Three-dimensional tissue culture based on magnetic cell levitation

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Abstract

Cell culture is an essential tool in drug discovery, tissue engineering and stem cell research. Conventional tissue culture produces two-dimensional cell growth with gene expression, signalling and morphology that can be different from those found in vivo, and this compromises its clinical relevance1,2,3,4,5. Here, we report a three-dimensional tissue culture based on magnetic levitation of cells in the presence of a hydrogel consisting of gold, magnetic iron oxide nanoparticles and filamentous bacteriophage. By spatially controlling the magnetic field, the geometry of the cell mass can be manipulated, and multicellular clustering of different cell types in co-culture can be achieved. Magnetically levitated human glioblastoma cells showed similar protein expression profiles to those observed in human tumour xenografts. Taken together, these results indicate that levitated three-dimensional culture with magnetized phage-based hydrogels more closely recapitulates in vivo protein expression and may be more feasible for long-term multicellular studies.

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Figure 1: Magnetic iron oxide-containing hydrogels.
Figure 2: Three-dimensional cell culture with magnetic-based levitation.
Figure 3: Comparison of three-dimensional cell growth with standard two-dimensional tissue culture.
Figure 4: Shape control of magnetically levitated culture.
Figure 5: Confrontation assay of magnetically levitated multicellular spheroids.

References

  1. 1

    Cukierman, E., Pankov, R., Stevens, D. R. & Yamada, K. M. Taking cell-matrix adhesions to the third dimension. Science 294, 1708–1712 (2001).

  2. 2

    Abbott, A. Biology's new dimension. Nature 424, 870–872 (2003).

  3. 3

    Pampaloni, F., Reynaud, E. G. & Stelzer, E. H. The third dimension bridges the gap between cell culture and live tissue. Nature Rev. Mol. Cell Biol. 8, 839–845 (2007).

  4. 4

    Griffith, L. G. & Swartz, M. A. Capturing complex 3D tissue physiology in vitro. Nature Rev. Mol. Cell Biol. 7, 211–224 (2006).

  5. 5

    Atala, A. Engineering tissues, organs and cells. J. Tissue Eng. Regen. Med. 1, 83–96 (2007).

  6. 6

    Coleman, C. B. et al. Diamagnetic levitation changes growth, cell cycle and gene expression of Saccharomyces cerevisiae. Biotechnol. Bioeng. 98, 854–863 (2007).

  7. 7

    Dobson, J. Remote control of cellular behaviour with magnetic nanoparticles. Nature Nanotech. 3, 139–143 (2008).

  8. 8

    Ito, A., Shinkai, M., Honda, H. & Kobayashi, T. Medical application of functionalized magnetic nanoparticles. J. Biosci. Bioeng. 100, 1–11 (2005).

  9. 9

    Souza, G. R. et al. Networks of gold nanoparticles and bacteriophage as biological sensors and cell targeting agents. Proc. Natl Acad. Sci. USA 103, 1215–1220 (2006).

  10. 10

    Souza, G. R. et al. Bottom-up assembly of hydrogels from bacteriophage and Au nanoparticles: the effect of cis- and trans-acting factors. PLoS ONE 3, e2242 (2008).

  11. 11

    Petersen, O. W., Ronnov-Jessen, L., Howlett, A. R. & Bissell, M. J. Interaction with basement membrane serves to rapidly distinguish growth and differentiation pattern of normal and malignant human breast epithelial cells. Proc. Natl Acad. Sci. USA 89, 9064–9068 (1992).

  12. 12

    Mikos, A. G. et al. Engineering complex tissues. Tissue Eng. 12, 3307–3339 (2006).

  13. 13

    Ito, A., Ino, K., Kobayashi, T. & Honda, H. The effect of RGD peptide-conjugated magnetite cationic liposomes on cell growth and cell sheet harvesting. Biomaterials 26, 6185–6193 (2005).

  14. 14

    Pankhurst, Q., Connolly, J., Jones, S. K. & Dobson, J. Applications of magnetic nanoparticles in biomedicine. J. Phys. D 36, R167–R181 (2003).

  15. 15

    Alsberg, E., Feinstein, E., Joy, M. P., Prentiss, M. & Ingber, D. E. Magnetically-guided self-assembly of fibrin matrices with ordered nano-scale structure for tissue engineering. Tissue Eng. 12, 3247–3256 (2006).

  16. 16

    Dobson, J., Cartmell, S. H., Keramane, A. & El Haj, A. J. Principles and design of a novel magnetic force mechanical conditioning bioreactor for tissue engineering, stem cell conditioning and dynamic in vitro screening. IEEE Trans. Nanobiosci. 5, 173–177 (2006).

