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Quantitative Analysis of Melanocyte Migration in vitro Based on Automated Cell Tracking under Phase Contrast Microscopy Considering the Combined Influence of Cell Division and Cell-Matrix Interactions

Published online by Cambridge University Press:  03 February 2010

V. Letort
Affiliation:
Laboratory of Applied Mathematics, Ecole Centrale Paris, F-92295 Chatenay-Malabry, France
S. Fouliard
Affiliation:
Laboratory of Applied Mathematics, Ecole Centrale Paris, F-92295 Chatenay-Malabry, France
G. Letort
Affiliation:
Laboratory of Applied Mathematics, Ecole Centrale Paris, F-92295 Chatenay-Malabry, France
I. Adanja
Affiliation:
Laboratory of Image Synthesis and Analysis (LISA), Faculty of Applied Sciences, Universite Libre de Bruxelles, 1050 Brussels, Belgium
M. Kumasaka
Affiliation:
Institut Curie, Centre Recherche, Developmental Genetics of Melanocytes, F-91405 Orsay, France CNRS UMR146, F-91405 Orsay, France
S. Gallagher
Affiliation:
Institut Curie, Centre Recherche, Developmental Genetics of Melanocytes, F-91405 Orsay, France CNRS UMR146, F-91405 Orsay, France
O. Debeir
Affiliation:
Laboratory of Image Synthesis and Analysis (LISA), Faculty of Applied Sciences, Universite Libre de Bruxelles, 1050 Brussels, Belgium
L. Larue
Affiliation:
Institut Curie, Centre Recherche, Developmental Genetics of Melanocytes, F-91405 Orsay, France CNRS UMR146, F-91405 Orsay, France
F. Xavier*
Affiliation:
Laboratory of Applied Mathematics, Ecole Centrale Paris, F-92295 Chatenay-Malabry, France
*
* Corresponding author. E-mail: francoise.xavier@ecp.fr
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Abstract

The aim of this study was to describe and analyze the regulation and spatio-temporal dynamics of melanocyte migration in vitro and its coupling to cell division and interaction with the matrix. The melanocyte lineage is particularly interesting because it is involved in both embryonic development and oncogenesis/metastasis (melanoma). Biological experiments were performed on two melanocyte cell lines established from wild-type and β-catenin-transgenic mice (bcat*). The multi-functional β-catenin molecule is known to be able to regulate the transcription of various genes involved in cell proliferation and migration, particularly in the melanocyte lineage. We also investigated fibronectin, an extra-cellular matrix protein that binds integrins, thereby providing adhesion points for cells and encouraging migration. As the migration of individual cells were followed, automated methods were required for processing the large amount of data generated by the time-lapse video-microscopy. A model-based approach for automated cell tracking was evaluated on a sample by comparison with manual tracking. This method was found reliable in studying overall cell behaviour. Its application to all the observed sequences provided insight into the factors affecting melanocyte migration in vitro: melanocytes of mutated form of β-catenin showed higher division rates and no contact inhibition of growth was induced by the resulting increase in cell density. However, cell density limited the amplitude of cell displacements, although their motility was less affected. The high fibronectin concentration bound to substratum promoted cell migration and motility, the effect being more intense for wild-type cells than for cells with β-catenin over-expression. During the division process, cell migration speed increased rapidly then decreased slowly. Analyses of such data is expected to lead both to biological answers and to a framework for a better modeling processes in the future.

