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Regulation of cell proliferation in a stratified culture system of epithelial cells from prostate tissue

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Abstract

Mechanisms controlling epithelial proliferation and differentiation in the prostate have been primarily investigated in mouse models. The regulation of proliferation and differentiation is poorly understood in human prostate epithelial cells. In vivo, the glandular prostate epithelium consists of a p63-positive proliferating basal cell layer and a post-mitotic p27-positive secretory cell layer. We have established an organized stratified culture system of human primary prostate epithelial cells to gain insight into mechanisms regulating proliferation and differentiation. In this system, expression of p63 is observed in the bottom layer. In addition, BrdU incorporation persists even though cells are confluent. In contrast, in the upper layer, p63 expression is greatly diminished, p27 is expressed, and the cells are growth arrested. Overexpression of cyclin D1 or knockdown of p27 does not increase proliferation. After inactivation of the nuclear phosphoprotein Rb, the cell layers remain organized and cell proliferation increases only in the bottom layer. Furthermore, the expression of p63 remains confined to the bottom layer after Rb inactivation. Altogether, this in vitro model recapitulates certain aspects of in vivo growth regulation and differentiation and suggests that the loss of Rb family proteins in human cells trigger hyperplasia but is not sufficient for transformation.

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References

  • Barnes DM, Gillett CE (1998) Cyclin D1 in breast cancer. Breast Cancer Res Treat 52:1–15

    Article  PubMed  CAS  Google Scholar 

  • Bayne CW, Ross M, Donnelly F, Chapman K, Buck C, Bollina P, Habib FK (1998) Selective interactions between prostate fibroblast and epithelial cells in co-culture maintain the BPH phenotype. Urol Int 61:1–7

    Article  PubMed  CAS  Google Scholar 

  • Bonkhoff H, Stein U, Remberger K (1994) The proliferative function of basal cells in the normal and hyperplastic human prostate. Prostate 24:114–118

    Article  PubMed  CAS  Google Scholar 

  • Bowen C, Bubendorf L, Voeller HJ, Slack R, Willi N, Sauter G, Gasser TC, Koivisto P, Lack EE, Kononen J, Kallioniemi OP, Gelmann EP (2000) Loss of NKX3.1 expression in human prostate cancers correlates with tumor progression. Cancer Res 60:6111–6115

    PubMed  CAS  Google Scholar 

  • Brooks JD, Bova GS, Isaacs WB (1995) Allelic loss of the retinoblastoma gene in primary human prostatic adenocarcinomas. Prostate 26:35–39

    Article  PubMed  CAS  Google Scholar 

  • Claudio PP, Zamparelli A, Garcia FU, Claudio L, Ammirati G, Farina A, Bovicelli A, Russo G, Giordano GG, McGinnis DE, Giordano A, Cardi G (2002) Expression of cell-cycle-regulated proteins pRb2/p130, p107, p27(kip1), p53, mdm-2, and Ki-67 (MIB-1) in prostatic gland adenocarcinoma. Clin Cancer Res 8:1808–1815

    PubMed  CAS  Google Scholar 

  • Collins AT, Habib FK, Maitland NJ, Neal DE (2001) Identification and isolation of human prostate epithelial stem cells based on alpha(2)beta(1)-integrin expression. J Cell Sci 114:3865–3872

    PubMed  CAS  Google Scholar 

  • Cunha GR (1984) Androgenic effects upon prostatic epithelium are mediated via trophic influences from stroma. Prog Clin Biol Res 145:81–102

    PubMed  CAS  Google Scholar 

  • Day KC, McCabe MT, Zhao X, Wang Y, Davis JN, Phillips J, Von Geldern M, Ried T, KuKuruga MA, Cunha GR, Hayward SW, Day ML (2002) Rescue of embryonic epithelium reveals that the homozygous deletion of the retinoblastoma gene confers growth factor independence and immortality but does not influence epithelial differentiation or tissue morphogenesis. J Biol Chem 277:44475–44484

    Article  PubMed  CAS  Google Scholar 

  • De Marzo AM, Meeker AK, Epstein JI, Coffey DS (1998a) Prostate stem cell compartments: expression of the cell cycle inhibitor p27Kip1 in normal, hyperplastic, and neoplastic cells. Am J Pathol 153:911–919

    Google Scholar 

  • De Marzo AM, Nelson WG, Meeker AK, Coffey DS (1998b) Stem cell features of benign and malignant prostate epithelial cells. J Urol 160:2381–2392

