No abstract

Ovarian carcinomas are thought to arise in the ovarian surface epithelium (OSE). Although this tissue forms a simple epithelial covering on the ovarian surface, OSE cells exhibit some mesenchymal characteristics and contain little or no E-cadherin. However, E-cadherin is present in metaplastic OSE cells that resemble the more complex epithelia of the oviduct, endometrium and endocervix, and in primary epithelial ovarian carcinomas. To determine whether E-cadherin was a cause or consequence of OSE metaplasia, we expressed this cell-adhesion molecule in simian virus 40-immortalized OSE cells. In these cells the exogenous E-cadherin, all three catenins, and F-actin localized at sites of cell-cell contact, indicating the formation of functional adherens junctions. Unlike the parent OSE cell line, which had undergone a typical mesenchymal transformation in culture, E-cadherin-expressing cells contained cytokeratins and the tight-junction protein occludin. They also formed cobblestone monolayers in two-dimensional culture and simple epithelia in three-dimensional culture that produced CA125 and shed it into the culture medium. CA125 is a normal epithelial-differentiation product of the oviduct, endometrium, and endocervix, but not of normal OSE. It is also a tumor antigen that is produced by ovarian neoplasms and by metaplastic OSE. Thus, E-cadherin restored some normal characteristics of OSE, such as keratin, and it also induced epithelial-differentiation markers associated with weakly preneoplastic, metaplastic OSE and OSE-derived primary carcinomas. The results suggest an unexpected role for Ecadherin in ovarian neoplastic progression.

Our group recently described recurrent somatic mutations of the miRNA processing gene DICER1 in non-epithelial ovarian cancer. Mutations appeared to be clustered around each of four critical metal-binding residues in the RNase IIIB domain of DICER1. This domain is responsible for cleavage of the 3' end of the 5p miRNA strand of a pre-mRNA hairpin. To investigate the effects of these cancer-associated 'hotspot' mutations, we engineered mouse DICER1-deficient ES cells to express wild-type and an allelic series of the mutant DICER1 variants. Global miRNA and mRNA profiles from cells carrying the metal-binding site mutations were compared to each other and to wild-type DICER1. The miRNA and mRNA profiles generated through the expression of the hotspot mutations were virtually identical, and the DICER1 hotspot mutation-carrying cells were distinct from both wild-type and DICER1-deficient cells. Further, miRNA profiles showed that mutant DICER1 results in a dramatic loss in processing of mature 5p miRNA strands but were still able to create 3p strand miRNAs. Messenger RNA (mRNA) profile changes were consistent with the loss of 5p strand miRNAs and showed enriched expression for predicted targets of the lost 5p-derived miRNAs. We therefore conclude that cancer-associated somatic hotspot mutations of DICER1, affecting any one of four metal-binding residues in the RNase IIIB domain, are functionally equivalent with respect to miRNA processing and are hypomorphic alleles, yielding a global loss in processing of mature 5p strand miRNA. We further propose that this resulting 3p strand bias in mature miRNA expression likely underpins the oncogenic potential of these hotspot mutations.

The ovarian surface epithelium (OSE) is the precursor of common epithelial ovarian carcinomas. In the present study, we examined the molecular mechanisms and possible physiological basis for the propensity of OSE cells to undergo epithelio-mesenchymal transition (EMT) in response to environmental influences. We hypothesized that EMT may be a homeostatic mechanism that permits displaced OSE to assume a stromal phenotype within the ovarian cortex. We report that EGF in conjunction with hydrocortisone is the EMT-inducing factor of OSE as shown by changes to a fibroblast-like morphology and growth pattern. EGF increased cell motility, enhanced the activities of secreted pro-matrix metalloproteinase (MMP)-2 and -9, and enhanced expression and activation of Erk and integrin-linked kinase (ILK). Increased ILK expression correlated with the activation of PKB/Akt, the phosphorylation of GSK-3␤, and the increased expression of cyclin E and cdk2 kinase. EGF withdrawal resulted in a more epithelial morphology and reversal of the EGF-induced activation of signaling pathways and pro-MMP activity. In contrast, treatment of EGF-treated cells with specific inhibitors of phosphatidylinositol 3-kinase, Mek, or ILK inhibited the inhibitor-specific pathways. The inhibitors caused suppression of EGF-induced migration and pro-MMP-2/-9 activities but did not lead to any change in EGF-induced mesenchymal morphology. ILK small interfering RNA inhibited Akt phosphorylation and reduced pro-MMP-2/-9 activities but had no effect on Erk activation or cell morphology. These results indicate that the EGF-induced morphological and functional changes in OSE cells are controlled by distinct signaling mechanisms working in concert. EMT of OSE cells displaced by ovulation likely permits their survival and integration with a fibroblast-like identity within the stroma. Failure to do so may lead to the formation of epithelium-derived inclusion cysts, which are known preferential sites of malignant transformation. epidermal growth factor; migration; invasion THE OVARIAN SURFACE EPITHELIUM (OSE) is a single, flat to cuboid layer of cells supported on the ovarian surface by a basement membrane and tunica albuginea. The cells are held together laterally by desmosomes and tight junctions (31). Surface epithelial cells are continuous with the mesothelium of the ovarian ligament and the peritoneum. This single layer of epithelial cells contributes to ovulation by lysis and reconstruction of the ovarian cortex and is of major importance in gynecological pathology because it is thought to be the source of 90% of ovarian neoplasms, the epithelial ovarian carcinomas (5, 28, 30). OSE-derived epithelial inclusion cysts are the preferred sites for the origin of epithelial ovarian cancer. These observations emphasize the need to determine the mechanisms regulating the postovulatory fate of OSE and to better understand the initiation of its neoplastic transformation as well as the biology of ovulation.Sequential interaction of cellular signaling pathways is operative ...