Monoclonal antibody characterization of progesterone receptors, estrogen receptors and the stress-responsive protein of 27 kDa (SRP27) in human uterine leiomyoma

Abstract

Uterine leiomyoma occurs in one of every four or five women during their reproductive life. Its origin is unknown but it is accepted that estrogens play a significant role in its development. In order to learn more about the estrogen dependency of leiomymoma, the biochemical and immunological properties of two markers of estrogen response in target cells (the progesterone receptor (PR) and the stress-responsive protein of 27 kDa (SRP27)) were studied in leiomyoma. The ER (estrogen receptor) and PR content were determined by conventional DCC exchange assays. Specific anti-ER, anti-PR and anti-SRP27 monoclonal antibodies were used in immunoblots and immunohistochemical (IHC) studies. The binding properties of PR from cytosol of leiomyoma showed a Kd of 0.8-1.3 nM, which is in the range described for other human tissues. 80% of all studied leiomyoma contained PR, in a range of 805-2000 fmol/mg protein. The Kd for leiomyoma ER was 0.1-0.9 nM, and 84% of the samples were positive for ER. The PR of leiomyoma has the two A and B forms of 120 and 94 kDa, as shown in the immunoblot using the AB52 anti-PR monoclonal antibody. The IHC study revealed that the PR is concentrated in the cell nuclei, in the form of perinuclear bodies, with a homogeneous staining pattern from cell to cell. The leiomyoma fibres contain SRP27 in a higher concentration than the healthy myometrium. The leiomyoma SRP27 shows a typical doublet of 24 kDa and 27 kDa in immunoblot, the same as in MCF-7 cells. The IHC study revealed a high degree of organization of SRP27 in leiomyoma cells, suggesting that this protein may be part of the cytoskeleton. The results obtained show that human leiomyomas contain ER, PR and RSP27 with similar immunological and biochemical properties to those of other human tissues, including the MCF-7 breast cancer cell line.