    Article  Google Scholar 

  • Drobnjak M, Osman I, Scher HI, Fazzari M, Cordon-Cardo C (2000) Overexpression of cyclin D1 is associated with metastatic prostate cancer to bone. Clin Cancer Res 6:1891–1895

    PubMed  CAS  Google Scholar 

  • Erdamar S, Yang G, Harper JW, Lu X, Kattan MW, Thompson TC, Wheeler TM (1999) Levels of expression of p27KIP1 protein in human prostate and prostate cancer: an immunohistochemical analysis. Mod Pathol 12:751–755

    PubMed  CAS  Google Scholar 

  • Fry PM, Hudson DL, O'Hare MJ, Masters JR (2000) Comparison of marker protein expression in benign prostatic hyperplasia in vivo and in vitro. BJU Int 85:504–513

    Article  PubMed  CAS  Google Scholar 

  • Gao H, Ouyang X, Banach-Petrosky W, Borowsky AD, Lin Y, Kim M, Lee H, Shih WJ, Cardiff RD, Shen MM, Abate-Shen C (2004) A critical role for p27kip1 gene dosage in a mouse model of prostate carcinogenesis. Proc Natl Acad Sci USA 101:17204–17209

    Article  PubMed  CAS  Google Scholar 

  • Garraway LA, Lin D, Signoretti S, Waltregny D, Dilks J, Bhattacharya N, Loda M (2003) Intermediate basal cells of the prostate: in vitro and in vivo characterization. Prostate 55:206–218

    Article  PubMed  Google Scholar 

  • Geng Y, Yu Q, Sicinska E, Das M, Bronson RT, Sicinski P (2001) Deletion of the p27Kip1 gene restores normal development in cyclin D1-deficient mice. Proc Natl Acad Sci USA 98:194–199

    Article  PubMed  CAS  Google Scholar 

  • Gmyrek GA, Walburg M, Webb CP, Yu HM, You X, Vaughan ED, Vande Woude GF, Knudsen BS (2001) Normal and malignant prostate epithelial cells differ in their response to hepatocyte growth factor/scatter factor. Am J Pathol 159:579–590

    PubMed  CAS  Google Scholar 

  • Grandori C, Wu KJ, Fernandez P, Ngouenet C, Grim J, Clurman BE, Moser MJ, Oshima J, Russell DW, Swisshelm K, Frank S, Amati B, Dalla-Favera R, Monnat RJ Jr (2003) Werner syndrome protein limits MYC-induced cellular senescence. Genes Dev 17:1569–1574

    Article  PubMed  CAS  Google Scholar 

  • Gumbiner LM, Gumerlock PH, Mack PC, Chi SG, deVere White RW, Mohler JL, Pretlow TG, Tricoli JV (1999) Overexpression of cyclin D1 is rare in human prostate carcinoma. Prostate 38:40–45

    Article  PubMed  CAS  Google Scholar 

  • Halbert CL, Demers GW, Galloway DA (1991) The E7 gene of human papillomavirus type 16 is sufficient for immortalization of human epithelial cells. J Virol 65:473–478

    PubMed  CAS  Google Scholar 

  • Hall JA, Maitland NJ, Stower M, Lang SH (2002) Primary prostate stromal cells modulate the morphology and migration of primary prostate epithelial cells in type 1 collagen gels. Cancer Res 62:58–62

    PubMed  CAS  Google Scholar 

  • Hanahan D, Weinberg RA (2000) The hallmarks of cancer. Cell 100:57–70

    Article  PubMed  CAS  Google Scholar 

  • Hayward SW, Haughney PC, Rosen MA, Greulich KM, Weier HU, Dahiya R, Cunha GR (1998) Interactions between adult human prostatic epithelium and rat urogenital sinus mesenchyme in a tissue recombination model. Differentiation 63:131–140

    Article  PubMed  CAS  Google Scholar 

  • Henshall SM, Quinn DI, Lee CS, Head DR, Golovsky D, Brenner PC, Delprado W, Stricker PD, Grygiel JJ, Sutherland RL (2001) Overexpression of the cell cycle inhibitor p16INK4A in high-grade prostatic intraepithelial neoplasia predicts early relapse in prostate cancer patients. Clin Cancer Res 7:544–550

    PubMed  CAS  Google Scholar 

  • Hudson DL, O'Hare M, Watt FM, Masters JR (2000) Proliferative heterogeneity in the human prostate: evidence for epithelial stem cells. Lab Invest 80:1243–1250

