Analysis of secreted proteins for the study of bladder cancer cell aggressiveness

Manousos Makridakis, Maria G. Roubelakis, Vasiliki Bitsika, Veronica Dimuccio, Martina Samiotaki, Sophia Kossida, George Panayotou, Jonathan Coleman, Giovanni Candiano, Nikolaos P. Anagnou, Antonia Vlahou

Research output: Contribution to journalArticle

Abstract

Secreted proteins play a key role in cell signaling, communication, and migration. We recently described the development of an aggressive variant (T24M) of the bladder cancer cell line T24. Using this cell line model, the objective of our work was the identification of secreted proteins involved in the acquisition of the aggressive phenotype. Using in vitro assays, we demonstrate that conditioned media of the T24M cells promote motility of the parental less aggressive T24 cells. Proteomic analysis of cell culture conditioned media by the use of 2-dimensional gel electrophoresis coupled to MALDI TOF MS and LC-MS approaches resulted in enrichment and detection of multiple classical extracellular and secreted proteins such as fibronectin, cystatin, fibrillin, fibulin, interleukin 6, etc. Comparison of the secretome of the T24 and T24M cells indicated differences in proteins with potential involvement in the mechanisms of cell aggressiveness including SPARC, tPA, and clusterin. These findings were further confirmed by Western blot analysis. In the case of SPARC, further studies involving transwell assays indicated that blockage of the protein in the presence of SPARC-specific Abs results in decreased cell motility. Collectively, our study provides a 2DE-based comprehensive analysis of bladder cancer cell secretome. The results indicate various secreted proteins with potential involvement in bladder cancer cell aggressiveness and more specifically provide initial evidence for special role of SPARC in bladder cancer cell motility and invasiveness.

Original languageEnglish
Pages (from-to)3243-3259
Number of pages17
JournalJournal of Proteome Research
Volume9
Issue number6
DOIs
Publication statusPublished - Jun 4 2010

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Urinary Bladder Neoplasms
Cells
Cell Movement
Proteins
Conditioned Culture Medium
Assays
Far-Western Blotting
Cystatins
Clusterin
Cell signaling
Cell Line
Matrix-Assisted Laser Desorption-Ionization Mass Spectrometry
Electrophoresis
Fibronectins
Cell culture
Cell Communication
Proteomics
Culture Media
Interleukin-6
Identification (control systems)

Keywords

  • 2DE
  • bladder cancer
  • proteomics
  • secretome
  • SPARC
  • T24

ASJC Scopus subject areas

  • Biochemistry
  • Chemistry(all)

Cite this

Makridakis, M., Roubelakis, M. G., Bitsika, V., Dimuccio, V., Samiotaki, M., Kossida, S., ... Vlahou, A. (2010). Analysis of secreted proteins for the study of bladder cancer cell aggressiveness. Journal of Proteome Research, 9(6), 3243-3259. https://doi.org/10.1021/pr100189d

Analysis of secreted proteins for the study of bladder cancer cell aggressiveness. / Makridakis, Manousos; Roubelakis, Maria G.; Bitsika, Vasiliki; Dimuccio, Veronica; Samiotaki, Martina; Kossida, Sophia; Panayotou, George; Coleman, Jonathan; Candiano, Giovanni; Anagnou, Nikolaos P.; Vlahou, Antonia.

In: Journal of Proteome Research, Vol. 9, No. 6, 04.06.2010, p. 3243-3259.

Research output: Contribution to journalArticle

Makridakis, M, Roubelakis, MG, Bitsika, V, Dimuccio, V, Samiotaki, M, Kossida, S, Panayotou, G, Coleman, J, Candiano, G, Anagnou, NP & Vlahou, A 2010, 'Analysis of secreted proteins for the study of bladder cancer cell aggressiveness', Journal of Proteome Research, vol. 9, no. 6, pp. 3243-3259. https://doi.org/10.1021/pr100189d
Makridakis M, Roubelakis MG, Bitsika V, Dimuccio V, Samiotaki M, Kossida S et al. Analysis of secreted proteins for the study of bladder cancer cell aggressiveness. Journal of Proteome Research. 2010 Jun 4;9(6):3243-3259. https://doi.org/10.1021/pr100189d
Makridakis, Manousos ; Roubelakis, Maria G. ; Bitsika, Vasiliki ; Dimuccio, Veronica ; Samiotaki, Martina ; Kossida, Sophia ; Panayotou, George ; Coleman, Jonathan ; Candiano, Giovanni ; Anagnou, Nikolaos P. ; Vlahou, Antonia. / Analysis of secreted proteins for the study of bladder cancer cell aggressiveness. In: Journal of Proteome Research. 2010 ; Vol. 9, No. 6. pp. 3243-3259.
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