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Biologically Active Nanofibers – Paralyzed Limbs Move Again: November 26, 2008

Researchers have developed a new material that enables mice with severe spinal cord injuries to regain movement in their paralyzed limbs within weeks. In combination with existing technologies, this new approach has the potential to considerably improve functional recovery in people following spinal cord injury.

(a) Schematic representation of self-assembling nanofibers with biologically active molecules on their surface. Nanofibers measure 6 to 8 nanometers in diameter – approximately 6,000 times thinner than a human hair. (b) Bundles of individual nanofibers forming a network (magnification bar = 200 nanometers) seen through a microscope. These networks assemble spontaneously when the molecules are injected into the site of spinal cord injury, creating therapy for damaged nerve extensions.

Laminin, a protein found in connective tissue and space between cells, encourages nerve cells to grow new extensions during development or after injury. Scientists identified a string of five amino acids (IKVAV) in laminin that binds to cell surface proteins known as integrin receptors, which relay growth signals into the cell. Samuel Stupp, Board of Trustees Professor of Materials Science, Chemistry, and Medicine at Northwestern University, is working with colleagues to harness the potential of this biological repair mechanism. Stupp’s team engineered a biologically active material, termed IKVAV peptide amphiphile (IKVAV PA), that organizes into nanostructures with an extremely high concentration of the IKVAV peptides on their surfaces. When exposed to salts in the body, IKVAV PA self-assembles into cylinder (tube)-shaped nanofibers. If the fibers are sufficiently concentrated, they form gels.

Nanofibers Enable Recovery after Spinal Cord Injury

Previously, Stupp’s team demonstrated that in a test tube IKVAV PA nanofibers stimulated outgrowth of nerve cell extensions. This suggests that injection of bioactive material might promote recovery of damaged nerves in the body. Stupp’s recent study with colleague John Kessler at Northwestern University, supported in part by the National Institute of Biomedical Imaging and Bioengineering (NIBIB) and the National Institute of Neurological Disorders and Stroke, examined whether IKVAV PA could assist in recovery of nerve function after spinal cord injury. Mice received an injection of IKVAV PA 24 hours after severe spinal cord injury that left their hind limbs paralyzed. The timing was chosen to mimic the delay of medical intervention people might experience after a serious injury. “Patients have to get to the hospital or might have to be stabilized before intervention,” says Stupp. Nine weeks after spinal cord injury, some of the mice treated with the nanofibers began using their hind legs again. These astounding results suggested that IKVAV PA had profound biological effects.

IKVAV PA – Modes of Action

The bioactive scaffold improved growth of nerve extensions (axons) up and down the spinal cord, leading to recovery of sensation and movement. At 11 weeks after injury, new nerve fibers grew through the injury site and connected with the spinal cord. This long-term effect surprised researchers because it is known that the scaffold is biodegraded within a couple of weeks.

It was also noted that, in addition to promoting survival of cells that help repair damaged nerves (oligodendrocytes) and directly enhancing nerve extension growth, IKVAV PA suppressed growth of scar-producing cells (astrocytes), thereby reducing scar formation. Scar tissue that forms after injury to or surgery on the spinal cord impedes nerve regeneration. The nanofiber therapy would be most helpful if administered soon after injury so it can diminish scar tissue formation and rescue damaged nerves while they are still alive.

Hope for the Paralyzed

In a test tube, the bioactive nanofibers encouraged neural progenitor cells, the immature predecessors of cells in the central nervous system, to differentiate into neurons. If IKVAV PA can do the same in the human body, there is hope for recovery from chronic nerve injury, such as long-term paralysis. By stimulating these progenitor cells, even dead neurons could potentially be replaced. “It could be the same therapy, but a different strategy. After injury, some axons may die completely, and some may not be making the same connections. If you have a therapy that can somehow improve axonal regeneration, it would be worthwhile to do surgery that removes the scar and then apply therapy,” explains Stupp. The treatment could be applied to treat both central (brain) or peripheral (limbs) nervous system injuries.

Unique Features of IKVAV PA

Self-assembly into nanofibers is a unique feature of IKVAV PA. Injection of IKVAV or laminin without the PA scaffold did not produce the same recovery benefit as IKVAV PA. Importantly, scar reduction occurred only with IKVAV PA, suggesting that the nanostructure of the scaffold itself plays a biological role, possibly by exposing cells to a high density of IKVAV molecules.

Applications and Future Directions

Previously, polymer scaffolds were studied for tissue regeneration as a way to mechanically support cells, but those scaffolds were not bioactive. Stupp and his team are among the pioneers in the field of bioactive nanomaterials for tissue regeneration. Although at this stage treatment with IKVAV PA enables only partial recovery of function after spinal cord injury, improving the stability and mechanical properties of the scaffold could significantly enhance its performance. “Our relatively recent work, also supported by NIBIB, has shown that we can control the mechanical properties of the scaffold by changing sequences in the domains where there isn’t supposed to be any bioactivity. The IKVAV is the bioactive domain, but there is also another domain, which is used to place amino acids to promote formation of cylindrical fibers. I would say that, from a bioengineering perspective, that was a very critical discovery that we made,” explains Stupp.

In the future, researchers will be able to design scaffolds with various bioactive molecules on their surface, instructing cells to behave in a desired way. “We have PAs that are of interest in the formation of blood vessel, bone, and cartilage, as well as in wound healing and Parkinson’s disease. In addition, we have a project on PAs that are cytotoxic and that kill cancer cells,” says Stupp.

Thus, although it might not yet represent a miraculous cure for paralysis, the biologically active nanofiber technology could become an important tool in the treatment of a broad range of medical conditions.

This work is supported in part by the National Institute of Biomedical Imaging and Bioengineering.

References

Huang Z, Sargeant T, Hulvat JF, Mata A, Bringas Jr P, Koh C-Y, Stupp SI, and Snead ML. Bioactive nanofibers instruct cells to proliferate and differentiate during enamel regeneration. Journal of Bone and Mineral Research. In press.

Tysseling-Mattiace VM, Sahni V, Niece KL, Birch D, Czeisler C, Fehlings MG, Stupp SI, Kessler JA. Self-assembling nanofibers inhibit glial scar formation and promote axon elongation after spinal cord injury. J Neurosci. 2008 Apr 2;28(14):3814–23.

Capito RM, Azevedo HS, Velichko YS, Mata A, Stupp SI. Self-assembly of large and small molecules into hierarchically ordered sacs and membranes. Science. 2008 Mar 28;319(5871):1812–6.

Samuel Stupp