PURPOSE: A cell-cell interaction in which in vivo inoculation of androgen-dependent, non-tumorigenic LNCaP and human bone fibroblast resulted in derivation of androgen-independent and metastatic LNCaP subline(C4-2) in castrated hosts. The purpose of this study is to evaluate if human prostate fibroblasts when grown together with LNCaP may promote androgen-independent growth and enhance metastatic potential. MATERIALS AND METHODS: LNCaP cells and human prostate fibroblasts derived either from peripheral or transition zone co-inoculated in athymic mice for 8 weeks, and then mice were castrated. The chimeric tumors were maintained for additional 4 weeks. The LNCaP sublines, designated P4 and T4, were established and characterized. These sublines were co-inoculated again in castrated mice with human prostate fibroblasts for 8-12 weeks. And then second generation LNCaP sublines, P4-2 and T4-2, were established and also characterized. RESULTS: Marked cytogenetic alterations were observed in P4-2, P4, T4-2 and T4 LNCaP sublines in comparison to parental LNCaP. Although LNCaP cells injected orthotopically did not form tumors in castrated hosts, LNCaP sublines formed PSA-producing tumors and had metastatic potentials to lymph node, lung, liver and bone. These P and T sublines had androgen-independent growth characteristics and metastatic potential. CONCLUSIONS: Cell-cell interactions between prostatic epithelium and their surrounding fibroblasts could contribute to androgen-independent characteristics and enhanced metastatic potential of localized prostate cancer in vivo.
Animals ; Cytogenetics ; Epithelium ; Fibroblasts* ; Humans* ; Liver ; Lung ; Lymph Nodes ; Mice ; Mice, Nude ; Parents ; Prostate* ; Prostatic Neoplasms

PURPOSE: Three-dimensional(3-D) organization is critical for both the normal development and, tumor growth and progression. Cell-cell and cell-matrix interactions determine normal prostate development and its subsequent neoplastic transformation. To understand the epigenetic factors that lead to cell transformation, a 3-D human prostate cell culture was established with prostate epithelial cells grown in a rotating-wall vessel(RWV) under microgravity-simulated conditions with either alone or with prostate or bone stromal cells. We tested the hypothesis of whether phenotypic and genotypic alterations of LNCaP cells may be achived when grown as 3-D organoids. MATERIALS AND METHODS: LNCaP cells were seeded in RWV alone and with either human prostate or bone fibroblasts. After period of 2 months, RWV1, 2, and 3 cell lines were established from the prostate organoids and were characterized. RESULTS: While LNCaP cells injected orthotopically failed to form tumors in castrated mouse, RWV-derived cell lines formed PSA-producing tumors and metastasized to lymph node, bone, lung and liver, which stained positively by PSA antibody. RWV cells grew faster than parental LNCaP, especially in sex hormone-free conditions. Unlike parental LNCaP cells which respond positively to androgen and estrogen-induced growth and PSA expression, RWV cells are insensitive to sex steroid-induced growth, but remain sensitive to androgen for induction of PSA expression. Comparative genomic hybridization(CGH) results demonstrated that RWV cell lines have different chromosomal gain and loss each other as compared to those of LNCaP. CONCLUSIONS: Cell-cell and cell-matrix interactions of androgen-dependent and non-metastatic LNCaP cultured alone or with either prostate or bone fibroblasts in 3-D culture under microgravity-simulated conditions resulted in induction of androgen- independent and metastatic LNCaP sublines, RWV cell lines, meaning androgen- independent progression. Phenotype and genotype of RWV cell lines are definitely dissimilar to those of parental LNCaP. 3-D culture of prostatic epithelial cells under microgravity-simulated conditions could be novel approach to the study of normal development and cancer of prostate as an ideal in vitro model and, will be further exploited.
Animals ; Cell Culture Techniques ; Cell Line ; Epigenomics ; Epithelial Cells ; Fibroblasts ; Genotype ; Humans ; Liver ; Lung ; Lymph Nodes ; Mice ; Organoids* ; Parents ; Phenotype ; Prostate* ; Prostatic Neoplasms ; Stromal Cells

