EGF-Induced EMT and Invasiveness in Serous Borderline Ovarian Tumor Cells: A Possible Step in the Transition to Low-Grade Serous Carcinoma Cells?

In high-grade ovarian cancer cultures, it has been shown that epidermal growth factor (EGF) induces cell invasion by activating an epithelial-mesenchymal transition (EMT). However, the effect of EGF on serous borderline ovarian tumors (SBOT) and low-grade serous carcinomas (LGC) cell invasion remains unknown. Here, we show that EGF receptor (EGFR) was expressed, that EGF treatment increased cell migration and invasion in two cultured SBOT cell lines, SBOT3.1 and SV40 large T antigen-infected SBOT cells (SBOT4-LT), and in two cultured LGC cell lines, MPSC1 and SV40 LT/ST-immortalized LGC cells (ILGC). However, EGF induced down-regulation of E-cadherin and concurrent up-regulation of N-cadherin in SBOT cells but not in LGC cells. In SBOT cells, the expression of the transcriptional repressors of E-cadherin, Snail, Slug and ZEB1 were increased by EGF treatment. Treatment with EGF led to the activation of the downstream ERK1/2 and PI3K/Akt. The MEK1 inhibitor PD98059 diminished the EGF-induced cadherin switch and the up-regulation of Snail, Slug and ZEB1 and the EGF-mediated increase in SBOT cell migration and invasion. The PI3K inhibitor LY294002 had similar effects, but it could not block the EGF-induced up-regulation of N-cadherin and ZEB1. This study demonstrates that EGF induces SBOT cell migration and invasion by activating EMT, which involves the activation of the ERK1/2 and PI3K/Akt pathways and, subsequently, Snail, Slug and ZEB1 expression. Moreover, our results suggest that there are EMT-independent mechanisms that mediate the EGF-induced LGC cell migration and invasion

Interaction of cellular and environmental factors in tumor histogenesis

The histogenesis of two squamous-carcinoma cell lines, originally derived from the same human cervical tumor but with different morphology in vitro and in vivo, was compared histochemically and electron microscopically. Inoculation into hamster cheek pouches was used to distinguish in vitro from in vivo responses in the two cell lines. Normal cervical epithelium was grown in vitro to determine which histogenetic characteristics were due to the malignant nature of the cell lines. Possible mechanisms underlying histogenetic variation were investigated by chemical and physical modifications of the in vitro environment. In vitro cells of line C-4c were more cohesive, less deformable and less adhesive to substrata than the cells of line C-4s and they formed more compact and highly stratified colonies. Line C-4s responded to crowding in vitro by cell separation (dispersal) while line C-4c increased stratification. Histochemically the cell lines were essentially similar but they differed in ultrastructure, particularly at the level of tissue organization. The C-4s cells were columnar, with a higher nucleo-cytoolasmic ratio, a polarity of the organelles, dispersed cytoplasmic filaments, little stratification and with terminal bars between superficial cells. In contrast, C-4c cells were oval, stratified and with superficial cells flattened, with a lower nucleo-cytoplas-mic ratio, without polarity of organelles, and with more cytoplasmic filaments condensed into bundles that were associated with desrnosomes. In both lines, but particularly in line C-4c, interdigitating microvilli provided the main intercellular contact and the cell surfaces were modified in association with both substrata and the growth medium. Most in vitro differences between the cell lines were retained in vivo, and, in addition, incomplete basement membranes were formed in both lines, though more extensively in line C-4.s. It appeared that the only major difference in vitro between benign squamous cells and the carcinoma cells lay in the ability of the latter to maintain an intercellular organization in the complete absence of any supporting tissue. Possible mechanisms underlying these observed differences in histogenesis were experimentally investigated. Ferritin uptake in vitro indicated that the difference in stratification was not due to more efficient intercellular circulation in the more highly stratified C-4c line. No difference in cell-surface charges or distribution of cell-surface acid mucosubstances could be demonstrated, suggesting no difference in contact inhibition to be associated with the stratification. It was found that C-4c cells were more cohesive and that this cohesion, in both lines, depended predominantly on the presence of divalent cations. In contrast, adhesion to the substrata required extracellular proteins, probably accompanied by the masking of acid groups on the cell surface. These and other described observations suggested a defect in deformability in these cell lines with the probability that the stronger cohesion in line C-4c was due to the more extensive microvillus population. The in vitro maintenance of cell shape, of tissue organization and of cell separation (dispersal) seemed to involve an interaction between these intercellular adhesive forces and the cytoplasmic filaments. On the other hand, the specific, modifications, such as terminal-bar formation and cell flattening, which appeared in relation to growth medium, to other cells and to substrata could be shown to be responses of the cells to the viscosity of the immediate environment and thus experimentally modifiable. The results of this investigation are discussed relative to the specific suggestion that much of the histogenetic variation could be explained by considering that the more cohesive cell line (C-4c) had retained properties of normal epithelium stratum spinosum cells, while the spreading cell line (C-4s) exhibited characteristics of basal cells, and relative to the interaction of differentiation capacity and malignancy in tumor development.Science, Faculty ofZoology, Department ofGraduat

