title: Enrichment of prostate cancer stem cells from primary prostate cancer cultures of biopsy samples.
abstract: This study was to enrich prostate cancer stem cells (PrCSC) from primary prostate cancer cultures (PPrCC). Primary prostate cancer cells were amplified in keratinocyte serum-free medium with epidermal growth factor (EGF) and bovine pituitary extract (BPE), supplemented with leukemia inhibitory factor (LIF), stem cell factor (SCF) and cholera toxin. After amplification, cells were transferred into ultra-low attachment dishes with serum-free DMEM/F12 medium, supplemented with EGF, basic fibroblast growth factor (bFGF), bovine serum albumin (BSA), insulin, and N2 nutrition. Expression of cell-type-specific markers was determined by RT-qPCR and immunostaining. Tumorigenicity of enriched PrCSC was determined by soft agar assay and xenograft assay in NOD/SCID mice. Biopsy samples from 19 confirmed prostate cancer patients were used for establishing PPrCC, and 18 cases (95%) succeeded. Both basal marker (CK5) and luminal markers (androgen receptor and CK8) strongly co-expressed in most of PPrCC, indicating their basal epithelial origin. After amplification under adherent culture condition in vitro, transient amplifying cells were the dominant cells. Sphere formation efficiency (SFE) of passaged PPrCC was about 0.5%, which was 27 times lower than SFE of LNCaP (13.67%) in the same condition. Compared with adherent cells from PPrCC, prostasphere from PPrCC showed up regulated stem cell markers and increased tumorigenic potential in soft-agar assay. However, spheroid cells from PPrCC prostasphere failed to initiate tumor in xenograft assay in 6 months. Thus, PPrCC can be established and amplified from prostate cancer biopsy samples. Our modified sphere culture system can enrich PrCSC from PPrCC.
authors.affiliation: Institute of Life Science, College of Life Sciences and Food Engineering, Nanchang University Nanchang, Jiangxi 330031, China ; State Key Laboratory of Molecular Biology, Shanghai Key Laboratory of Andrology, Institute of Biochemistry and Cell Biology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences Shanghai 200031, China.
authors.forename: Shunqi
authors.initials: S
authors.lastname: Wang
authors.affiliation: Department of Urology, Affiliated Tongji Hospital of Tongji Universit Shanghai, Shanghai 200065, China.
authors.forename: Shengsong
authors.initials: S
authors.lastname: Huang
authors.affiliation: Department of Urology, Affiliated Tongji Hospital of Tongji Universit Shanghai, Shanghai 200065, China.
authors.forename: Xin
authors.initials: X
authors.lastname: Zhao
authors.affiliation: Department of Urology, Affiliated Tongji Hospital of Tongji Universit Shanghai, Shanghai 200065, China.
authors.forename: Qimin
authors.initials: Q
authors.lastname: Zhang
authors.affiliation: Department of Urology, Affiliated Tongji Hospital of Tongji Universit Shanghai, Shanghai 200065, China.
authors.forename: Min
authors.initials: M
authors.lastname: Wu
authors.affiliation: State Key Laboratory of Molecular Biology, Shanghai Key Laboratory of Andrology, Institute of Biochemistry and Cell Biology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences Shanghai 200031, China.
authors.forename: Feng
authors.initials: F
authors.lastname: Sun
authors.affiliation: State Key Laboratory of Molecular Biology, Shanghai Key Laboratory of Andrology, Institute of Biochemistry and Cell Biology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences Shanghai 200031, China.
authors.forename: Gang
authors.initials: G
authors.lastname: Han
authors.affiliation: Department of Urology, Affiliated Tongji Hospital of Tongji Universit Shanghai, Shanghai 200065, China.
authors.forename: Denglong
authors.initials: D
authors.lastname: Wu
mesh_terms: D000818:Animals; D001706:Biopsy; D018929:Cell Culture Techniques / Q000379:methods*; D064593:Heterografts; D006801:Humans; D007150:Immunohistochemistry; D008297:Male; D051379:Mice; D016688:Mice, Inbred NOD; D016513:Mice, SCID; D014411:Neoplastic Stem Cells / Q000166:cytology*; D011471:Prostatic Neoplasms; D060888:Real-Time Polymerase Chain Reaction; D014407:Tumor Cells, Cultured
keywords: Prostate cancer (PrCa); primary prostate cancer cultures (PPrCC); prostate cancer stem cells (PrCSC); sphere culture