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Title: The molecular basis of steroid hormone promoted growth
Author: Love, Colin Andrew
ISNI:       0000 0001 3613 0366
Awarding Body: University of Glasgow
Current Institution: University of Glasgow
Date of Award: 1982
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1. The literature concerning measurement and stability of the oestrogen nuclear receptor has been reviewed in relation to the use of receptor assays for management of breast cancer. 2. The influence of temperature and ionic strength on the stability of the human nuclear oestrogen receptor complex was studied in relation to the major methods used to assay for nuclear receptor. The exchange rate of (3H)-oestradiol from the nuclear receptor was measured as a decrease in specifically bound labelled oestradiol during an incubation with an excess of unlabelled oestradiol. 3. When the nuclear oestrogen receptor complex was bound to intact nuclei, the rate of exchange increased with temperature but a significant exchange could be measured over 24 hours at 4°c. The use of protease inhibitors together with re-labelling of nuclear receptor after incubation confirmed that the observations represented exchange of hormone rather than degradation of the hormone receptor complex. 4. Measurement of the on-rate of ( 3H) - oestradiol to the oestrogen nuclear receptor, prefilled with unlabelled oestradiol, also demonstrated that there was a significant rate of exchange of hormone at 4^c. 5. The exchange rate from salt extracted, hydroxylappatite precipitated receptor also increased with temperature. However, this extracted complex was more stable and no exchange could be detected at 4°c. 6. Only 50% of total detectable human nuclear oestrogen receptor was solubilised by the standard method of salt extraction in 0.6m KCl. As the salt concentration was raised (0-0.6M KCl) an increase in stripping of oestradiol from the hormone receptor complex was observed. This would create a false impression of the proportion of "empty nuclear receptor". 7. These results were compared with other reports of the stability of the hormone receptor complex. The consequences of these results were discussed in relation to current nuclear oestrogen receptor assays and their use in stratifying therapy for patients with breast cancer. 8. The literature concerning current iu vitro models of human breast cancer has been reviewed and the merits of these models have been compared. 9. A method has been developed for the routine primary culture of cells from breast cancer biopsies. This method involved digestion with collagenase followed by growth on a feeder layer. Approximately 85% of biopsies gave rise to viable primary cultures. 10. Growth of these cells was in the form of ring colonies which could be maintained in culture for up to 3 months. Cells were characterised as epithelial by electron microscopy and immunofluorescence and tests were carried out to demonstrate the retention of malignant potential. 11. The effect of hormones and growth factors on the growth of cells in the ring colonies has been investigated. In particular cortisol and oestradiol were found to act synergistically to stimulate growth. Cholera toxin, prolactin and cortisol were found to stimulate growth or plating efficiency to a lesser extent. Epidermal growth factor did not stimulate growth of breast epithelial cells. The effect of these hormones was compared with published reports of other in vitro systems. 12. Differences between primary tumours (such as oestrogen receptor status or growth rate) were reflected by the growth and the response to hormonal stimulation of the ring colonies, 13. A method was developed, using autoradiography, to measure the effect of hormones and growth factors on (3H) thymidine incorporation in a small number of cultured breast epithelial cells. The possible applications of this system have been discussed.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available
Keywords: Biochemistry