Proteomic Analysis of Stromal and Epithelial Cell Communications in Human Endometrial Cancer Using a Unique 3D Co-Culture Model.

Epithelial and stromal communications are essential for normal uterine functions and their dysregulation contributes to the pathogenesis of many diseases including infertility, endometriosis, and cancer. Although many studies have highlighted the advantages of culturing cells in 3D compared to the conventional 2D culture system, one of the major limitations of these systems is the lack of incorporation of cells from non-epithelial lineages. In an effort to develop a culture system incorporating both stromal and epithelial cells, 3D endometrial cancer spheroids are developed by co-culturing endometrial stromal cells with cancerous epithelial cells. The spheroids developed by this method are phenotypically comparable to in vivo endometrial cancer tissue. Proteomic analysis of the co-culture spheroids comparable to human endometrial tissue revealed 591 common proteins and canonical pathways that are closely related to endometrium biology. To determine the feasibility of using this model for drug screening, the efficacy of tamoxifen and everolimus is tested. In summary, a unique 3D model system of human endometrial cancer is developed that will serve as the foundation for the further development of 3D culture systems incorporating different cell types of the human uterus for deciphering the contributions of non-epithelial cells present in cancer microenvironment.

Human ovarian cancer stem/progenitor cells are stimulated by doxorubicin but inhibited by Mullerian inhibiting substance.

Women with late-stage ovarian cancer usually develop chemotherapeutic-resistant recurrence. It has been theorized that a rare cancer stem cell, which is responsible for the growth and maintenance of the tumor, is also resistant to conventional chemotherapeutics. We have isolated from multiple ovarian cancer cell lines an ovarian cancer stem cell-enriched population marked by CD44, CD24, and Epcam (3+) and by negative selection for Ecadherin (Ecad-) that comprises less than 1% of cancer cells and has increased colony formation and shorter tumor-free intervals in vivo after limiting dilution. Surprisingly, these cells are not only resistant to chemotherapeutics such as doxorubicin, but also are stimulated by it, as evidenced by the significantly increased number of colonies in treated 3+Ecad- cells. Similarly, proliferation of the 3+Ecad- cells in monolayer increased with treatment, by either doxorubicin or cisplatin, compared with the unseparated or cancer stem cell-depleted 3-Ecad+ cells. However, these cells are sensitive to Mullerian inhibiting substance (MIS), which decreased colony formation. MIS inhibits ovarian cancer cells by inducing G1 arrest of the 3+Ecad- subpopulation through the induction of cyclin-dependent kinase inhibitors. 3+Ecad- cells selectively expressed LIN28, which colocalized by immunofluorescence with the 3+ cancer stem cell markers in the human ovarian carcinoma cell line, OVCAR-5, and is also highly expressed in transgenic murine models of ovarian cancer and in other human ovarian cancer cell lines. These results suggest that chemotherapeutics may be stimulative to cancer stem cells and that selective inhibition of these cells by treating with MIS or targeting LIN28 should be considered in the development of therapeutics.

The Drosophila homolog of human tumor suppressor TSC-22 promotes cellular growth, proliferation, and survival.

TSC22D1, which encodes transforming growth factor beta-stimulated clone 22 (TSC-22), is thought to be a tumor suppressor because its expression is lost in many glioblastoma, salivary gland, and prostate cancers. TSC-22 is the founding member of the TSC-22/DIP/Bun family of leucine zipper transcription factors; its functions have not been investigated in a multicellular environment. Genetic studies in the model organism Drosophila melanogaster often provide fundamental insights into mechanisms disrupted in carcinogenesis, because of the strong evolutionary conservation of molecular mechanisms between flies and humans. Whereas humans and mice have four TSC-22 domain genes with numerous isoforms, Drosophila has only one TSC-22 domain gene, bunched (bun), which encodes both large and small protein isoforms. Surprisingly, Drosophila Bun proteins promote cellular growth and proliferation in ovarian follicle cells. Loss of both large isoforms has the strongest phenotypes, including increased apoptosis. Cultured S2 cells depleted for large Bun isoforms show increased apoptosis and less frequent cell division, with decreased cell size. Altogether, these data indicate that Drosophila TSC-22/DIP/Bun proteins are necessary for cellular growth, proliferation, and survival both in culture and in an epithelial context. Previous work demonstrated that bun prevents recruitment of epithelial cells to a migratory fate and, thus, maintains epithelial organization. We speculate that reduced TSC22D1 expression generally reduces cellular fitness and only contributes to carcinogenesis in specific tissue environments.

In vivo evidence of role of bone morphogenetic protein-4 in the mouse ovary.

The transition of a primordial follicle to a primary follicle is an early step in folliculogenesis. All female mammals are born with a fixed stock of primordial follicles, and exhaustion of that stock leads to menopause or infertility. Recently, several in vitro studies have indicated that BMP-4, BMP-7, and several other growth factors affect the transition of primordial to primary follicles. The aim of our present study was to investigate role of BMP-4 in this process using passive immunization to investigate the role of BMP-4 in a prepubertal mouse model. After seven days of treatment, the weight of antiBMP-4 treated ovaries was significantly lower than the ovaries from mice treated with nonimmune Ig. The number of primary follicles was lower, and the numbers of primordial follicles were higher in antiBMP-4 treated ovaries compared to control ovaries. Treatment with equine chorionic gonadotrophin (eCG) showed no influence on the effects of antiBMP-4 in the mouse ovary. Thus, the results of our study indicate that in vivo BMP-4 acts as transition factor in transition of primordial to primary follicle.

Drosophila follicle cells: morphogenesis in an eggshell.

Epithelial morphogenesis is important for organogenesis and pivotal for carcinogenesis, but mechanisms that control it are poorly understood. The Drosophila follicular epithelium is a genetically tractable model to understand these mechanisms in vivo. This epithelium of follicle cells encases germline cells to create an egg. In this review, we summarize progress toward understanding mechanisms that maintain the epithelium or permit migrations essential for oogenesis. Cell-cell communication is important, but the same signals are used repeatedly to control distinct events. Understanding intrinsic mechanisms that alter responses to developmental signals will be important to understand regulation of cell shape and organization.