Radioimmunoassay measurements of nuclear dihydrotestosterone in rat prostate. Relationship to androgen receptors and androgen-regulated responses

Abstract

The concentration of dihydrotestosterone was measured by radioimmunoassay in nuclear and cytoplasmic extracts from rat ventral prostates. In the regenerating prostates of castrated rats treated with dihydrotestosterone for 4 days, the nuclear concentration of this steroid increased from approx. 70nM to 800nM as a linear function of the injected dose, whereas the cytoplasmic concentration remained relatively constant (70-130nM). Isotope-exchange measurements of nuclear androgen receptors by using [3H]methyltrienolone indicated that, although the concentration of nuclear dihydrotestosterone was several-fold higher than the concentration of androgen receptors, they were logarithmically related. The recruitment of prostatic cells into the growth fraction and the stimulation of 5 a-reductase activity were more directly correlated to the nuclear concentration of androgen receptors than to the total nuclear concentration of dihydrotestosterone. Maximal restoration of a specific isoenzyme of acid phosphatase was achieved when approx. 2000 androgen receptors were present in the prostatic nuclei; higher concentrations of nuclear androgen receptors were associated with decreased amounts of this enzyme. Hence the results imply, first, that the total amount of dihydrotestosterone accumulated by nuclei is not a direct consequence of carrier-mediated transport by androgen receptors, and, secondly, that, whereas acid phosphatase may be differentially controlled by androgens in the regenerating prostate, increases in the amount of cell proliferation and 5a-reductase activity directly parallel increases in the nuclear concentration of androgen receptors.In the prostate, androgenic regulation of physiological functions and cellular growth is attributed largely to the formation, and subsequent binding to chromatin, of androgen-receptor complexes (King & Mainwaring, 1974 (Coffey, 1974;Rennie et al., 1975;Van Doorn et al., 1976). Although it has been postulated that androgen receptors are involved in the uptake and retention of dihydrotestosterone by target-cell * To whom correspondence should be addressed.Vol. 200 nuclei (Rennie & Bruchovsky, 1973;Bruchovsky et al., 1975a), a direct stoicheiometric relationship has not been demonstrated. When castrated rats are injected with radioactive androgen in the ventral prostate the concentration of radioisotope measured in nuclei exceeds that measured in the cytoplasm (Bruchovsky et al., 1975b). However, the concentration of androgen-receptor complexes recovered from the nuclei is insufficient to account for the nuclear influx of androgens if a mole-to-mole relationship is assumed. Furthermore, radioimmunoassay measurements of dihydrotestosterone in ventral prostates from non-castrated rats have indicated that only about 20% of the endogenous nuclear androgen is bound to androgen receptors (Bruchovsky et al., 1980). Whether the large nuclear concentration of non-receptor-bound dihydrotestosterone is achieved through a receptor mechanism or

Androgen metabolism and regulation of rat ventral prostate growth and acid phosphatase during sexual maturation

“…These results correlated well with other studies (Rennie et al 1978; De Larminat, Rennie & Bruchovsky, 1981;Downey et al 1983) using cas¬ trated adult rats injected with low or high doses of androgen and measuring the rate of cell proliferation and production of secretory AP. Low doses of andro¬ gen induced secretory AP without causing prostate enlargement.…”

Molecular Biology of Androgen Action in Genital Hypoplasia Associated with Congenital Growth Hormone Deficiency: A “Transitory Androgen Insensitivity Syndrome”?

“…The DHT production in our patient points toward an involvement of peripheral A5R [3] responsible for the major part of the DHT concentration measured. A5R activity depends on adequate substrate (T) concentration but is also regulated by the DHT receptor [5]. DHT binds to the cytosolic and nuclear fraction in a demonstrable and saturable manner.…”

Synthesis and Evaluation of Immobilized Androgens for Affinity Chromatography in the Purification of Nuclear Androgen Receptor

“…Nuclear extracts were prepared and concentrated by the following sequence: sonication of purified nuclei, digestion of the sonicated material with micrococcal nuclease, extraction of the digests with 0.6 M ammonium bicarbonate, and lyophilization of the extracts as described in detail elsewhere [4,5,9]. The rate of inactivation of lyophilized nuclear androgen receptor was approximately 6% per week.…”

“…(6) and its l7p-acetate (7) were synthesized by the same procedure using instead 4,6-androstadienone-19-nor, 17p-ol,3-oneacetate (8) as starting material prepared according to published procedures 161. Alternatively, reduction of alcohol (3) with lithium in liquid ammonia [8] yielded hydroxy-ketone (9) which was oxidized to give dihydrotestosterone-17P-acetoxy-7aundecanoic acid ( 10). Deacetylation of adduct (9) afforded dihydrotestosterone-7aundecanoic acid (11).…”

“…Alternatively, reduction of alcohol (3) with lithium in liquid ammonia [8] yielded hydroxy-ketone (9) which was oxidized to give dihydrotestosterone-17P-acetoxy-7aundecanoic acid ( 10). Deacetylation of adduct (9) afforded dihydrotestosterone-7aundecanoic acid (11).…”