    Article  PubMed  CAS  Google Scholar 

  • Ittmann MM, Wieczorek R (1996) Alterations of the retinoblastoma gene in clinically localized, stage B prostate adenocarcinomas. Hum Pathol 27:28–34

    Article  PubMed  CAS  Google Scholar 

  • King KE, Ponnamperuma RM, Yamashita T, Tokino T, Lee LA, Young MF, Weinberg WC (2003) deltaNp63alpha functions as both a positive and a negative transcriptional regulator and blocks in vitro differentiation of murine keratinocytes. Oncogene 22:3635–3644

    Article  PubMed  CAS  Google Scholar 

  • Koster MI, Kim S, Mills AA, DeMayo FJ, Roop DR (2004) p63 is the molecular switch for initiation of an epithelial stratification program. Genes Dev 18:126–131

    Article  PubMed  CAS  Google Scholar 

  • Kubota Y, Fujinami K, Uemura H, Dobashi Y, Miyamoto H, Iwasaki Y, Kitamura H, Shuin T (1995) Retinoblastoma gene mutations in primary human prostate cancer. Prostate 27:314–320

    Article  PubMed  CAS  Google Scholar 

  • Kurita T, Medina RT, Mills AA, Cunha GR (2004) Role of p63 and basal cells in the prostate. Development 131:4955–4964

    Article  PubMed  CAS  Google Scholar 

  • Lang SH, Sharrard RM, Stark M, Villette JM, Maitland NJ (2001) Prostate epithelial cell lines form spheroids with evidence of glandular differentiation in three-dimensional Matrigel cultures. Br J Cancer 85:590–599

    Article  PubMed  CAS  Google Scholar 

  • Lee CT, Capodieci P, Osman I, Fazzari M, Ferrara J, Scher HI, Cordon-Cardo C (1999) Overexpression of the cyclin-dependent kinase inhibitor p16 is associated with tumor recurrence in human prostate cancer. Clin Cancer Res 5:977–983

    PubMed  CAS  Google Scholar 

  • Leenders G van, Dijkman H, Hulsbergen-van de Kaa C, Ruiter D, Schalken J (2000) Demonstration of intermediate cells during human prostate epithelial differentiation in situ and in vitro using triple-staining confocal scanning microscopy. Lab Invest 80:1251–1258

    PubMed  Google Scholar 

  • Leenders G van, Balken B van, Aalders T, Hulsbergen-van de Kaa C, Ruiter D, Schalken J (2002) Intermediate cells in normal and malignant prostate epithelium express c-MET: implications for prostate cancer invasion. Prostate 51:98–107

    Article  PubMed  CAS  Google Scholar 

  • Leenders GJ van, Gage WR, Hicks JL, Balken B van, Aalders TW, Schalken JA, De Marzo AM (2003) Intermediate cells in human prostate epithelium are enriched in proliferative inflammatory atrophy. Am J Pathol 162:1529–1537

    PubMed  Google Scholar 

  • Liefer KM, Koster MI, Wang XJ, Yang A, McKeon F, Roop DR (2000) Down-regulation of p63 is required for epidermal UV-B-induced apoptosis. Cancer Res 60:4016–4020

    PubMed  CAS  Google Scholar 

  • Maddison LA, Sutherland BW, Barrios RJ, Greenberg NM (2004a) Conditional deletion of Rb causes early stage prostate cancer. Cancer Res 64:6018–6025

    Article  CAS  Google Scholar 

  • Maddison LA, Huss WJ, Barrios RM, Greenberg NM (2004b) Differential expression of cell cycle regulatory molecules and evidence for a "cyclin switch" during progression of prostate cancer. Prostate 58:335–344

    Article  CAS  Google Scholar 

  • Oyama T, Kashiwabara K, Yoshimoto K, Arnold A, Koerner F (1998) Frequent overexpression of the cyclin D1 oncogene in invasive lobular carcinoma of the breast. Cancer Res 58:2876–2880

    PubMed  CAS  Google Scholar 

  • Parsons JK, Gage WR, Nelson WG, De Marzo AM (2001) p63 protein expression is rare in prostate adenocarcinoma: implications for cancer diagnosis and carcinogenesis. Urology 58:619–624

    Article  PubMed  CAS  Google Scholar 

  • Peehl DM, Leung GK, Wong ST (1994) Keratin expression: a measure of phenotypic modulation of human prostatic epithelial cells by growth inhibitory factors. Cell Tissue Res 277:11–18

    Article  PubMed  CAS  Google Scholar 

  • Petre CE, Wetherill YB, Danielsen M, Knudsen KE (2002) Cyclin D1: mechanism and consequence of androgen receptor co-repressor activity. J Biol Chem 277:2207–2215