PURPOSE: Cell-cell interactions determine normal prostate development and subsequent neoplastic transfor mation. The progression of prostate cancer from androgen-dependent to androgen-independent states involves multiple steps of genetic changes mediated by tumor-microenvironment interactions. To understand the epigenetic factors that lead to progression, we studied if 1) androgen-dependent and non-metastatic LNCaP may interact with prostate or bone fibroblasts under microgravity-simulated conditions in vitro. 2) LNCaP may interact with prostate fibroblasts in vivo, and acquire androgen-independence and metastatic potential. MATERIALS AND METHODS: The LNCaP sublines were generated as follows. 1) LNCaP cells were grown in vitro either alone or with prostate or bone fibroblasts under microgravity-simulated conditions. 2) LNCaP cells were grown in vivo as chimeric tumors with prostate fibroblasts. The LNCaP sublines were characterized by studies of chromosomal analysis, comparative genomic hybridization and, in vivo tumorigenicity and metastatic potential. RESULTS: In comparison to the parental LNCaP cells, the LNCaP sublines underwent permanent genotypic and phenotypic changes manifested in androgen-independence and metastatic potential. CONCLUSION: These results emphasize the importance of cell-cell interaction as a critical determinant that could "induce" or "select" progenies favoring enhanced prostate cancer growth and progression. This concept favors the development of toxic gene therapy targeting both prostate cancer epithelium and supporting bone stroma for an effective eradication and prevention of prostate cancer bone metastasis.
Cell Line* ; Comparative Genomic Hybridization ; Epigenomics ; Epithelium ; Fibroblasts* ; Genetic Therapy ; Humans ; Neoplasm Metastasis ; Parents ; Prostate* ; Prostatic Neoplasms*

OBJECTIVESTo examine the anti-oncogenic effects of promyelocytic leukemia (PML) on bladder cancer and to explore its molecular mechanisms of growth suppression.

METHODSWild-type PML was transfected into bladder cancer cells (5637 cell) and expressed in a replication-deficient adenovirus-mediated gene delivery system and introduced into human bladder cancer cells (5637 cell) in vitro and in vivo. The effect and mechanisms of the PML gene in cell growth, clonogenicity, and tumorigenicity of bladder cancer cells were studied using in vitro and in vivo growth assays, soft agar colony-forming assay, cell cycle analysis, apoptosis assay and in vivo tumorigenicity assay.

RESULTSOverexpression of PML in 5637 cells significantly reduced their growth rate and clonogenicity on soft agar. PML suppressed bladder cancer cell growth by inducing G1 cell cycle arrest and apoptosis. Adenovirus-mediated PML (Ad-PML) significantly suppressed the tumorigenicity and growth of bladder cancer cells. Intratumoral injection of Ad-PML into tumors induced by 5637 cells dramatically suppressed their growth.

CONCLUSIONSThe results indicated that overexpression of PML protein may promote efficient growth inhibition of human bladder cancer cells by inducing G1 cell cycle arrest and apoptosis, and adenovirus-mediated PML (Ad-PML) expression efficiently suppresses human bladder cancer growth.

Adenoviridae ; Animals ; Apoptosis ; physiology ; Cell Division ; physiology ; Cells, Cultured ; Humans ; Male ; Mice ; Mice, Nude ; Neoplasm Proteins ; analysis ; Nuclear Proteins ; Promyelocytic Leukemia Protein ; Transcription Factors ; analysis ; Transfection ; Tumor Cells, Cultured ; Tumor Suppressor Proteins ; Urinary Bladder Neoplasms ; pathology

OBJECTIVETo investigate whether the sonic hedgehog signaling pathway is involved in the development of human fetal prostate, and to evaluate the changing staining patterns of its molecules, sonic hedgehog (SHH), patchedl (PTC1), smoothened (SMO), and GLI1, in the human fetal prostate at various gestation stages.

METHODSFifteen human fetal prostate specimens at various developmental stages (10 - 39 weeks) were included in this study. SHH, PTC1, SMO and GLI1 were detected in all the specimens by immunohistochemical technique. All the slides were observed and assessed under the light microscope.

RESULTSSHH, PTC1, SMO and GLI1 could be detected in human fetal prostate tissues, and their expression formed two surges, the former at week 16, and the latter at week 28. The staining of SHH and SMO was distributed only in the ductal epithelium but not in the stroma. The expression of PTC1 and GLI1 could be found mainly in the epithelium, with minimal staining in the stroma.

CONCLUSIONThe sonic hedgehog signaling pathway is involved in the development of the human fetal prostate. The high expression of its molecules at early gestation stages might be associated with the induction of prostatic buds, while their abundant expression at later gestation stages might be related to the prostate ductal branching, growth, differentiation and morphogenesis.

Gene Expression Regulation, Developmental ; physiology ; Hedgehog Proteins ; biosynthesis ; Humans ; Male ; Oncogene Proteins ; biosynthesis ; Patched Receptors ; Prostate ; embryology ; metabolism ; Receptors, Cell Surface ; biosynthesis ; Receptors, G-Protein-Coupled ; biosynthesis ; Signal Transduction ; physiology ; Smoothened Receptor ; Trans-Activators ; biosynthesis ; Zinc Finger Protein GLI1