of neonatal mouse wild-type uterine mesenchyme plus human fetal uterine tube epithelium (wt UtM+hUtE) (A &B) and aERKO uterine mesenchyme plus human uterine tube epithelium Esr1KO UtM+ h UtE) (C & D) grown in DES-treated hosts and immunostained for ESR1 (A & C) and PGR (B & D). DES induced ESR1 and PGR even when the mesenchyme was genetically devoid of ESR1. Sections (C) and (D) are adjacent sections stained for ESR1 (C) and PGR (D). (E) is a section of a graft of Esr1KO UtM+hUtE tissue recombinants stained with Hoechst dye 33258 to verify the tissue origin.

Figure 5

Sections of non-grafted human female fetal reproductive tracts (A and B) at the ages specified immunostained for PGR. (C) A 13 week human fetal uterine corpus grown for 4 weeks in DES-treated ovariectomized hosts and immunostained for PGR. Note DES induction of epithelial and stromal PGR.