Joy Mulholland

Progesterone Directed Gene Expression in Rat Uterine Stromal Cells

When embryos attach to the luminal epithelium in the rat uterus they somehow induce differentiation of the stromal cells underlying the attachment site, giving rise to an enormous increase in the size of the stromal cell compartment through both cell proliferation and cell growth. This reaction, decidualization, is localized to areas of embryo attachment (reviewed in DeFeo, 1967). Both attachment of the embryo to the luminal epithelium and differentiation of the stromal cells in response to embryo attachment require conditioning of the uterus with steroid hormones (Psychoyos, 1973). If the uterus is hormonally prepared, decidualization can be obtained using an intraluminal traumatic or chemical artificial stimulus. In response to these stimuli stromal cells along the entire length of the uterus will differentiate (DeFeo, 1967).

Morphological and immunohistochemical differentiation patterns of rabbit uterine epithelium in vitro

We describe morphological and immunohistochemical changes of uterine epithelium from immature rabbits in vitro in response to hormonal treatments, using a matrix-coated semipermeable filter. These investigations were compared to in vivo studies of uterine epithelium from immature rabbits treated with estrogen and/or progesterone. In vitro, polarization of the epithelium seems to be best developed under progesterone dominance, and the pattern of cell organelles is similar to those seen in vivo. Two types of apical protrusions could be observed in cultures treated with progesterone, some shaped like domes, containing cell organelles, and some irregular in shape with small lucent vesicles. Both types of apical differentiation are typical for the in vivo situation. In vitro, estrogen leads to a more pseudostratified growth pattern of the cells. They develop apical protrusions with big vesicles probably containing mucin, as in vivo. Treatment with both steroid hormones leads to a heterogeneous response of the uterine epithelial cells in culture, some cells responding more to the estrogen, others to the progesterone whereas in vivo the progesterone-dominant features are obvious. Immunohistochemistry of uteroglobin in monensin-treated cultures gives evidence for uteroglobin secretion in all cultures, but to a lesser extent in the untreated, and this is strongly increased in cultures treated with estrogen and progesterone. These results correspond to observations made in vivo. This in vitro cell culture method seems therefore to provide a useful model for investigating the regulatory mechanisms of sexual steroid hormones and the cell biology of uterine receptivity.

In Vitro Models of Implantation

Blastocysts are capable of attaching to a uterus only after it has been appropriately prepared by ovarian steroid hormones (receptive uterus). 1,2 They can, however, attach to extrauterine sites regardless of the hormonal environment.3 These data suggest that under certain conditions the apical surface of the uterine epithelial (UE) cell expresses molecules (ligands) which do not permit blastocyst attachment (non-receptive). It has been proposed that interactions between UE cells and various regulatory agents (hormones, growth factors) effect (induce, stimulate, repress) structural and functional changes at the apical surface of the UE cell that allow nidation.

Effects of tamoxifen and ICI 164384 on protein synthesis and vectorial secretion in polarized rat uterine epithelial cells

A polarized, primary cell, bicameral culture system was utilized to test the effects of two types of antiestrogens, tamoxifen (class I) and ICI 164384 (class II) on post-mitotic, uterine epithelial cells. The results demonstrate that in addition to blocking estrogen action in a dose-dependent manner, each of these compounds has independent effects on protein synthesis and secretion in these cells. The effects of both tamoxifen and ICI 164384 on vectorial protein secretion were completely opposite to those of estradiol. Both antiestrogens given alone repressed apical secretion and stimulated basal secretion, whereas estradiol stimulated apical and repressed basal secretion. Furthermore, specific protein bands in both the apical and basal secretory compartments responded differently according to dose to each compound. These experiments using polarized uterine epithelial cell cultures have identified apical and basal protein secretion as two cellular response with increased sensitivity to steroids and antisteroids.

Uterine Preparation for Blastocyst Attachment

Existing models used to analyze uterine receptivity to blastocyst attachment have thus far failed to account for non-genotypic embryonic loss following in vitro fertilization and embryo transfer in any species. Although application of the technology for biochemistry and molecular biology has yielded useful information about the uterus at a molecular level, the regulatory mechanisms of ovoreceptivity and implantation remain elusive.

Development of suspension cultures for the study of epithelial cell polarity

Abstract Polarity is an important characteristic of all epithelial cells which plays a role in the regulation of vectorial transport of ions, proteins, and carbohydrates. Examination of cells exposed to microgravity during spaceflight indicates that cell polarity may be affected by the loss of gravity, but these effects have not yet been investigated directly. To maintain polarized cell functions in epithelial cells grown in cell culture, special cell culture substrata are usually required. As it is difficult to adapt this culture technique to the constraints of spaceflight hardware, alternative methods to maintain polarized epithelial cells in culture would facilitate the study of cell polarity in microgravity. The principal aim of the research project described below is to develop a novel cell culture system which allows hormon‐ally responsive, secretory epithelial cells to develop and maintain polarity when grown in suspension. If successful, it will be possible to utilize this culture system with exis...

Growth factor effects on endometrial epithelial cell differentiation and protein synthesis in vitro*†*Supported by The Women’s Fund, Houston, Texas, and by grant HD25189 from the National Institutes of Health, Bethesda, Maryland.†Presented at the 47th Annual Meeting of The American Fertility Society, Orlando, Florida, October 19 to 24, 1991.

OBJECTIVE To develop a baseline for projected studies of a rat endometriosis model. DESIGN We investigated the effects of two macrophage-related growth factors, platelet-derived growth factor (PDGF) and transforming growth factor-beta (TGF-beta), on proliferation, in vitro differentiation, and protein secretion of uterine epithelial cells from immature rats. Uterine epithelial cells grown on matrix covered filters were treated with growth factors (GFs) or estrogen and/or P. Incorporation of [35S]methionine by polarized uterine epithelial cell proteins and secretion of labeled proteins into apical and basal culture medium were examined. SETTING Department of Cell Biology, Baylor College of Medicine, Houston, Texas. MAIN OUTCOME MEASURES Cell associated and secreted proteins were resolved by gel electrophoresis, fluorography, and immunoblotting. Proliferation was quantified by cell counts in parallel cultures by hemocytometer. RESULTS Estrogen and P increase protein synthesis by uterine epithelium. Transforming growth factor-beta depressed protein synthesis and secretion in uterine epithelial cells. Platelet-derived growth factor appears to have no effect on epithelial protein synthesis or secretion and does not modulate the effect of TGF-beta. Estrogen and P increase complement component 3 (C3) production by epithelial cells. CONCLUSION Macrophage-secreted GFs may play a role in the development and maintenance of ectopic endometrial tissue. Both TGF-beta and ovarian steroids may participate in the dynamic regulation of protein synthesis by ectopic uterine epithelium. These molecules may indirectly affect the macrophage-stromal axis through nonspecific modulation of C3 secretion. Platelet-derived growth factor appears to have no direct effect on uterine epithelial cells. The recognized effect of PDGF on ectopic endometrial tissue is most likely mediated via the stromal component.