Dr. Remigio Picone


EPSRC PhD Plus 2010 Project: The Development of a Novel Molecular Patterning Technique for Nerve Tissues Regeneration



Background

The cell is the basic unit of life. Understanding how cells organise themselves, and in turn, how they self- assemble into healthy functional tissues, has profound implications for our understanding of disease and the development of drugs and tools for tissue regeneration. Conventional cell biology and micro/nano-fabrication tools and techniques have provided powerful insights into the understanding of these mechanisms (1, 2). The main aim of my project is to develop novel two- and three-dimensional micropatterning techniques. To explore its potential for biomedical applications, I will fabricate a novel device to guide regeneration of nerve tissues following severe injuries. This highly interdisciplinary proposal builds on my recent findings and may lead to a new approach to treat nerve injuries.


Over the past 10 years, micro/nano-fabrication techniques, have made substantial contributions to the understanding of fundamental cell biology, stem cell research and tissue engineering (1-6). Molecular micropatterning(7), which consists of positioning of molecules in geometrical patches onto different type of surfaces has been especially useful. However, techniques based upon conventional micropattering are limited to 2D and present many problems when applied to biological problems(8). After encountering many of these problems during the course of my PhD, I pioneered the development of a novel micropatterning method,  which I successfully applied during recent pilot experiments in a previous work(5). The technique has three main steps: pre-patterning surface coating; patterning by ablation; and post-patterning surface coating. Important advantages over existing micropatterning techniques are:

  1. highly uniform and reproducible patterning in 2D and 3D (Figure A)
  2. control over molecule-surface binding strength
  3. patterning of different type of molecules onto the same surface
  4. preservation of the surface bulk material
  5. cost effectiveness
  6. shorter processing times

These features open up exciting new possibilities for the generation of very large surface patterned areas on biomaterials for biomedical applications.

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Project

The peripheral nervous system has remarkable regenerative potential and Schwann cells play a key role in this regeneration. The Schwann cells provide a physical “conduit” to direct and stimulate axonal regrowth, but cannot heal large gaps produced by severe nerve injuries. Recently, Alison Lloyd and collaborators at University College London (UCL) found a new extracellular signalling molecule (Ephrin-B) responsible for driving Schwann cells directional growing (9) (Figure A in vitro). This result potentially offers an important application for the regeneration of nerve tissues following severe injuries, which are normally impeded by the large distance between the damaged nerve ends.

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By patterning stripes of Ephrin-B inside a silicon tubular 3D structure (Figure B), it may be possible to connect together the two cut nerve ends and promote guidance of the Schwann cells from one end to the other, thereby promoting the entire axonal nerve regeneration. By applying the above micropatterning technique I will pattern 2D surfaces (Figure A) and fabricate and pattern the 3D structure (Figure A). This will allow me to test technique reproducibility, stability (the pattern must be stable for days) on different 2D surfaces and 3D large structures. The test, in well established cellular (in vitro, Figure B) and animal systems (in vivo, Figure B), will be conducted in the laboratory of Alison Lloyd at MRC-Laboratory for Molecular Cell Biology (LMCB) at UCL.


References

  1. R. Singhvi, A. Kumar, G. P. Lopez, G. N. Stephanopoulos, D. I. Wanget al., Science 264, 696 (1994).
  2. L. G. Griffith, G. Naughton, Science 295, 1009 (2002).
  3. W. Wang, K. Itaka, S. Ohba, N. Nishiyama, U. Chunget al., Biomaterials 30, 2705 (2009).
  4. C. S. Chen, M. Mrksich, S. Huang, G. M. Whitesides, D. E. Ingber, Science 276, 1425 (1997).
  5. Picone R, Ren X, Clarke J D W, McKendry R. A, Buzz Baum (2010). A Polarised Population of Dynamic Microtubules Mediates Homeostatic Length Control in Animal Cells. Plos Biology 8: e1000542. Highlighted in “Faculty of 1000"
  6. C. S. Chen, M. Mrksich, S. Huang, G. M. Whitesides, D. E. Ingber, Biotechnol Prog 14, 356 (1998).
  7. R. S. Kane, S. Takayama, E. Ostuni, D. E. Ingber, G. M. Whitesides, Biomaterials 20, 2363 (1999).
  8. J. Fink, M. Théry, A. Azioune, R. Dupont, F. Chatelainet al., Lab Chip 7, 672 (2007).
  9. Parrinello, S., Napoli, I., Ribeiro, S., Digby, P. W. W., Fedorova, M., Parkinson, D. B., Doddrell, R. D., Nakayama, M., Adams, R. H., Lloyd, A. C. (2010). EphB signaling directs peripheral nerve regeneration through Sox2-dependent Schwann cell sorting. Cell 143, 145–155


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