Androgen binding site in J111 cell line.
BACKGROUND
The finding of sex steroid receptor protein in non classical reproductive tissues suggested the possibility that sex steroids may have a relevance to the immune system.


METHODS
The J111 cells were maintained in RPMI 1640 complete medium at 37 degrees C in 5% CO2 in air. Cells were resuspended at 1x10(6) cells in 0.2 ml complete medium in 1.5 ml eppendorf tubes. A single saturating concentration 1x10(-9) M of [3H]5alpha-DHT was added to the cells suspension. Unlabelled steroids (5alpha-DHT, 17-beta estradiol, or the synthetic glucocorticoid triamcinolone acetonid) were added over the range 1x10(-8) to 1x10(-9) M. Duplicate tubes were incubated at 37 degrees C for 1h. For autoradiography, the supernatant was discarded and the pellet resuspended in 0.2 ml medium. For binding assay, Labeled cells were separated from unbound steroid by immunomagnetic bead using anti-CD68 antibody.


RESULTS
In autoradiography, a population of approximately 96% of J111 cells that contain receptors for androgen has been demonstrated. The results of immunomagnetic showed that binding identified in the J111 cells was modest selective towards androgenic compounds. Schatchard analysis of data showed the KD value of 2.5x10(-9) M and the number of receptor in each cell was found to be 257+/-1. Little competition was seen from 17 beta estradiol or the synthetic glucocorticoid triamcinolone acetonid.


CONCLUSIONS
These data indicate that androgen binding in J111 cells is of modest affinity and specific, due to the inability of 100-fold molar excess of estradiol to displace bound [3H]-5alphaDHT.
Ahmadi
Kazem
K
Department of Immunology and Research Centre of Molecular Biology, Baqiyatallah University of Medical Sciences, Tehran, I.R. Iran. K.Ahmadi@bmsu.ac.ir
McCruden
Alan B
AB
D013741:3-Oxo-5-alpha-Steroid 4-Dehydrogenase / Q000378:metabolism; D000728:Androgens / Q000378:metabolism*; D001345:Autoradiography; D001665:Binding Sites; D004958:Estradiol / Q000378:metabolism; D006801:Humans; D014222:Triamcinolone Acetonide / Q000378:metabolism
D000728:Androgens; D004958:Estradiol; D013741:3-Oxo-5-alpha-Steroid 4-Dehydrogenase; D014222:Triamcinolone Acetonide