Serafini P, Ablan F, Lobo R A
J Clin Endocrinol Metab. 1985 Feb;60(2):349-55. doi: 10.1210/jcem-60-2-349.
A simplified, rapid, and highly reproducible technique is described for measuring 5 alpha-reductase activity (5 alpha RA) in small skin biopsies. Human genital skin was obtained from 23 nonhirsute and 20 hirsute premenopausal women (HW) and 5 normal men. Skin samples were minced at 4 C and incubated with RPMI-1640 in the presence of 95% O2-5% CO2 and 4.15 nmol [14C]testosterone ([14C]T) for 2 h at 37 C. Steroids were extracted with diethyl ether and separated by Celite and paper chromatography. Radioactivity in specific eluates was quantified, and the mass of each steroid was measured by RIA. The separate formation of 5 alpha-androstane-17 beta-ol-3-one (DHT), 5 alpha-androstane-3 alpha, 17B diol (3 alpha diol), androstenedione, and androsterone from [14C]T was measured. In separate experiments it was demonstrated that an incubation time of 2 h was optimum and that the addition of cofactors was unnecessary. Radiochemical purity was confirmed after chromatography. The mean +/- SE conversion ratio (CR) of T to DHT (in 2 h) in HW was higher than that in normal women (16.80 +/- 1.62% vs. 4.48 +/- 0.36%; P less than 0.01). In men, the CR of T to DHT averaged 31.60 +/- 3.96%. Individual values for the CR of T to DHT in HW and normal women did not overlap. The CR of T to 3 alpha diol was significantly higher in HW (9.66 +/- 0.86%) and men (15.98 +/- 2.0%) compared to that in normal women (2.96 +/- 0.32%; P less than 0.05). The CR of T to androstenedione was significantly greater in HW and men (6.18 +/- 0.42 and 7.28 +/- 1.92%) compared to that in normal women (2.64 +/- 0.64%; P less than 0.05). The CR of T to androsterone was very low and was similar in the three groups. The production of DHT in HW (4.50 +/- 1.0 pmol/mg X 2 h) was significantly greater than that in normal women (0.48 +/- 0.08; P less than 0.01) and was similar to the production in men (6.18 +/- 1.94 pmol/mg X 2 h). There was a significant correlation between the CR of T to DHT and DHT production, and the CR of T to 3 alpha diol and 3 alpha diol production as well as between the CRs of T to DHT and T to 3 alpha diol. These data suggest that measurements of DHT formation are best suited for the assessment of 5 alpha RA and that the measurement of 5 alpha RA in vitro from small skin biopsies is suitable for the clinical evaluation of hirsutism.
本文描述了一种用于测量小块皮肤活检组织中5α-还原酶活性（5αRA）的简化、快速且高度可重复的技术。从23名非多毛和20名多毛的绝经前女性（HW）以及5名正常男性身上获取人类生殖器皮肤。将皮肤样本在4℃下切碎，在95% O₂-5% CO₂和4.15 nmol [¹⁴C]睾酮（[¹⁴C]T）存在的情况下，于37℃用RPMI-1640孵育2小时。用乙醚提取类固醇，并通过硅藻土和纸色谱法进行分离。对特定洗脱液中的放射性进行定量，并通过放射免疫分析（RIA）测量每种类固醇的质量。测量了[¹⁴C]T分别生成5α-雄烷-17β-醇-3-酮（双氢睾酮，DHT）、5α-雄烷-3α,17β-二醇（3α-二醇）、雄烯二酮和雄酮的情况。在单独的实验中表明，2小时的孵育时间是最佳的，并且无需添加辅因子。色谱分析后确认了放射化学纯度。HW中T转化为DHT的平均±标准误转化率（CR）（2小时内）高于正常女性（16.80±1.62%对4.48±0.36%；P<0.01）。在男性中，T转化为DHT的CR平均为31.60±3.96%。HW和正常女性中T转化为DHT的个体值没有重叠。与正常女性相比，HW（9.66±0.86%）和男性（15.98±2.0%）中T转化为3α-二醇的CR显著更高（正常女性为2.96±0.32%；P<0.05）。与正常女性相比，HW和男性中T转化为雄烯二酮的CR显著更高（分别为6.18±0.42和7.28±1.92%）（正常女性为2.64±0.64%；P<0.05）。T转化为雄酮的CR非常低，且在三组中相似。HW中DHT的生成量（4.50±1.0 pmol/mg×2小时）显著高于正常女性（0.48±0.08；P<0.01），且与男性中的生成量相似（6.18±1.94 pmol/mg×2小时）。T转化为DHT的CR与DHT生成量之间、T转化为3α-二醇的CR与3α-二醇生成量之间以及T转化为DHT的CR与T转化为3α-二醇的CR之间均存在显著相关性。这些数据表明，测量DHT的生成最适合用于评估5αRA，并且从小块皮肤活检组织中体外测量5αRA适用于多毛症的临床评估。