    Article  PubMed  CAS  Google Scholar 

  • Petre-Draviam CE, Cook SL, Burd CJ, Marshall TW, Wetherill YB, Knudsen KE (2003) Specificity of cyclin D1 for androgen receptor regulation. Cancer Res 63:4903–4913

    PubMed  CAS  Google Scholar 

  • Phillips SM, Barton CM, Lee SJ, Morton DG, Wallace DM, Lemoine NR, Neoptolemos JP (1994) Loss of the retinoblastoma susceptibility gene (RB1) is a frequent and early event in prostatic tumorigenesis. Br J Cancer 70:1252–1257

    PubMed  CAS  Google Scholar 

  • Rhim JS (2000) In vitro human cell culture models for the study of prostate cancer. Prostate Cancer Prostatic Dis 3:229–235

    Article  PubMed  Google Scholar 

  • Robinson EJ, Neal DE, Collins AT (1998) Basal cells are progenitors of luminal cells in primary cultures of differentiating human prostatic epithelium. Prostate 37:149–160

    Article  PubMed  CAS  Google Scholar 

  • Sarkar FH, Sakr WA, Li YW, Jacobs J, Crissman JD (1996) Tumor suppressor p53 gene mutation in squamous cell carcinoma of the larynx. Diagn Mol Pathol 5:201–205

    Article  PubMed  CAS  Google Scholar 

  • Shaffer DR, Viale A, Ishiwata R, Leversha M, Olgac S, Manova K, Satagopan J, Scher H, Koff A (2005) Evidence for a p27 tumor suppressive function independent of its role regulating cell proliferation in the prostate. Proc Natl Acad Sci USA 102:210–215

    Article  PubMed  CAS  Google Scholar 

  • Sherr CJ (1996) Cancer cell cycles. Science 274:1672–1677

    Article  PubMed  CAS  Google Scholar 

  • Sherr CJ, Roberts JM (2004) Living with or without cyclins and cyclin-dependent kinases. Genes Dev 18:2699–2711

    Article  PubMed  CAS  Google Scholar 

  • Sicinski P, Donaher JL, Parker SB, Li T, Fazeli A, Gardner H, Haslam SZ, Bronson RT, Elledge SJ, Weinberg RA (1995) Cyclin D1 provides a link between development and oncogenesis in the retina and breast. Cell 82:621–630

    Article  PubMed  CAS  Google Scholar 

  • Signoretti S, Waltregny D, Dilks J, Isaac B, Lin D, Garraway L, Yang A, Montironi R, McKeon F, Loda M (2000) p63 is a prostate basal cell marker and is required for prostate development. Am J Pathol 157:1769–1775

    PubMed  CAS  Google Scholar 

  • Sutherland RL, Musgrove EA (2002) Cyclin D1 and mammary carcinoma: new insights from transgenic mouse models. Breast Cancer Res 4:14–17

    Article  PubMed  CAS  Google Scholar 

  • Thomas DM, Johnson SA, Sims NA, Trivett MK, Slavin JL, Rubin BP, Waring P, McArthur GA, Walkley CR, Holloway AJ, Diyagama D, Grim JE, Clurman BE, Bowtell DD, Lee JS, Gutierrez GM, Piscopo DM, Carty SA, Hinds PW (2004) Terminal osteoblast differentiation, mediated by runx2 and p27KIP1, is disrupted in osteosarcoma. J Cell Biol 167:925–934

    Article  PubMed  CAS  Google Scholar 

  • Tonini T, Bagella L, D'Andrilli G, Claudio PP, Giordano A (2004) Ezh2 reduces the ability of HDAC1-dependent pRb2/p130 transcriptional repression of cyclin A. Oncogene 23:4930–4937

    Article  PubMed  CAS  Google Scholar 

  • Tricoli JV, Gumerlock PH, Yao JL, Chi SG, D'Souza SA, Nestok BR, deVere White RW (1996) Alterations of the retinoblastoma gene in human prostate adenocarcinoma. Genes Chromosomes Cancer 15:108–114

    Article  PubMed  CAS  Google Scholar 

  • Tsihlias J, Kapusta LR, DeBoer G, Morava-Protzner I, Zbieranowski I, Bhattacharya N, Catzavelos GC, Klotz LH, Slingerland JM (1998) Loss of cyclin-dependent kinase inhibitor p27Kip1 is a novel prognostic factor in localized human prostate adenocarcinoma. Cancer Res 58:542–548

