Gene expression profiles of human BPH (II): Optimization of laser-capture microdissection and utilization of cDNA microarray.
Laser capture microdissection (LCM) enables the dissection of heterogeneous components of tissues, helping to investigate the molecular properties of these tissues. We have reported gene expression profiles in human benign prostatic hyperplasia (BPH) using LCM and quantitative real-time PCR. In the current work, we extended the previous observations. Firstly, we studied the relationship between the number of dissected acini and amplification by PCR, and found that androgen receptor (AR) and 18S rRNA transcripts were successfully amplified from total RNA obtained from one prostate acinus. Furthermore, LCM-dissected samples were applicable to methylation-specific PCR of E-cadherin promoter gene after bisulfite modification of genomic DNA. Next, we performed cDNA microarray analysis to screen gene expression profiles in the epithelium and stroma. RNA was amplified by T7-RNA polymerase and labeled with Cy3 and Cy5. Epithelium-related or stroma-related genes were identified through cDNA microarray. We confirmed true gene expression levels by quantitative real-time PCR. In epithelium, E-cadherin and serine protease 2, Kunitz-type gene expression levels were significantly elevated, while the connective tissue growth factor gene expression level was significantly elevated in stroma. Thus, LCM-dissected samples were applicable to various molecular examinations including methylation-specific PCR and cDNA microarray, and this will contribute to a precise understanding of the molecular profiles of prostate glands.
Matsui
Hiroshi
H
Department of Urology, Gunma University School of Medicine, 3-39-22 Showa-machi, Maebashi-city, Gunma 371-8511, Japan.
Suzuki
Kazuhiro
K
Hasumi
Masaru
M
Koike
Hidekazu
H
Okugi
Hironobu
H
Nakazato
Haruki
H
Yamanaka
Hidetoshi
H
D004247:DNA / Q000032:analysis; D004210:Dissection / Q000379:methods; D020869:Gene Expression Profiling; D006801:Humans; D007834:Lasers; D008297:Male; D008846:Micromanipulation / Q000379:methods*; D020411:Oligonucleotide Array Sequence Analysis / Q000379:methods*; D011470:Prostatic Hyperplasia / Q000235:genetics* / Q000378:metabolism; D012313:RNA / Q000032:analysis
D012313:RNA; D004247:DNA