29 Jul 2005

The allure of stem cells lies in their potential to treat a plethora of human degenerative diseases, including neurologic disorders such as Alzheimer and Parkinson diseases. However, realizing this potential requires taming the little dynamos by gently but firmly steering them down desired paths of functional differentiation. Two papers appearing this week present new solutions to this delicate problem. In the first, Scott Whittemore and colleagues at the University of Louisville in Kentucky describe how forced expression of a neurotrophin enhances the differentiation and therapeutic effectiveness of embryonic glial precursor cells transplanted into rats after spinal cord injury. The second, from Michal Stachowiak and coworkers at the State University of New York, Buffalo, reports a somewhat different route, showing that injection of DNA-coated nanoparticles into mouse brain can efficiently transfect progenitors with a receptor gene that slows their proliferation. Because the nanoparticle approach can be used to deliver DNA to mature neurons, it also has potential for neuronal gene therapy.

The use of glial progenitors and neural stem cells in animal models shows the potential of stem cells to treat spinal cord injury. That glial precursor transplants can restore at least some function after such trauma is partially a result of their ability to differentiate into oligodendrocytes, which promote remyelination of damaged nerve fibers. In Whittemore’s study, which appeared in the July 27 issue of the Journal of Neuroscience, first author Qilin Cao showed that the therapeutic effects of glial progenitor cells could be boosted by transducing the cells with the gene for the neurotrophin, D15A, before transplantation. D15A displays both brain-derived neurotrophic factor (BDNF) and neurotrophin 3 (NT3)-like activities—both important for myelination during development and for axon regeneration after injury. The authors found that adding D15A increased the survival of the transplanted precursor cells, elevated their differentiation into mature oligodendrocytes, and enhanced remyelination of axons in the area of the injury.

This repair was good enough to restore nerve conductance across the injury site in eight out of 12 rats. Although the amplitude of the conductance was only a tenth of that seen in uninjured rats, no restoration was seen in any of the animals grafted with precursor cells only, or with D15A-expressing fibroblasts. Hind limb function was also improved by the souped-up precursors compared to rats transplanted with plain precursor cells, leading the authors to conclude that combined treatment of progenitor cells plus neurotrophic support could be a useful strategy for rescuing injured spinal cord. The approach might also prove useful for tackling diseases where myelin has degenerated, such as multiple sclerosis and Alzheimer disease.

An alternative to stem cell transplants would be to mobilize progenitors already present in the spinal cord or brain. In theory, this could be done by genetically modifying the cells, but this has proven difficult, not least because the viral vectors often used to deliver the genetic material can be inefficient, or worse, toxic. But the paper by Stachowiak and colleagues in this week’s PNAS Early Edition online suggests that nanoparticles can get around these major problems. First author Dhruba Bharali and colleagues injected modified silica nanoparticles, loaded with DNA coding for enhanced green fluorescent protein (EGFP), into the ventricular space of mouse brains. When they examined the brain tissue about 7 days later, they found EGFP was expressed in surrounding structures, including the striatum, motor cortex, and hippocampus. Using a novel confocal fiber optic probe, they detected fluorescent cells surrounding the ventricle in live animals, including in the subventricular zone, a region rich in neuronal progenitor cells.

To try to encourage progenitor cell differentiation in vivo, the researchers coated nanoparticles with DNA coding for a nuclear-targeted fibroblast growth factor receptor 1 (FGFR1) and delivered it to the lateral ventricle—it was previously demonstrated that nuclear accumulation of FGFR1 prompts precursor cells to exit the cell cycle and begin differentiation. Ten days later, expression of FGFR1 in the region was accompanied by lower incorporation of bromodeoxyuridine, indicating that cell proliferation had been slowed. No toxicity was seen from the procedure, even after two sequential intraventicular injections.

Bharali and colleagues also showed that the nanoparticles could also be used to deliver DNA into brain tissues; when the EGFP construct was injected into the substantia nigra, robust expression was widespread in dopaminergic neurons. The new nanoparticles could be useful for investigating Parkinson disease, determining the signals controlling stem cell proliferation, and possibly even delivering gene therapies.—Pat McCaffrey

Comments

1. • Mahendra Rao
     Life Technologies Corporation
   • Posted: 29 Jul 2005

   The authors of this paper make an important point in a very nice and elegant way. Glial progenitors require an additional signal to complete the process of myelination. They showed that a neurotrophin signal is what may be missing, as making a modified progenitor that expresses a neurotrophin capable of binding to multiple receptors can enhance myelination and functional repair.

   View all comments by Mahendra Rao
2. • Ronald Klein
     
   • Posted: 05 Aug 2005

   The nanoparticle gene transfer described by Bharali et al. is an exciting new avenue for genetic modification in the brain. The study suggests the organically modified nanoparticles are easy to make, are safe, and provide efficient and widespread transgene expression. The spread of expression after ventricular injections and the live imaging technique were impressive. I would have liked to see if the nanoparticles are more efficient or less toxic than other nonviral systems. The current nanoparticle preparations are probably not as effective for gene transfer as the most efficient viral vectors—as GFP fluorescence was rather weak—but further improvements on the new technology are to be expected. One potential advantage of nonviral techniques is for efficient glial expression, as most viral vectors have a strong tropism for neurons, although glial expression was not demonstrated in this study.

   View all comments by Ronald Klein
3. • Jacob Mack
     bio-chemistry-psychology/neuroscience graduate.
   • Posted: 11 Aug 2005

   I think the new applications utilizing D15A looks very promising, but the nanotechnolgy has more broad-based, long-term implications. Showing no toxicity is an amazing leap forward as usually nanotech experiments show similar effects to silicosis. Perhaps a combination of both treatments will show the most therapeutic promise.

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