Bruchovsky N, Sadar M D, Akakura K, Goldenberg S L, Matsuoka K, Rennie P S
Department of Cancer Endocrinology, British Columbia Cancer Agency, Vancouver, Canada.
J Steroid Biochem Mol Biol. 1996 Dec;59(5-6):397-404. doi: 10.1016/s0960-0760(96)00125-2.
The expression of 5alpha-reductase type 1 and type 2 isoenzymes in hyperplastic human prostate tissue and several human prostate cell lines was investigated by Northern blot analyses, reverse transcription-polymerase chain reaction (RT-PCR), and enzyme activity. Separation of stroma and epithelium was confirmed histologically and only preparations with no apparent contamination were employed in the subsequent studies. Poly(A)+ RNA was isolated from stromal and epithelial fractions and analysed by Northern blot and RT-PCR. Inhibition of epithelial and stromal 5alpha-reductase activities by 17beta-N,N-diethylcarbamoyl-4-methyl-4-aza-5alpha-androstan- 3-one (4MA) was assessed using a range of concentrations between 10(-13) and 10(-5) M. Results from Northern blot analyses and RT-PCR showed that the prostate stroma expressed 5alpha-reductase type 1 and type 2 isoenzymes, whereas the prostate epithelium only expressed 5alpha-reductase type 1. This was consistent with biphasic inhibition of 5alpha-reductase activity by 4MA in stroma and monophasic inhibition in epithelium. Cultured epithelial cells derived from human prostate only expressed 5alpha-reductase type 1 and had Vmax and Km values that approximated the lower end of the range reported for surgically removed prostate epithelium. The foregoing data explains the disparate activities of 5alpha-reductase, previously reported, in stroma and epithelium. The differential localization of these isoenzymes in the prostate suggests that future therapy of androgen-sensitive disease may be more successful through the use of selective inhibitors of the different 5alpha-reductase isoenzymes.
通过Northern印迹分析、逆转录-聚合酶链反应（RT-PCR）和酶活性检测，研究了5α-还原酶1型和2型同工酶在人前列腺增生组织及几种人前列腺细胞系中的表达情况。通过组织学方法确认了基质和上皮的分离，后续研究仅采用无明显污染的样本。从基质和上皮部分分离出poly(A)+ RNA，并通过Northern印迹和RT-PCR进行分析。使用浓度范围在10^(-13)至10^(-5) M之间的17β-N,N-二乙基氨基甲酰基-4-甲基-4-氮杂-5α-雄甾烷-3-酮（4MA）评估其对上皮和基质5α-还原酶活性的抑制作用。Northern印迹分析和RT-PCR结果显示，前列腺基质表达5α-还原酶1型和2型同工酶，而前列腺上皮仅表达5α-还原酶1型。这与4MA对基质中5α-还原酶活性的双相抑制以及对上皮中5α-还原酶活性的单相抑制相一致。来源于人前列腺的培养上皮细胞仅表达5α-还原酶1型，其Vmax和Km值接近手术切除的前列腺上皮报道范围的下限。上述数据解释了先前报道的5α-还原酶在基质和上皮中不同的活性。这些同工酶在前列腺中的差异定位表明，未来通过使用不同5α-还原酶同工酶的选择性抑制剂治疗雄激素敏感性疾病可能会更成功。