HOXA4 protein levels and localization in the aorta and in human abdominal aortic aneurysms

Abstract This report presents evidence for the specificities of select commercially available HOXA4 antibodies in regards to concerns about the specificity of the HOXA4 antibody used by Lillvis et al. (Regional expression of HOXA4 along the aorta and its potential role in human abdominal aortic aneurysms. BMC Physiol 2011, 11:9). Using an antibody characterized extensively by us, Lillvis et al. report detecting HOXA4 at a size of 33 kDa despite our previous reports that HOXA4 is detected at ~37-39 kDa and that the ~30-33 kDa band is non-specific. Using small interfering RNA targeting HOXA4, forced expression of full-length HOXA4 and HOXA4-positive and -negative ovarian cancer cell lines, we confirm our previous findings that the ~30-33 kDa band is non-specific and that HOXA4 is detected at ~37-39 kDa. Moreover, we demonstrate that HOXA4 small interfering RNA reduces the ~37-39 kDa HOXA4 band, but not the ~30-33 kDa non-specific band, in a human acute monocytic leukemia cell line used by Lillvis et al. Western blot analysis performed with two additional commercially available HOXA4 antibodies also detected HOXA4 at ~37-39 kDa. Lastly, immunofluorescent staining of a HOXA4-negative ovarian cancer cell line with the antibody used by Lillvis et al. yields strong perinuclear staining, similar to that observed by Lillvis et al., which cannot be attributed to HOXA4. Our results highlight and briefly discuss the importance of careful antibody validation and selection for use in various applications.Medicine, Faculty ofObstetrics and Gynaecology, Department ofReviewedFacult

Ovarian surface epithelium: family history and early events in ovarian cancer

Ovarian cancer is the most common cause of death from gynecological cancers in the Western world. There are many genetic and environmental factors which can influence a woman's risk of getting ovarian cancer. A strong family history of breast or ovarian cancer is definitely one of the most important and best-defined epidemiological risk factors. This review evaluates current knowledge of hereditary ovarian cancer. Histologic, cytologic and molecular studies on the ovarian surface epithelium (OSE), which is the origin of ovarian epithelial carcinomas, from women with a strong family history for ovarian carcinomas or with a mutation in one of the two known cancer susceptibility genes – BRCA1 and BRCA2, provide a background to facilitate understanding of the early changes in ovarian carcinogenesis. This overview is followed by a discussion of recent hypotheses and research on two questions. First, is there a mutational hotspot of BRCA mutation for ovarian cancer? Second, why do mutations in BRCA1 and BRCA2, which are ubiquitously expressed genes that participate in general cellular activities, lead preferentially to breast and ovarian cancer?Medicine, Faculty ofObstetrics and Gynaecology, Department ofNon UBCReviewedFacult

Mitogen-activated protein kinases in normal and (pre)neoplastic ovarian surface epithelium

Mitogen-activated protein kinases (MAPKs) are a group of serine/threonine kinases which are activated in response to a diverse array of extracellular stimuli and mediate signal transduction from the cell surface to the nucleus. It has been demonstrated that MAPKs are activated by external stimuli including chemotherapeutic agents, growth factors and reproductive hormones in ovarian surface epithelial cells. Thus, the MAPK signaling pathway may play an important role in the regulation of proliferation, survival and apoptosis in response to these external stimuli in ovarian cancer. In this article, an activation of the MAPK signaling cascade by several key reproductive hormones and growth factors in epithelial ovarian cancer is reviewed.Medicine, Faculty ofObstetrics and Gynaecology, Department ofNon UBCReviewedFacult