Paracrine mediators of mouse uterine epithelial cell transepithelial resistance in culture.

Epithelial cell integrity at mucosal surfaces provides an effective physical barrier against potential pathogens that threaten reproductive health. We have used polarized epithelial cells from adult mouse uteri to investigate the roles of TNFalpha and TGFbeta, which are produced by uterine epithelial and stromal cells, and hepatocyte growth factor (HGF), produced by uterine stromal cells, in regulating epithelial cell integrity measured as transepithelial electrical resistance (TER). Exposure of epithelial cells to TNFalpha, TGFbeta, and HGF have profound effects on TER that are different from their known actions on TER at other mucosal surfaces. When incubated with TNFalpha, TER increased in a dose-dependent manner. In contrast, when cells were incubated with TGFbeta, TER was markedly but reversibly suppressed. Interestingly, HGF, when placed in the basolateral compartment, increased TER. Based on these findings, we conclude that TNFalpha, TGFbeta and HGF may play regulatory roles in modulating epithelial cell tight junctions. These studies suggest that factors, such as hormone balance, pathogen exposure as well as pregnancy, which affect cytokine and growth factor secretion, influence epithelial cell barrier protection within the female reproductive tract.

Effect of oestradiol on mouse uterine epithelial cell tumour necrosis factor-alpha release is mediated through uterine stromal cells.

Oestradiol-17beta (Oe(2)) stimulates uterine epithelial cell proliferation and is critical for normal uterine differentiation and secretory function. Oe(2) can act directly on the epithelium via the epithelial oestrogen receptor (OR) or indirectly via the OR-positive underlying stroma. A primary role for epithelial-stromal interactions has been established for mediating steroid hormone action in the uterus. This study was undertaken to determine the mode of Oe(2) action in regulating epithelial cell cytokine release in the uterus. Mouse uterine epithelial and stromal cells were isolated and cultured separately. Transepithelial resistance (TER) was monitored with an EVOM voltohmmeter to determine monolayer polarity and integrity. Epithelial cells grown alone or in coculture with stromal cells were treated with Oe(2). Supernatants collected were assayed for transforming growth factor-beta (TGF-beta) and tumour necrosis factor-alpha (TNF-alpha) by bioassay and enzyme-linked immunosorbent assay, respectively. While Oe(2) treatment of epithelial cells led to a significant decrease in TER, the amount of TNF-alpha released was not altered. However, when epithelial cells were cocultured with stromal cells and treated with Oe(2), apical TNF-alpha release was significantly decreased, compared to cells not treated with hormone. As determined by oestrogen receptor antagonist studies, Oe(2) primed epithelial cells for the action of the stromal paracrine factor(s). In contrast, TGF-beta release by epithelial cells was not affected by Oe(2) when grown alone or in the presence of stromal cells. These studies indicate that Oe(2) has both direct and indirect effects on the uterine epithelium. While epithelial monolayer integrity is directly influenced by Oe(2), TNF-alpha release in response to Oe(2) is dependent on the presence of stromal cells, indicating that paracrine communication is necessary for steroid regulation of some but not all cytokines.

Epithelial cells in the female reproductive tract: a central role as sentinels of immune protection.

The continued presence of bacterial and viral antigens in the lumen of the vagina coupled with the periodic presence of antigens in the lumen of the upper reproductive tract provide an ongoing challenge that can compromise female reproductive health and threaten life. Separating underlying tissues from luminal antigens, polarized epithelial cells of the cervix, uterus and Fallopian tubes have evolved to protect against potential pathogens. Once thought to function exclusively by providing a crucial barrier, mucosal epithelial cells are now known to function as sentinels that recognize antigens, respond in ways that lead to bacterial and viral killing, as well as signal to underlying immune cells when pathogenic challenge exceeds their protective capacity. Unique to epithelial cells of the female reproductive tract is the regulatory control of the female sex hormones. Acting both directly and indirectly through underlying stromal cells, estradiol and progesterone regulate epithelial cell innate and adaptive immune functions to protect against potential pathogens while providing an environment that supports an allogeneic fetus. In this article, we will outline how polarized epithelial cells function as the first line of defense against potential pathogens in the female reproductive tract.

Hepatocyte growth factor regulation of uterine epithelial cell transepithelial resistance and tumor necrosis factor alpha release in culture.

Underlying stromal cells are essential for the normal development of epithelial cells (ECs) at mucosal surfaces. Recent studies from our laboratory have shown that uterine stromal cells regulate EC integrity, measured as transepithelial resistance (TER) as well as tumor necrosis factor (TNF) alpha alpha secretion by ECs in culture. Using stromal cells in coculture with polarized ECs grown on inserts, we found that stromal cells produce soluble mediators that increase TER and decrease TNFalpha secretion. The purpose of the present study was to identify the mechanisms whereby stromal cells exert their effects on uterine epithelium. We report that hepatocyte growth factor (HGF), a known mesenchymal growth factor that mediates EC proliferation, increases TER but, at the same time, decreases apical TNFalpha release. When ECs and/or stromal cells were incubated with anti-HGF or anti-HGF receptor (HGFR) antibody before HGF, the effects of HGF were blocked. These findings indicate that ECs express the HGFR at their basolateral surfaces and that HGFR mediates the effects of HGF on TER and TNFalpha. Neutralization of stromal cell secretions with antibodies for HGF and HGFR demonstrate that stromal-derived HGF is the mediator of EC TER. In contrast, neither anti-HGF antibody nor HGFR antibody had any effect on stromal cell-induced decreases in TNFalpha secretion. From these results, we conclude that stromal cell regulation of EC TER is mediated through the secretion of stromal HGF. Furthermore, because neutralization of stromal media failed to affect TNFalpha secretion, these findings suggest that other growth factors, in addition to HGF, affect EC cytokine production.

Effect of estradiol on mouse uterine epithelial cell transepithelial resistance (TER).

PROBLEM: The effects of estradiol on epithelial cell function in the uterus may either be direct or indirect through the paracrine effects of underlying stromal cells. The aim of this study was to test whether estradiol-17beta (E(2)) acts directly to regulate uterine epithelial cell monolayer integrity. METHODS OF STUDY: Mouse uterine epithelial cells were isolated and grown on cell culture inserts to form confluent, polarized monolayers, as indicated by the development of high transepithelial resistance (TER). RESULTS: When polarized epithelial cells were treated with E(2), TER was significantly decreased within 24 hr of exposure. Epithelial cells remained hormonally responsive in culture for at least 10 days. In contrast to estradiol, incubation with progesterone, cortisol, aldosterone, and DHT had no effect on uterine epithelial cell TER. The ability of E(2) to decrease TER was inhibited following co-incubation with ICI 182,780, a pure estrogen receptor antagonist. To further investigate the mechanism involved in estradiol-induced decreases in TER, we tested the effect of TAPI-0, an inhibitor of matrix metalloproteinases. Our findings indicate that TAPI-0 reversed the inhibitory effect of E(2) on TER. CONCLUSIONS: These studies demonstrate that epithelial monolayer integrity is directly influenced by E(2) and ER mediated. Further, it suggests that the mechanism through which estradiol decreases TER is mediated by matrix metalloproteinases.