  17. 17

    Meyer, C. J. et al. Mechanical control of cyclic AMP signaling and gene transcription through integrins. Nature Cell Biol. 2, 666–668 (2000).

  18. 18

    Matthews, B. D., La Van, D. A., Overby, D. R., Karavitis, J. & Ingber, D. E. Electromagnetic needles with submicron pole tip radii for nanomanipulation for biomolecules and living cells. Appl. Phys. Lett. 85, 2968–2970 (2004).

  19. 19

    Arap, W., Pasqualini, R. & Ruoslahti, E. Cancer treatment by targeted drug delivery to tumor vasculature in a mouse model. Science 279, 377–380 (1998).

  20. 20

    Hajitou, A. et al. A hybrid vector for ligand-directed tumor targeting and molecular imaging. Cell 125, 385–398 (2006).

  21. 21

    Nam, K. T. et al. Virus-enabled synthesis and assembly of nanowires for lithium ion battery electrodes. Science 312, 885–888 (2006).

  22. 22

    Langer, R. & Tirrell, D. A. Designing materials for biology and medicine. Nature 428, 487–492 (2004).

  23. 23

    Hautot, D. et al. Preliminary observation of elevated levels of nanocrystalling iron oxide in the basal ganglia of neuroferritinopathy patients. Biochim. Biophys. Acta 1772, 21–25 (2007).

  24. 24

    Snyder, E. Y. et al. Multipotent neural cell lines can engraft & participate in development of mouse cerebellum. Cell 68, 33–51 (1992).

  25. 25

    Wan, X., Li, Z. & Lubkin, S. R. Mechanics of mesenchymal contribution to clefting force in branching morphogenesis. Biomech. Model. Mechanobiol. 7, 417–426 (2008).

  26. 26

    Csikasz-Nagy, A., Battogtokh, D., Chen, K. C., Novak, B. & Tyson, J. J. Analysis of a generic model of eukaryotic cell-cycle regulation. Biophys. J. 90, 4361–4379 (2006).

  27. 27

    Ofek, G. et al. Matrix development in self-assembly of articular cartilage. PLoS ONE 3, e2795 (2008).

  28. 28

    Bhowmick, D. A., Zhuang, Z., Wait, S. D. & Weil, R. J. A functional polymorphism in the EGF gene is found with increased frequency in glioblastoma multiforme patients and is associated with more aggressive disease. Cancer Res. 64, 1220–1223 (2004).

  29. 29

    Chicoine, M. R. & Silbergeld, D. L. Mitogens as motogens. J. Neurooncol. 35, 249–257 (1997).

  30. 30

    Georgescu, M. M., Kirsch, K. H., Akagi, T., Shishido, T. & Hanafusa, H. The tumor-suppressor activity of PTEN is regulated by its carboxyl-terminal region. Proc. Natl Acad. Sci. USA 96, 10182–10187 (1999).

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Acknowledgements

The authors would like to thank R.R. Brentani, N.R. Pellis and E.H. Sage for helpful discussions and K. Dunner Jr and D. Bier for technical assistance. G.R.S. was supported by the Odyssey Scholar Program of the University of Texas M.D. Anderson Cancer Center and by the Breast Cancer Research Program of the US Department of the Defense (DOD). D.J.S. received support from the National Science Foundation. T.C.K. received support from the David and Lucille Packard Foundation. W.A. and R.P. received support from the Gillson-Longenbaugh Foundation, the Marcus Foundation, AngelWorks, DOD, National Institutes of Health (NIH) and National Cancer Institute.

Author information

G.R.S., J.R.M., R.M.R., D.J.S., C.S.L, J.M., T.C.K., W.A. and R.P. conceived and designed the experiments. G.R.S., J.R.M., T.C.K., D.J.S., C.S.L., J.S.A. and J.M. performed the experiments. G.R.S., M.G.O., D.J.S., C.S.L., L.F.B., J.S.A., J.A.B., T.C.K., W.A. and R.P. analysed the data. R.M.R., M.M.G., J.A.B., J.G.G., T.C.K., W.A. and R.P. contributed materials and analysis tools. G.R.S., R.M.R., M.G.O., L.F.B., J.A.B., J.G.G., T.C.K., W.A. and R.P. co-wrote the paper.

Correspondence to T. C. Killian or Wadih Arap or Renata Pasqualini.

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Competing interests

The University of Texas M. D. Anderson Cancer Center (UTMDACC) and Rice University (RU), along with their researchers, have filed patents on the technology and intellectual property reported here. If licensing or commercialization occurs, the researchers are entitled to standard royalties. G.R.S., R.M.R., C.S.L. and T.C.K. have equity in Nano3D Biosciences, Inc. UTMDACC and RU manage the terms of these arrangements in accordance to their established institutional conflict-of-interest policies.

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