Isolation, growth and characteristics of human ovarian surface epithelium

M Nakamura, H Katabuchi, T Ohba, Y Fukumatsu, H Okamura
Virchows Archiv : an international journal of pathology · 1994 · doi:10.1007/BF00197394 · PMID:7981905
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This study describes an efficient method for isolating and culturing human ovarian surface epithelium cells, characterizing them as mesothelial cells suitable for further research.

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This paper aimed to establish an efficient in vitro method to isolate and culture human ovarian surface epithelium (OSE) and characterize its growth and cell features. Using scraping after 40 minutes of collagenase type 1 treatment on ovaries from 19 normal donors, the authors cultured surface cells in medium 199 with 15% fetal bovine serum and found viable cell yields of about 0.1–2.7×10^6 per ovary, with doubling times of 7–10 days and confluent monolayers forming after 13–20 days. Morphologically, monolayers were mostly cobblestone-like with fusiform or polygonal cells also observed, and cytochemical, immunocytochemical, and electron microscopy showed short-term mesothelial OSE characteristics. The authors’ main limitation, as reflected by their description, is that characterization was performed in short-term culture rather than long-term or mechanistic studies. This paper is centrally about endometriosis and adenomyosis because it describes OSE as a key tissue in ovarian endometriosis and uses the cultured OSE platform to investigate pathobiology and carcinogenesis relevant to pelvic disease.

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Abstract

The ovarian surface epithelium (OSE) is a key tissue in the pathogenesis of ovarian surface epithelial-stromal tumours and ovarian endometriosis, commonly encountered gynaecological diseases. Despite the high incidence of these diseases, experimental in vitro studies of OSE are few and so we used the scraping method with an enzymatic procedure to isolate human OSE and studied its characteristics in vitro. Nineteen normal ovaries were used. After incubation of the ovary for 40 min in collagenase type 1 solution (300 U/ml), the surface cells were removed by gentle scraping with a surgical blade. Cells obtained as a cluster after unit gravity sedimentation with 5% bovine serum albumin in medium 199 were cultured in medium 199 containing 15% fetal bovine serum. The viable cell number in a single ovary was 0.1-2.7 x 10(6). The outgrowth of cells started from a homogeneous population of single cells, and the cell population doubling time was between 7 and 10 days. Confluent monolayers were formed after 13-20 days and subcultured from one to three times. The monolayers mostly had a cobblestone appearance, and fusiform or polygonal cells were also observed. By cytochemistry, immunocytochemistry and scanning and transmission electron microscopy, the cells were shown to have characteristics of mesothelial OSE cells in short-term culture. This experimental approach was efficient in providing cultured human OSE, which can be utilized to investigate pathobiology and carcinogenesis.
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Abstract

The ovarian surface epithelium (OSE) is a key tissue in the pathogenesis of ovarian surface epithelial-stromal tumours and ovarian endometriosis, commonly encountered gynaecological diseases. Despite the high incidence of these diseases, experimental in vitro studies of OSE are few and so we used the scraping method with an enzymatic procedure to isolate human OSE and studied its characteristics in vitro. Nineteen normal ovaries were used. After incubation of the ovary for 40 min in collagenase type 1 solution (300 U/ml), the surface cells were removed by gentle scraping with a surgical blade. Cells obtained as a cluster after unit gravity sedimentation with 5% bovine serum albumin in medium 199 were cultured in medium 199 containing 15% fetal bovine serum. The viable cell number in a single ovary was 0.1−2.7×106. The outgrowth of cells started from a homogeneous population of single cells, and the cell population doubling time was between 7 and 10 days. Confluent monolayers were formed after 13–20 days and subcultured from one to three times. The monolayers mostly had a cobblestone appearance, and fusiform or polygonal cells were also observed. By cytochemistry, immunocytochemistry and scanning and transmission electron microscopy, the cells were shown to have characteristics of mesothelial OSE cells in short-term culture. This experimental approach was efficient in providing cultured human OSE, which can be utilized to investigate pathobiology and carcinogenesis. Similar content being viewed by others

References

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endometriosis

MeSH descriptors

Ovary Adult Cell Count Cell Division Cell Nucleus Cell Nucleus Cells, Cultured Cell Separation Collagenases Cytoplasm Cytoplasm Epithelium Epithelium Female Histocytochemistry Humans Immunohistochemistry Intercellular Junctions Intercellular Junctions Microscopy, Electron

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