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dc.contributor.authorBrown, Michael D
dc.contributor.authorGilmore, Paul E
dc.contributor.authorHart, Claire A
dc.contributor.authorSamuel, Joanne D
dc.contributor.authorRamani, Vijay A C
dc.contributor.authorGeorge, Nicholas J
dc.contributor.authorClarke, Noel W
dc.date.accessioned2009-07-07T16:18:44Z
dc.date.available2009-07-07T16:18:44Z
dc.date.issued2007-09-15
dc.identifier.citationCharacterization of benign and malignant prostate epithelial Hoechst 33342 side populations. 2007, 67 (13):1384-96 Prostateen
dc.identifier.issn0270-4137
dc.identifier.pmid17639507
dc.identifier.doi10.1002/pros.20620
dc.identifier.urihttp://hdl.handle.net/10541/72878
dc.description.abstractBACKGROUND: The prostate epithelial stem cell has been proposed as the primary origin of neoplastic change in prostate cancer. However, the isolation and characterization of unexpanded prostate epithelial stem cells have proven problematic. METHODS: A prostate epithelial side population (SP) has been isolated utilizing a modified Hoechst 33342 dye efflux assay from both benign and malignant prostate tissue. CD45(-ve), integrin alpha2(+ve) Hoechst 33342 SP and NSP cells were isolated by FACS, immunophenotyped and functionally characterized in 3D culture. RESULTS: FACS analysis revealed a verapamil sensitive SP accounting for 0.93 +/- 0.12% and 0.57 +/- 0.11% of the total epithelial population from both benign and malignant prostates. The benign SP phenotype revealed a heterogeneous cell population consisting predominantly of small basal cells containing minimal cytoplasm. Conversely, the malignant SP was of undetermined acinar origin and with a complete loss of expression of the CDK2 inhibitor p21(WAF1/Cip1). In vitro androgen-enhanced 3D culture of the benign and malignant SP cells led to the production of spheroids which had acinus like morphology and expressed primitive and basal cell markers. Incorporation of the CD133 marker isolated a further SP sub-fraction accounting for 0.037 +/- 0.01% of epithelial cells. CONCLUSIONS: Our observations are consistent with the Hoechst 33342 dye efflux assay isolating a stem cell enriched population which can be further sub-fractionated by CD133 selection. Moreover, the loss of the CDK inhibitor in malignancy is consistent with the hypothesis that neoplastic change originates in the stem cell compartment.
dc.language.isoenen
dc.subjectProstatic Canceren
dc.subject.meshAdult Stem Cells
dc.subject.meshAntigens, CD
dc.subject.meshAntigens, CD45
dc.subject.meshBenzimidazoles
dc.subject.meshCell Fractionation
dc.subject.meshCell Growth Processes
dc.subject.meshCell Line
dc.subject.meshEpithelial Cells
dc.subject.meshFlow Cytometry
dc.subject.meshFluorescent Dyes
dc.subject.meshGlycoproteins
dc.subject.meshHumans
dc.subject.meshImmunohistochemistry
dc.subject.meshImmunophenotyping
dc.subject.meshMale
dc.subject.meshMicroscopy, Confocal
dc.subject.meshPeptides
dc.subject.meshProstatic Hyperplasia
dc.subject.meshProstatic Neoplasms
dc.titleCharacterization of benign and malignant prostate epithelial Hoechst 33342 side populations.en
dc.typeArticleen
dc.contributor.departmentProMPT Genito-Urinary Cancer Research Group, Paterson Institute for Cancer Research, University of Manchester, Manchester, UK. mbrown@picr.man.ac.uken
dc.identifier.journalThe Prostateen
html.description.abstractBACKGROUND: The prostate epithelial stem cell has been proposed as the primary origin of neoplastic change in prostate cancer. However, the isolation and characterization of unexpanded prostate epithelial stem cells have proven problematic. METHODS: A prostate epithelial side population (SP) has been isolated utilizing a modified Hoechst 33342 dye efflux assay from both benign and malignant prostate tissue. CD45(-ve), integrin alpha2(+ve) Hoechst 33342 SP and NSP cells were isolated by FACS, immunophenotyped and functionally characterized in 3D culture. RESULTS: FACS analysis revealed a verapamil sensitive SP accounting for 0.93 +/- 0.12% and 0.57 +/- 0.11% of the total epithelial population from both benign and malignant prostates. The benign SP phenotype revealed a heterogeneous cell population consisting predominantly of small basal cells containing minimal cytoplasm. Conversely, the malignant SP was of undetermined acinar origin and with a complete loss of expression of the CDK2 inhibitor p21(WAF1/Cip1). In vitro androgen-enhanced 3D culture of the benign and malignant SP cells led to the production of spheroids which had acinus like morphology and expressed primitive and basal cell markers. Incorporation of the CD133 marker isolated a further SP sub-fraction accounting for 0.037 +/- 0.01% of epithelial cells. CONCLUSIONS: Our observations are consistent with the Hoechst 33342 dye efflux assay isolating a stem cell enriched population which can be further sub-fractionated by CD133 selection. Moreover, the loss of the CDK inhibitor in malignancy is consistent with the hypothesis that neoplastic change originates in the stem cell compartment.


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