Androgen receptor in serially subcultured human endometrial fibroblasts

Specific dihydrotestosterone (DHT) binding was demonstrated in the whole cell sonicates obtained from serially subcultured human endometrial fibroblasts. This binding showed a high affinity and specificity for DHT (Kd=3.3 X 10(-10) M). Various kinds of excess nonradioactive steroids were arranged, as a result of competition among them, in the following decreasing order of binding affinity: DHT, testosterone, cyproterone acetate, 17 beta-estradiol, progesterone, and 5 alpha-androstane-3 alpha, 17 beta-diol. Androsterone, 5 alpha-androstanedione, cortisol, androstenedione and estrone hardly affected the binding even at 200 fold excess. Saturation analysis indicated that cultured endometrial and genital skin fibroblasts have a similar number of binding sites for DHT (14.3 +/- 8.5 vs 11.5 +/- 8.6 fmoles/mg cell protein). Sucrose density gradient centrifugation brought about a sharp peak of 3H-DHT in the 4S region in high ionic strength buffer. This binding was abolished by heating sonicates at 37 degrees C for 1 hr, treating them with pronase or adding to them excess nonradioactive DHT. Endometrial fibroblasts also contained testosterone binding components which formed a peak in the 4S region and showed a similar binding affinity to that for DHT.