    PubMed  CAS  Google Scholar 

  • Uzgare AR, Isaacs JT (2004) Enhanced redundancy in Akt and mitogen-activated protein kinase-induced survival of malignant versus normal prostate epithelial cells. Cancer Res 64:6190–6199

    Article  PubMed  CAS  Google Scholar 

  • Uzgare AR, Xu Y, Isaacs JT (2004) In vitro culturing and characteristics of transit amplifying epithelial cells from human prostate tissue. J Cell Biochem 91:196–205

    Article  PubMed  CAS  Google Scholar 

  • Varambally S, Dhanasekaran SM, Zhou M, Barrette TR, Kumar-Sinha C, Sanda MG, Ghosh D, Pienta KJ, Sewalt RG, Otte AP, Rubin MA, Chinnaiyan AM (2002) The polycomb group protein EZH2 is involved in progression of prostate cancer. Nature 419:624–629

    Article  PubMed  CAS  Google Scholar 

  • Wang TC, Cardiff RD, Zukerberg L, Lees E, Arnold A, Schmidt EV (1994) Mammary hyperplasia and carcinoma in MMTV-cyclin D1 transgenic mice. Nature 369:669–671

    Article  PubMed  CAS  Google Scholar 

  • Wang Y, Hayward SW, Donjacour AA, Young P, Jacks T, Sage J, Dahiya R, Cardiff RD, Day ML, Cunha GR (2000) Sex hormone-induced carcinogenesis in Rb-deficient prostate tissue. Cancer Res 60:6008–6017

    PubMed  CAS  Google Scholar 

  • Wang Y, Hayward S, Cao M, Thayer K, Cunha G (2001) Cell differentiation lineage in the prostate. Differentiation 68:270–279

    Article  PubMed  CAS  Google Scholar 

  • Webber MM, Bello D, Kleinman HK, Wartinger DD, Williams DE, Rhim JS (1996) Prostate specific antigen and androgen receptor induction and characterization of an immortalized adult human prostatic epithelial cell line. Carcinogenesis 17:1641–646

    Article  PubMed  CAS  Google Scholar 

  • Webber MM, Bello D, Kleinman HK, Hoffman MP (1997) Acinar differentiation by non-malignant immortalized human prostatic epithelial cells and its loss by malignant cells. Carcinogenesis 18:1225–1231

    Article  PubMed  CAS  Google Scholar 

  • Xin L, Ide H, Kim Y, Dubey P, Witte ON (2003) In vivo regeneration of murine prostate from dissociated cell populations of postnatal epithelia and urogenital sinus mesenchyme. Proc Natl Acad Sci USA 100 (Suppl 1):11896–11903

    Article  PubMed  CAS  Google Scholar 

  • Yang RM, Naitoh J, Murphy M, Wang HJ, Phillipson J, deKernion JB, Loda M, Reiter RE (1998) Low p27 expression predicts poor disease-free survival in patients with prostate cancer. J Urol 159:941–945

    Article  PubMed  CAS  Google Scholar 

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Acknowledgements

The authors thank Ivy Yu and Weldon Debusk for excellent technical assistance, and Leona Cohen-Gould and the Medical Imaging Core at FHCRC for help with the acquisition and analysis of confocal images. B.S.K. is grateful to Selina Chen-Kiang for her encouragement to establish and explore primary cell cultures. The authors are also grateful to Donna Peehl for helpful comments on the manuscript.

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Authors and Affiliations

  1. Division of Public Health Sciences, Program in Cancer Biology, PHS Building, M5-A864, Fred Hutchinson Cancer Research Center, 1100 Fairview Avenue N., Seattle, WA 98109, USA

    Chang Xu & Beatrice S. Knudsen

  2. Clinical Research and Human Biology, Fred Hutchinson Cancer Research Center, Seattle, WA 98109, USA

    Michael P. Gustafson, Jonathan E. Grim & Bruce E. Clurman

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  1. Michael P. Gustafson
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  2. Chang Xu
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  3. Jonathan E. Grim
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Correspondence to Beatrice S. Knudsen.

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This work was supported by the Departments of Pathology and Urology at Weill Medial College, by grants DAMD-17-02-1-0159, MEDC-GR-355, and P30 CA015704-30, and by grant RO1CA84069 to B.E.C.

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Gustafson, M.P., Xu, C., Grim, J.E. et al. Regulation of cell proliferation in a stratified culture system of epithelial cells from prostate tissue. Cell Tissue Res 325, 263–276 (2006). https://doi.org/10.1007/s00441-005-0093-0

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