Type
Research Article
Copyright
© EDP Sciences, 2010

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References

Abal, M., Piel, M., Bouckson-Castaing, V., Mogensen, M., Sibarita, J.-B. Bornens, M.. Microtubule release from the centrosome in migrating cells . The Journal of Cell Biology, 159 (2002), 731737 CrossRefGoogle ScholarPubMed
Allard, S., Adin, P., Goudard, L., di Clemente, N., Josso, N., Orgebin-Crist, M.-C., Picard, J.-Y. Xavier, F.. Molecular mechanisms of hormone-mediated Müllerian duct regression: Involment of beta-catenin . Development, 127 (2000), 3349360 Google Scholar
Busam, K.J., Charles, C., Lee, G. Halpern, A.C.. Morphologic Features of Melanocytes, Pigmented Keratinocytes, and Melanophages by In Vivo Confocal Scanning Laser Microscopy . Mod. Pathol., 14 (2001), 862868 CrossRefGoogle ScholarPubMed
Cheng, Y.. Mean shift, mode seeking, and clustering . IEEE Trans. Pattern Anal. Mach. Intell., 17 (1995), No. 8, 790799 CrossRefGoogle Scholar
Dammermann, A., Desai, A. Oegema, K.. The minus end in sight . Current Biology, 13 (2003), 614624 CrossRefGoogle ScholarPubMed
Debeir, O., Camby, I., Kiss, R., Van Ham, P. Decaescker, C.. A Model-Based Approach for Automated In Vitro Cell Tracking and Chemotaxis Analyses . Cytometry, 60A (2004), 2940 CrossRefGoogle Scholar
Debeir, O., Van Ham, P., Kiss, R. Decaestecker, C.. Tracking of Migrating Cells under Phase-contrast Video Microscopy with Combined Mean-Shift Processes . IEEE Transaction on Medical Imaging., 24 (2005), No. 6, 697711 CrossRefGoogle ScholarPubMed
Debeir, O., Mégalizzi, V., Warzée, N., Kiss, R. Decaestecker, C.. Videomicroscopic extraction of specific information on cell proliferation and migration in vitro . Experim. Cell Res., 314 (2008), 29852998 CrossRefGoogle ScholarPubMed
Delmas, V., Beermann, F., Martinozzi, S., Carreira, S., Ackermann, J., Kumasaka, M., Denat, L., Goodall, J., Luciani, F., Viros, A., Demirkan, N., Bastian, B.C., Goding, C.R. Larue, L.. beta-catenin induces immortalisation of melanocytes by suppressing p16INK4a expression and co-operates with N-Ras in melanoma development . Genes & Dev., 21 (2007), 2923-2935 CrossRefGoogle Scholar
Desban, N., Duband, J-L.. Avian neural crest cell migration on laminin: interaction of the a11 integrin with distinct laminin-1 domains mediates different adhesive responses . J. Cell Sci. 110 (1997), 27292744 Google Scholar
Decaestecker, C., Debeir, O., Van Ham, P. Kiss, R.. Can anti-migratory drugs be screened in vitro ? A review of 2D and 3D assays for the quantitative analysis of cell migration . Inc. Med. Res. Rev., 27 (2007), No. 2, 149176 CrossRefGoogle ScholarPubMed
R Development Core Team. R: A Language and Environment for Statistical Computing. R Foundation for Statistical Computing, Vienna, Austria., (2008), http://www.R-project.org
DiMilla, P.A., Stone, J.A., Quinn, J.A., Albelda, S.M. Lauffenburger, D.A.. Maximal Migration of Human Smooth Muscle Cells on Fibronectin and Type IV Collagen Occurs at an Intermediate Attachment Strength . J Cell Biol., 122 (1993), 72937 CrossRefGoogle ScholarPubMed
H, A.J., Sun, T., Barber, D.C., Hose, D.R. Macneil, S.. Automated tracking of migrating cells in phase-contrast video microscopy sequences using image registration . J. Microscopy., 234 (2009), 6279 Google Scholar
Hamilton, N.. Quantification and its Applications in Fluorescent Microscopy Imaging . Traffic, 10 (2009), 951961 CrossRefGoogle ScholarPubMed
