In recent years, three dimensional (3D)-culture technique has emerged as a very popular approach to reconstruct tissue architectures and develop experimental models for studying epithelial cancers. However, 3D culture models of prostate epithelial cells to mimic prostate cancer development are relatively rare, making it highly desirable to develop and characterize novel 3D culture models suitable for studying prostate cancer. Recently, epithelial-mesenchymal transition (EMT) has emerged as an important mechanism for cancer cell invasion. The zinc finger transcriptional factor Snail as a key regulator of EMT has been found to contribute to aggressive progression in many types of neoplasms. Even though several studies corroborated that EMT is implicated in prostate cancer, the expression patterns of Snail in normal prostate and prostate cancer, and the functional role of Snail in prostate cancer as well as its relation with EMT are still unknown. Based on this background, my major efforts were to establish a 3D culture model of human prostatic epithelial cells with structural and functional relevance to prostate gland and to employ this model to study the functional role of Snail in the prostate cancer. / When embedded in Matrigel for 3D culture, BPH-1 cells developed into growth-arrested acinar structures with a hollow lumen. Ultrastructural examination of BPH-1 spheroids by electricon microscopy indicated that BPH-1 spheroids displayed a polarized differentiation phenotype. Immunoflurescence analysis of polarized epithelial markers further confirmed that BPH-1 spheroids were polarized. In contrast, tumorigenic BPH-1CAFTD cells exhibited disorganized and continuously proliferating structures in Matrigel, with polarized epithelial markers randomly diffused or completely lost. In addition, BPH-1 CAFTD cells displayed significantly higher invasive capacity in comparison to BPH-1 cells by transwell invasion assay. Moreover, LY294002 treatment of BPH-1CAFTD1 and BPH-1CAFTD3 cells in 3D cultures resulted in impaired cell proliferation as evidenced by reduced colony size and decreased Ki-67 index, and western blot analysis showed that cyclin D1 protein levels were significantly decreased, while p21 protein levels were slightly up-regulated in LY294002-treated 3D cultures. Additionally, LY94002 significantly decreased the invasive capacity of BPH-1CAFTD1 and BPH-1CAFTD3 cells. Interestingly, LY294002 treatment completely reverted the disorganized non-polar 3D structures of BPH-1CAFTD1 cells to well-organized polarized spheroid structures in Matrigel, but failed to restore the polarized differentiation in 3D cultures of BPH-1CAFTD3 cells, which still formed compact aggregates as shown by confocal immunofluorescence analysis. Snail protein was barely detected in the epithelial cells of human benign prostatic tissue but significantly elevated as nuclear protein in primary prostate cancer and bone metastatic specimens by immunohistochemical analysis. Snail transcript levels were weakly expressed in a majority of nonmalignant prostatic epithelial cell lines, while markedly increased in almost all tested cancer cell lines. Snail expression induced a morphological switch to more scattered and spindle-shaped appearance in BPH-1 and BPH-1CAFTD1 cells in 2D culture, and immunofluorescence analysis of several EMT specific markers indicated that Snail-expressing cells underwent EMT. In 3D contexts, Snail-expressing cells developed into more disorganized structures with many cords or protrusions, with a concurrent EMT change as evidenced by reduced E-cadherin and increased vimentin expression. In addition, Snail expression augmented the invasive capacities in both BPH-1 cells and BPH-lCAFTD1 cells, but did not significantly affect the migratory capacities. Snail expression enhanced the MMP2 activity in BPH-1 cells and promoted both MMP-2 and MMP-9 activities in BPH-1CAFTD1 cells. Moreover, Snail expression enhanced anchorage-independent growth capability in BPH-1 cells, but failed to initiate tumor formation in nude-mice. Lastly, Snail expression induced a dramatic increase in FoxC2 and SPARC transcripts but a marked decrease in claudin-1 and p63 transcripts. / Chu, Jianhong. / Adviser: Franky Chan Leung. / Source: Dissertation Abstracts International, Volume: 70-06, Section: B, page: 3448. / Thesis (Ph.D.)--Chinese University of Hong Kong, 2008. / Includes bibliographical references (leaves 143-166). / Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Electronic reproduction. [Ann Arbor, MI] : ProQuest Information and Learning, [200-] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Abstracts in English and Chinese. / School code: 1307.
| Identifer | oai:union.ndltd.org:cuhk.edu.hk/oai:cuhk-dr:cuhk_344238 |
| Date | January 2008 |
| Contributors | Chu, Jianhong., Chinese University of Hong Kong Graduate School. Division of Anatomy. |
| Source Sets | The Chinese University of Hong Kong |
| Language | English, Chinese |
| Detected Language | English |
| Type | Text, theses |
| Format | electronic resource, microform, microfiche, 1 online resource (xiii, 166 leaves : ill.) |
| Rights | Use of this resource is governed by the terms and conditions of the Creative Commons “Attribution-NonCommercial-NoDerivatives 4.0 International” License (http://creativecommons.org/licenses/by-nc-nd/4.0/) |
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