Project description:Background. Fallopian tube secretory epithelial cells (FTSECs) have been implicated as a cell-of-origin for high-grade serous epithelial ovarian cancer. However, there are relatively few in vitro models of this tissue type available for use in studies of FTSEC biology and malignant transformation. In vitro three-dimensional (3D) cell culture models aim to recreate the architecture and geometry of tissues in vivo and restore the complex network of cell-cell/cell-matrix interactions that occur throughout the surface of the cell membrane. Results. We have established and characterized 3D spheroid culture models of primary FTSECs. FTSEC spheroids contain central cores of hyaline matrix surrounded by mono- or multi-layer epithelial sheets. We found that 3D culturing alters the molecular characteristics of FTSECs compared to 2D cultures of the same cells. Gene expression profiling identified more than a thousand differentially expressed genes between 3D and 2D cultures of the same FTSEC lines. Pathways significantly under-represented in 3D FTSEC cultures were associated with cell cycle progression and DNA replication. This was also reflected in the reduced proliferative indices observed in 3D spheroids stained for the proliferation marker MIB1. Comparisons with gene expression profiles of fresh fallopian tube tissues revealed that 2D FTSEC cultures clustered with follicular phase tubal epithelium, whereas 3D FTSEC cultures clustered with luteal phase samples. Conclusions. This 3D model of fallopian tube secretory epithelial cells will advance our ability to study the underlying biology and etiology of fallopian tube tissues and the pathogenesis of high-grade serous epithelial ovarian cancer. 3 primary FTSEC lines were plated in 2D, or in 3D on polyHEMA coated plates
Project description:Background: Fallopian tube secretory epithelial cells (FTSECs) have been implicated as a cell-of-origin for high-grade serous epithelial ovarian cancer. However, there are relatively few in vitro models of this tissue type available for use in studies of FTSEC biology and malignant transformation. In vitro three-dimensional (3D) cell culture models aim to recreate the architecture and geometry of tissues in vivo and restore the complex network of cell-cell/cell-matrix interactions that occur throughout the surface of the cell membrane. Results: We have established and characterized 3D spheroid culture models of primary FTSECs. FTSEC spheroids contain central cores of hyaline matrix surrounded by mono- or multi-layer epithelial sheets. We found that 3D culturing alters the molecular characteristics of FTSECs compared to 2D cultures of the same cells. Gene expression profiling identified more than a thousand differentially expressed genes between 3D and 2D cultures of the same FTSEC lines. Pathways significantly under-represented in 3D FTSEC cultures were associated with cell cycle progression and DNA replication. This was also reflected in the reduced proliferative indices observed in 3D spheroids stained for the proliferation marker MIB1. Comparisons with gene expression profiles of fresh fallopian tube tissues revealed that 2D FTSEC cultures clustered with follicular phase tubal epithelium, whereas 3D FTSEC cultures clustered with luteal phase samples. Conclusions: This 3D model of fallopian tube secretory epithelial cells will advance our ability to study the underlying biology and etiology of fallopian tube tissues and the pathogenesis of high-grade serous epithelial ovarian cancer.
Project description:The transcriptomes of three immortalized ovarian surface epithelial cell lines (iOSE, PMID: 17266044) and primary OSE cells (Innoprot, Derio, Spain) and four immortalized fallopian tube secretory epithelial (iFTE) cell lines (PMID: 21502498, 22936217) were compared. RNA-sequencing was done from rRNA depleted total RNA (Ribo-Zero rRNA Removal Kit) to approx. 20 million 50 bp paired end reads per sample. A discriminative gene expression signature comprised of 211 genes was developed and used to classify isolated and EpCAM enriched primary ovarian cancer cells (PMID: 25991672). Impact of this signature on overall survival was assessed from several publicly available ovarian cancer gene expression data sets. Background: High grade serous ovarian cancer (HGSOC) is characterized by extensive local, i.e. peritoneal, tumor spread, manifested in two different clinical presentations, miliary (many millet sized peritoneal implants) and non-miliary (few large exophytically growing peritoneal nodes), and an overall unfavorable outcome. HGSOC is thought to arise from fallopian tube secretory epithelial cells, via so called serous tubal intraepithelial carcinomas (STICs) but an ovarian origin was never ruled out for at least some cases. Comparative transcriptome analyses of isolated tumor cells from fresh HGSOC tissues and (immortalized) ovarian surface epithelial and fallopian tube secretory epithelial cell lines revealed a close relation between putative origin and tumor spread characteristic, i.e. miliary from tubes and non-miliary from ovaries.
