Effects of epidermal growth factor/hydrocortisone on the growth and differentiation of human ovarian surface epithelium.

OBJECTIVE: Ovarian surface epithelium (OSE), the precursor of the epithelial ovarian carcinomas, has limited growth potential in culture. Epidermal growth factor+hydrocortisone (EGF+HC) enhances its growth but induces epitheliomesenchymal transition (EMT). This study was undertaken to define the effects of EGF+HC and their reversibility, to optimize growth-promoting media, and to relate OSE phenotypes in vitro to physiologic states in vivo. METHODS: OSE was cultured in media 199/MDCB105 or EBM (Clonetics) with 2% or 10% fetal bovine serum with or without 10 ng/mL EGF, 1.0 microg/mL HC, and 1.0 microg/mL bovine brain extract. Growth rates and growth potentials (population doublings [PD] to senescence) were defined, and growth patterns and expression of keratin and collagen types III and IV were compared with the ovarian cancer cell lines OVCAR3 and SKOV3. RESULTS: EGF+HC increased growth potentials from 12-14 PD to 40-42 PD and reduced PD time from 53 hours to 20 hours. Without EGF+HC, OSE cells remained uniformly epithelial. EGF+HC induced EMT (mesenchymal shapes, reduced keratin, and production of collagenous extracellular matrix), but the EMT response varied greatly among OSE from different women. EMT was reversed over 1-2 weeks by subculture into EGF+HC-free medium in passage 1, but inconsistently thereafter. EGF+HC had no effect on the differentiation of ovarian carcinoma lines. CONCLUSION: The phenotype of intact OSE in vivo is most closely reproduced in media without EGF+HC. EGF+HC enhances growth but initiates EMT, which likely mimics a repair response. Variations in EGF+HC-induced phenotypes point to the existence of OSE subpopulations with differing responsiveness to growth factors or steroids, which may relate to their susceptibility to malignant transformation.

Epithelio-mesenchymal transition in a neoplastic ovarian epithelial hybrid cell line.

A hybrid cell line, IOSE-Ov29, was created through fusion of cells from the human ovarian adenocarcinoma line OVCAR3 and the non-tumorigenic SV40 Tag-transfected human ovarian surface epithelial line IOSE-29. OVCAR3 cells exhibit a differentiated epithelial phenotype, whereas line IOSE-29 expresses mesenchymal characteristics that were acquired in culture by epithelio-mesenchymal transition. Microsatellite analysis, comparative genomic hybridization (CGH), and MFISH showed the genotype of the IOSE-Ov29 cells to contain components of both parent cell lines, but to be predominantly OVCAR3 derived. IOSE-Ov29 resembled OVCAR3 and differed from IOSE-29 as shown by its unlimited life span, tumorigenicity, epithelial morphology, keratin, occludin, E-cadherin and CA125 expression, increased expression of kinases of the PI3K pathway, and loss of cGMP-dependent protein kinase expression. IOSE-29-derived properties included SV40 Tag expression, growth inhibition by activin, collagen type III secretion, increased adhesion and spreading on tissue culture plastic, and increased growth rate. Proliferation of all three lines was stimulated by FSH and ATP and inhibited by GnRH I and GnRH II. Interestingly, IOSE-Ov29 was more anchorage independent than either parent line and was the only line that invaded Matrigel in Boyden chambers and formed invasive branches in collagen gels. The results indicate that IOSE-Ov29 is an IOSE-29/OVCAR3 hybrid, which differs from both parent lines genetically and phenotypically. Unexpectedly, fusion with the non-tumorigenic IOSE-29 cells enhanced malignancy-associated characteristics of OVCAR3, presumably as a result of the expression of IOSE-29-derived mesenchymal properties that are usually acquired by carcinoma cells through epithelio-mesenchymal transition during metastatic progression.