Primary culture of uterine cells: Markers of growth and differentiation

Field, Raymond Paul (1985) Primary culture of uterine cells: Markers of growth and differentiation. PhD thesis, University of Glasgow.

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Abstract

Epithelial and fibroblastic cells were isolated from immature rat uteri by enzymic disaggregation, and established as primary cultures. Study of autoradiographs from cultures of epithelial cells that were labelled with tritiated thymidine, showed a variable heterogeneity in the distribution of labelled and unlabelled cells. This was interpreted as reflecting the distributions of proliferating and quiescent cells. In general, more flattened cells located towards the edge of colonies, proliferated more rapidly than more rounded cells towards the centre of colonies. Culture of epithelial cells on collagen gels resulted in reduced proliferative heterogeneity. Factors affecting cell proliferation of rat uterine epithelial cells in vitro are discussed. Proliferation of rat uterine fibroblastic cells in primary culture was studied using a fluoresence DNA microassay. Data confirming the validity of this assay are presented. The sensitivity of the proliferation of these fibroblastic cells to oestrogen was studied. The data are discussed in relation to current models which suggest an 'indirect' mechanism/action of oestrogen-induced proliferation of target cells. Oestrogen receptors were identified in cultured rat uterine fibroblastic cells, by a competitive binding assay using tritiated-oestradiol-17p and diethylstilboestrol. Conditions were established for a 'one-point' 'exchange' assay of oestrogen receptor levels. Using this assay, oestrogen receptor levels in cultured fibroblastic' cells were monitored in response to different serum treatments. A 2.5-fold difference in the level of specific oestradiol binding in the cytosol between cells cultured in whole and in heat-inactivated charcoal stripped foetal calf serum, indicates the presence of a factor, which depresses cellular binding levels. It is active in foetal calf serum (FCS), but is considerably less active in heat- inactivated charcoal stripped foetal calf serum (HIDCCFCS). This is discussed. Characteristics of rat uterine epithelial and fibroblastic cells were investigated by immunocytochemical detection of intermediate filaments. Epithelial cells were found to express cytokeratins, and fibroblastic cells express vimentin, both in vivo and in culture. However, vimentin staining also appears in epithelial cells in vitro. Desmin staining of fibroblastic cells was inconclusive. Epithelial cells derived from human endometrial carcinomas (HEC cells) were also established as primary cultures. Characteristic changes in cell- surface morphology were seen in response to oestrogen and a progestin in vitro, by using scanning electron microscopy. The data indicate that similar studies may be potentially useful in the dynamic evaluation of the hormone dependence of uterine cancer. Expression of a low molecular weight cytokeratin antigen ('simple epithelial antigen') was found to be altered in cultured HEC cells by retinoic acid. The implications of these observations are discussed.

Item Type: Thesis (PhD)
Qualification Level: Doctoral
Additional Information: Adviser: Robin Leake
Keywords: Developmental biology
Date of Award: 1985
Depositing User: Enlighten Team
Unique ID: glathesis:1985-71513
Copyright: Copyright of this thesis is held by the author.
Date Deposited: 10 May 2019 14:26
Last Modified: 10 May 2019 14:26
URI: http://theses.gla.ac.uk/id/eprint/71513

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