Project description:The human fallopian tube harbors the cell-of-origin for the majority of high-grade serous ‘ovarian’ cancers (HGSCs), but its cellular composition, particularly of the epithelial component, is poorly characterized. We performed single-cell transcriptomic profiling in 12 primary fallopian specimens from 8 patients, analyzing around 53,000 individual cells to map the major immune, fibroblastic and epithelial cell types present in this organ. We identified 10 epithelial sub-populations, characterized by diverse transcriptional programs including SOX17 (enriched in secretory epithelial cells), TTF3 and RFX3 (enriched in ciliated cells) and NR2F2 (enriched in early, partially differentiated secretory cells). Based on transcriptional signatures, we reconstructed a trajectory whereby secretory cells differentiate into ciliated cells via a RUNX3high intermediate. Computational deconvolution of the cellular composition of advanced HGSCs based on epithelial subset signatures identified the ‘early secretory’ population as a likely precursor state for the majority of HGSCs. The signature of this rare population of cells comprised both epithelial (EPCAM, KRT) and mesenchymal (THY1, ACTA2) features, and was enriched in mesenchymal-type HGSCs (P = 6.7 x 10-27), a group known to have particularly poor prognoses. This cellular and molecular compendium of the human fallopian tube in cancer-free women is expected to advance our understanding of the earliest stages of fallopian epithelial neoplasia. PAX8, SOX17, MECOM, and WT1 are critical genes in high-grade ovarian cancer tumorigenesis. However, the target genes of these factors in a normal to tumor context is unknown. Depletion of these factors in fallopian tube secretory epithelial and high-grade serous ovarian cancer cells revealed transcription factor and cell-type dependent differential expression patterns.
Project description:We established fallopian tube epithelial organoids from human fallopian tube tissues and cultured them in the organoid culturing cocktail medium. To investigate the heterogeneity of proliferation in the fallopian tube epithelial organoids, we stained the fallopian tube epithelial cells by a fluorescent membrane dye PKH26. As culturing them, the intensity of PKH26 was reduced in accordance with the proliferative level of each cell. We sorted the organoid cells by FACS into PKH26-retained slow cycling cells and PKH26-reduced proliferative cells, and compared their transcriptomic characteristics by bulk RNA sequencing and gene set enrichment analysis (GSEA). The result of GSEA showed that several pathways related to cell cycle were downregulated in PKH26-retained cells. So this dataset is supposed to be suitable for the analysis of quiescent cells in human fallopian tube epithelium.
Project description:The human fallopian tube harbors the cell-of-origin for the majority of high-grade serous “ovarian” cancers (HGSCs), but its cellular composition, particularly of the epithelial component, is poorly characterized. We subjected 12 fallopian specimens from 8 patients to single-cell transcriptomic profiling, analyzing around 53,000 individual cells to map the major immune, fibroblastic and epithelial cell types present in this organ. We identified 10 epithelial sub-populations, characterized by diverse transcriptional programs including SOX17 (enriched in secretory epithelial cells), TTF3 and RFX3 (enriched in ciliated cells) and NR2F2 (enriched in early, partially differentiated secretory cells). Based on transcriptional signatures, we reconstructed a trajectory whereby secretory cells differentiate into ciliated cells via a RUNX3high intermediate. Computational deconvolution of the cellular composition of advanced HGSCs based on epithelial subset signatures identified the ‘early secretory’ population as a likely precursor state for the majority of HGSCs. The signature of this rare population of cells comprised both epithelial (EPCAM, KRT) and mesenchymal (THY1, ACTA2) features, and was enriched in mesenchymal-type HGSCs (P = 6.7 x 10e-27), a group known to have particularly poor prognoses. This cellular and molecular compendium of the human fallopian tube in cancer-free women is expected to advance our understanding of the earliest stages of fallopian epithelial neoplasia.
Project description:To investigate the role of norepinephrine on 3D spheroid formation and survival in early high-grade serous ovarian cancer, we cultured immortalized human fallopian tube epithelial cell lines in ultra-low attachment conditions in the presence or absence of norepinephrine and/or propranolol over 2 different timepoints.
Project description:The purpose of this study is to understand the effects of adrenergic signaling on the transcriptome of cell line models postulated to be the cells of origin of epithelial ovarian cancers using RNA-Seq. Here we explored the effects of the stress-related hormone, norepinephrine, on normal human ovarian and fallopian tube surface epithelial cellss. We investigated the early transcriptional response to norepinephrine in normal immortalized ovarian surface epithelial cells and fallopian tube secretory cells. RNA-Seq data of treated and untreated cells were analyzed to identify genes with differential expression.
Project description:Fallopian tube epithelium is the tissue-of-origin of most high grade serous papillary ovarian carcinoma. This tumor has been exensively investigated and sequenced but expression profiling data of normal fallopian tube epithelial cells is still rare. This project compares the miRNA profiles of high grade serous papillary ovarian tumors (FFPE and fresh frozen) to that of normal unmatched epithelial cells from resected fallopian tubes.