Hazgui, S., Bonnet, N., Cutrona, J., Nawrocki-Raby, B., Polette, M., Chouchane, L., Birembaut, P. Zahm, J-M.. 3D culture model and computer assisted videomicroscopy to analyze migratory behavior of noninvasive and invasive bronchial epithelial cells . Am. J. Physiol. Cell Physiol., 289 (2005), 15471552 CrossRefGoogle ScholarPubMed
Holly, S.P., Larson, M.K. Parise, L.V.. Minireview. Multiple Roles of Integrins in Cell Motility . Exp. Cell Res., 261 (2000), 69-74 CrossRefGoogle Scholar
Jouneau, A., YU, Y.-Q., Pasdar, M. Larue, L.. Plasticity of Cadherin-Catenin Expression in the Melanocyte Lineage . Pigment Cell Res. 13 (2000), No. 4, 260272 CrossRefGoogle ScholarPubMed
Knust, E. Huttner, W.B.. Cell polarity from cell division . Dev. Cell., 12 (2007), 664666 CrossRefGoogle ScholarPubMed
Larue, L., Dougherty, N., Porter, S. Mintz, B.. Spontaneous malignant transformation of melanocytes explanted from Wf/Wf mice with a Kit kinase-domain mutation . Proc. Natl. Acad. Sci. USA., 89 (1992), 78167820 CrossRefGoogle ScholarPubMed
Larue, L., Kumasaka, M. Goding, C.R.. Beta-catenin in the melanocyte lineage . Pigment Cell R., 16 (2003), 312317 CrossRefGoogle ScholarPubMed
Lee, Y., McIntire, L.V. Zygourakis, K.. Analysis of endothelial cell locomotion: Differential effects of motility and contact inhibition . Biotechnol Bioeng. 43 (1994), 622634 CrossRefGoogle ScholarPubMed
Mayer, T.C.. The migratory pathway of neural crest cells into the skin of mouse embryos . Dev. Biol., 34 (1973), 3946 CrossRefGoogle Scholar
McCarthy, J. Turley, E.A.. Effects of Extracellular Matrix Components on Cell Locomotion . Critical Reviews in Oral Biology and Medecine., 4 (1993), 619637 CrossRefGoogle ScholarPubMed
Mooney, D.J., Langer, R. Ingber, D.E.. Cytoskeletal filament assembly and the control of cell spreading and function by extracellular matrix . J. Cell Sci., 108 (1995), 2311-2320 Google ScholarPubMed
Palecek, S.P., Loftus, J.C., Ginsberg, M.H., Lauffenburger, D.A. Horwitz, A.F.. Integrin-ligand binding properties govern cell migration speed through cell-substratum adhesiveness . Nature, 385 (1997), 537540 CrossRefGoogle ScholarPubMed
Ridley, A.J., Schwartz, M.A., Burridge, K., Firtel, R.A., Ginsberg, M.H., Borisy, G., Parsons, J.T. Horwitz, A.R.. Cell migration: integrating signals from front to back . Science., 302 (2003), 17041709 CrossRefGoogle ScholarPubMed
Rosello, C., Ballet, P., Planus, E. Tracqui, P.. Model driven quantification of individual and collective cell migration . Acta Biotheoretica, 52 (2004), 343-363 CrossRefGoogle ScholarPubMed
Scott, G., Leopardi, S., Printup, S. Madden, B.C.. Filopodia are conduits for melanosome transfer to keratinocytes . J. Cell Sci., 115 (2002), 14411451 Google ScholarPubMed
Stéphanou, A., Mylona, E., Chaplain, M. Tracqui, P.. A computational model of cell migration coupling the growth of focal adhesions with oscillatory cell protusions . Journal of Theor. Biol., 253 (2008), 701716 CrossRefGoogle Scholar
Tawk, M., Araya, C., Lyons, D.A., Reugels, A.M., Girdler, G.C., Bayley, P.R., Hyde, D.R., Tada, M. Clarke, J.D.. A mirror-symmetric cell division that orchestrates neuroepithelial morphogenesis . Nature, 446 (2007), No. 7137, 797-800 CrossRefGoogle ScholarPubMed
Théry, M., Racine, V., Pépin, A., Piel, M., Chen, Y., Sibarita, J.-B. Bornens, M.. The extracellular matrix guides the orientation of the cell division axis . Nature Cell Biology, 7 (2005), No. 10, 947953 CrossRefGoogle ScholarPubMed
Wilkie, A.L., Jordan, S.A. Jackson, I.J.. Neural crest progenitors of the melanocyte lineage: coat colour patterns revisited . Development, 129 (2002), 3349-57 Google ScholarPubMed