Hormone-Supplemented Medium Enhances Androgen Binding Protein Secretion in Sertoli Cell Cultures

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Abstract
Androgen binding protein (ABP) is synthesized and secreted into the seminiferous tubules by Sertoli cells. In cell culture, Sertoli cells will initially continue to synthesize ABP and to respond to stimulation by FSH [follitropin]. It was the objective of this study to determine cell culture conditions such that the production of ABP by Sertoli cells remained at a high level for a long period of time. Sertoli cell cultures from 20 day old Sprague-Dawley rats were initially plated in F-12 medium which contained 1% calf serum. The cells were incubated for 48 h at 32.degree. C and the medium was changed to F-12 lacking serum. Hormones or growth factors were added at this time. Thereafter, every 4 days the medium was replaced with fresh F-12 and hormones or growth factors were readded. Concentration of hormones in Serum Substitution Medium (SS-F-12): ovine FSH, 0.5 .mu.g/ml; testosterone, 2 .times. 10-9 M; insulin, 5 .mu.g/ml (0.1 IU/ml); epidermal growth factor, 1 .mu.g/ml; somatomedins, 1 .mu.g/ml; transferrin, 5 .mu.g/ml. The medium which contained a 4 day accumulation of ABP was assayed by equilibrium dialysis. The binding data is expressed as cpm 3H-testosterone bound per 100 .mu.l of culture fluid. The testosterone had a specific activity of 40 Ci/mmole and was present in the dialysis buffer at 2 nM. If no hormones were added, ABP secretion was greatly diminished and continued to decline. In the cultures which contained FSH and testosterone, the level of ABP activity was initially high but by the 3rd assay period had declined to less than 60% of the level of the first assay period. If the cells were maintained in SS-F-12, the ABP activity per 4 day period was sustained at a high level and actually tended to increase throughout the duration of the experiment. Additives, such as insulin, epidermal growth factor, transferrin and sommatomedins, extend the ability of the Sertoli cell cultures to secrete ABP for at least 2 wk. Experiments are currently underway to determine which of the hormone additives are most effective in promoting ABP synthesis.