Tissue culture media supplemented with 10% fetal calf serum contains a castrate level of testosterone

Abstract

Background Human prostate cancer cells are routinely maintained in media supplemented with 10% Fetal Calf Serum (FCS) to provide androgen. In the present study, total and free testosterone levels in 10%FCS supplemented tissue culture media were determined and compared to levels in intact and castrated human males. Dextran-coated charcoal stripped FCS (i.e., DC-FCS) is often used instead of FCS to minimize the level of androgen provided in 10% serum supplemented media. Therefore, total and free testosterone levels in 10%DC-FCS containing media were likewise determined. Methods Total testosterone, free testosterone, and total dihydrotestosterone (DHT) were determined on RPMI-1640 media supplemented with either 10%FCS or 10%DC-FCS by ELISA assays before and after exposure to LNCaP human prostate cancer cells in culture. The growth and PSA secretion by these cells was also determined. Results 10%FCS supplemented media contains a castrate level of testosterone. However, even with this castrate starting level of testosterone, LNCaP cells concentrate and metabolize the testosterone to produce a physiologic (i.e. 10 nM) level of intracellular DHT which optimally stimulates the growth of these cells in vitro. Conclusions The present studies document that prostate cancer cells auto-regulate their androgen metabolism so that an optimal level of DHT for growth is maintained during both up and down fluctuations in the supply of testosterone. These results have significant implications for whether exogenous androgen should be added to the 10%FCS supplemented media to grow prostate cancer cells from intact vs. castrated patients.

14-3-3ζ, a Novel Androgen-Responsive Gene, Is Upregulated in Prostate Cancer and Promotes Prostate Cancer Cell Proliferation and Survival

“…The cells were maintained in RPMI-1640 medium supplemented with 10% FBS. According to a previous report (25), the total testosterone is at the castrate level under this condition. LNCaP cells are considered to metabolize testosterone to produce a physiological (i.e., 10 nmol/L) level of DHT (25).…”

“…According to a previous report (25), the total testosterone is at the castrate level under this condition. LNCaP cells are considered to metabolize testosterone to produce a physiological (i.e., 10 nmol/L) level of DHT (25). The proliferation rate of LNCaP-FLAG-14-3-3z stable clones was significantly increased after 72-and 96-hour incubations, compared with that of the vector clones (P < 0.05; Fig.…”

Androgen activates β-catenin signaling in bladder cancer cells

“…There are several possibilities underlying this observation. First, remaining androgens in charcoal-stripped FBS (Sedelaar & Isaacs 2009) used for our cell culture led to AR activation since as low as 0.1 nM dihydrotestosterone was shown to regulate the growth of prostate cancer cell lines with endogenous or overexpressed AR (Mizokami et al 2004, Waltering et al 2009). Secondly, AR could be activated by non-androgenic compounds, such as growth factors (Culig et al 1994).…”

Development of &lt;i&gt;in vitro&lt;/i&gt; culture of rat Leydig cells after purification with Nycodenz gradient

“…Sedelaar and Isaacs [24] reported that hormone testosterone concentrations between 1.2 and 7.5 ng/ml were found in bovine serum from newborn to one year old assayed using the RIA method. Serum contains several nutritional components such as fatty acids, cholesterol, T3, insulin, IGF, EGF and androgen [25]. Moreover, the addition of serum is made to provide the hormones that stimulate cell growth and function.…”