Oestrogen and progesterone receptors in human term placenta. Measurement by binding assays and immunological methods.

Abstract

Oestrogen and progesterone receptors have been implicated in human placental steroidogenesis, but studies developed to measure their concentration in this tissue at term yielded discrepant results. Potential inaccuracy of the binding assays, the technique used in previous studies, might influence the discrepancy. In the present study, oestrogen and progesterone receptors were investigated by two different approaches: binding assays with radiolabelled ligand and immunoassays with monoclonal antibodies against receptors. Binding assays included techniques to quantify free and occupied sites in both the cytosolic and nuclear fraction. Conditions intended to block or decrease potential causes of inaccuracy, such as proteolysis or thermal denaturation of the sites, were included in the assays. Cytosolic specific binding was measured by two different techniques, dextran-coated charcoal and hydroxylapatite, whereas nuclear sites were studied on salt-extracts and by exchange. Negative or moderately positive results were obtained for either oestrogen or progesterone receptors, but high levels of non-specific binding made the technique unreliable. Immunoassays included the use of two techniques, histochemistry and enzyme-linked-immunosorbent-assay (ELISA). Both frozen and paraffin (Bouin solution or buffered formalin as fixatives) sections were stained with monoclonal antibodies against the oestrogen receptor. Results were always negative in all cases. Levels of oestrogen and progesterone receptors were measured in either cytosol or nuclear extracts by ELISA. This technique was highly sensitive and reproducible. Results were negative for both the cytosolic and nuclear fractions of oestrogen receptors. For progesterone receptors, low positivity was obtained in either crude (3.2 +/- 0.4 fmol/mg protein, mean +/- s.e.) or ammonium sulphate precipitated cytosols (12.6 +/- 3.5 fmol/mg protein, mean +/- s.e.), whereas the values for nuclear extracts were 62.0 +/- 9.0 fmol/mg DNA (mean +/- s.e.). We conclude that: (1) oestrogen receptors could not be detected in human term placenta by the methods used in this study, and (2) low levels of progesterone receptors were detected in both cytosolic and nuclear extracts.