Assessment of Global Methylation in Paraffin Embedded Prostatic Tissues and Cell Lines Using Flow Cytometry

Abstract

Background. The aim of this study is to measure global 5-methylcystosine (5MeC) methylation in paraffin embedded prostatic tissues and cell lines using flow cytometry. Methods. Cell/nuclei suspension from 10 cases of benign prostatic hyperplasia (BPH), 10 cases of prostatic adenocarcinoma, and two prostatic cell lines (PNT1A and LNCaP) were prepared using modified heat pretreatment technique. 5MeC global methylation was assessed by flow cytometry of cell/nuclei suspension and immunostaining of tissue sections. Results. Higher percentage of positively stained cells (PPSC) and mean channel fluorescence (MCF) were detected in PNT1A cell line and BPH cell/nuclei suspensions as compared to LNCaP cell lines and adenocarcinoma cell/nuclei suspensions. Lower scores of 5MeC immunostaining were observed in all prostate adenocarcinoma tissue sections as compared to BPH sections indicating global hypomethylation in prostate adenocarcinoma. Two distinctive populations of cells were detected in histograms generated from most of the BPH cell/nuclei suspensions. Conclusion. The study developed a novel technique that could measure 5MeC global methylation in paraffin embedded prostatic tissues. This represents a rapid and objective assessment of methylation and when combined with tissue micro-dissection and cell sorting, this technique could be applied to larger tissue samples such as post radical prostatectomy and transurethral resected specimens.