Sakamoto Shinji, Yokoyama Masahiro, Prakash Kulkarni, Tsuruha Jun-Ichiro, Masamoto Satoshi, Getzenberg Robert H, Kakehi Yoshiyuki
Pharmaceutical Frontier Research Laboratories, JT, Inc., Yokohama City, Japan.
J Biomol Screen. 2003 Dec;8(6):701-11. doi: 10.1177/1087057103259159.
Among urological diseases, benign prostatic hyperplasia (BPH) exhibits a high morbidity rate, afflicting approximately 50% of men older than age 50 years. Despite intense research efforts over the past decades, the etiology and mechanisms of BPH progression are only poorly understood. Employing oligonucleotide microarrays, the authors analyzed the gene expression profiles in normal and BPH prostate samples and found that CYR61, an immediate early gene, is markedly overexpressed in BPH. To quantify cellular CYR61 mRNA expression directly, the authors developed an assay using branched-chain DNA (bDNA) technology. A human prostatic epithelial cell line, BRF-55T, derived from a BPH patient, was treated with fetal bovine serum to stimulate gene expression, and then the induction profile of the CYR61 mRNA in these serum-stimulated cells was quantitated using both bDNA and quantitative reverse transcriptase-PCR (RT-PCR). The results obtained with the 2 detection systems were found to be very similar. The bDNA assay was also found to be sensitive and highly reproducible. To the authors' knowledge, this is the first time that identifying CYR61 as a novel marker for BPH and its quantitation has been reported. These detection methods not only may be useful for diagnostic purposes but may also be used to identify suppressors of CYR61 expression for BPH therapy employing high-throughput screening assays.
在泌尿系统疾病中,良性前列腺增生(BPH)发病率很高,约50%的50岁以上男性受其困扰。尽管在过去几十年中进行了大量研究,但BPH进展的病因和机制仍知之甚少。作者利用寡核苷酸微阵列分析了正常前列腺样本和BPH前列腺样本中的基因表达谱,发现即刻早期基因CYR61在BPH中明显过表达。为了直接定量细胞CYR61 mRNA表达,作者开发了一种使用分支DNA(bDNA)技术的检测方法。用胎牛血清处理源自一名BPH患者的人前列腺上皮细胞系BRF-55T以刺激基因表达,然后使用bDNA和定量逆转录聚合酶链反应(RT-PCR)对这些血清刺激细胞中CYR61 mRNA的诱导情况进行定量。发现两种检测系统获得的结果非常相似。还发现bDNA检测方法灵敏且重复性高。据作者所知,这是首次报道将CYR61鉴定为BPH的新型标志物并对其进行定量。这些检测方法不仅可能有助于诊断,还可用于通过高通量筛选试验鉴定用于BPH治疗的CYR61表达抑制剂。