Novel method for the isolation and characterisation of the putative prostatic stem cell.
BACKGROUND
Prostate stem cells, responsible for the development, maturation, and function of the prostate, have been implicated in the aetiology of both benign prostate hyperplasia (BPH) and prostate cancer (CaP). However, research has been hampered by the lack of a definitive stem cell marker. We have adapted the protocol for differential Hoechst 33342 uptake by hemopoietic stem cells to enable isolation of putative stem cells from the prostate.


METHODS
Prostate epithelial cells isolated from prostate tissue obtained from patients with BPH after transurethral resection of the prostate were stained with Hoechst 33342. The Hoechst 33342 Red/Blue flow cytometry profile was then determined. Hoechst 33342 and Pyronin Y staining was used to determined the cell cycle status.


RESULTS
A verapamil-sensitive side population (SP) can be isolated from primary prostate tissue accounting for 1.38% +/- 0.07% of prostate epithelial cells. Cell cycle analysis of this SP population revealed that the majority of SP cells are in either G0 (12.38 +/- 0.31%) or G1 (63.19 +/- 2.13%).


CONCLUSIONS
The Hoechst 33342 dye efflux protocol can be adapted for the isolation of a SP from primary prostate tissue.
Bhatt
Rupesh I
RI
Genito-Urinary Cancer Research Group, Cancer Research UK Paterson Institute for Cancer Research, Christie Hospital NHS Trust, Manchester, UK.
Brown
Michael D
MD
Hart
Claire A
CA
Gilmore
Paul
P
Ramani
Vijay A C
VA
George
Nicholas J
NJ
Clarke
Noel W
NW
D001562:Benzimidazoles / Q000493:pharmacokinetics; D002121:Calcium Channel Blockers / Q000494:pharmacology; D018929:Cell Culture Techniques / Q000379:methods*; D002453:Cell Cycle; D004847:Epithelial Cells / Q000166:cytology; D005434:Flow Cytometry / Q000379:methods*; D005456:Fluorescent Dyes / Q000494:pharmacology; D016193:G1 Phase; D006412:Hematopoietic Stem Cells / Q000166:cytology; D006801:Humans; D007633:Keratins / Q000378:metabolism; D017493:Leukocyte Common Antigens / Q000096:biosynthesis; D008297:Male; D010641:Phenotype; D011419:Propidium / Q000494:pharmacology; D011467:Prostate / Q000166:cytology*; D016192:Resting Phase, Cell Cycle; D013234:Stem Cells / Q000166:cytology*; D014700:Verapamil / Q000494:pharmacology
D001562:Benzimidazoles; D002121:Calcium Channel Blockers; D005456:Fluorescent Dyes; D011419:Propidium; D007633:Keratins; D014700:Verapamil; D017493:Leukocyte Common Antigens; C017807:bisbenzimide ethoxide trihydrochloride