Core Laboratories of the Reeve Foundation International Research Consortium on Spinal Cord Injury

The core laboratories of the Reeve Foundation International Research Consortium on Spinal Cord Injury were set up to create centralized resources that Consortium members could share, in order to improve the efficiency and the quality of the work of the Consortium. In addition, Reeve Foundation recognized that these laboratories could become invaluable training centers for young scientists.

The goals for the core laboratories are as follows:

• To facilitate collaborative experiments that involve some or all Consortium laboratories and alleviate the burden on any one laboratory to provide the personnel, resources, and space.
• To provide technical and analytical assistance to member laboratories.
• To train Consortium Associates and support their transition to independent investigators by providing them with continued access to core expertise and activities even after they leave.
• To ensure continuity within the Consortium and the integrity of collaborations and projects, particularly as some Associates cycle out and new Associates join.
• To enable the Consortium to undertake studies that might otherwise be too expensive for any individual laboratory to tackle because of the need for new equipment, staff, or other resources. This is particularly true when animal models are involved.
• To ensure standardization across Consortium laboratories.
• To perfect experimental techniques and assessment tools and to develop other resources that will improve the work of the Consortium and enrich the field of spinal cord research.

Read more about the Reeve Foundation International Research Consortium on Spinal Cord Injury.

Microarray Core (Salk Institute for Biological Studies)

Microarray technology was a logical choice for the first core laboratory of the Reeve Foundation International Research Consortium on Spinal Cord Injury. This technology has emerged as an exciting and aggressive research tool that enables researchers to screen thousands of genes simultaneously to see which ones are active, or expressed, and which ones were silent. Genes are arrayed on a microchip the size of a fingernail, and experiments that once took years to complete can be done in a matter of weeks.

The hope was - and continues to be - that by observing the patterns of gene expression to see how they changed after a spinal cord injury, scientists might identify therapeutic targets. For example, treatments might be devised to enhance the influence of beneficial genes or minimize the power of those that prevent the spinal axons from regenerating. Researchers also hoped to learn how gene activity changes during the development of the brain and spinal cord, which might offer clues for how to restart that creative process to heal an injury. Finally, microarray technology offers a new way to evaluate potential treatments by assessing their impact on gene activity.

Because microarray equipment was expensive and required dedicated, highly trained personnel, it made sense to create a central microarray laboratory for the Consortium. Under the direction of Fred H. Gage, Ph. D., this core opened at the Salk Institute in September 2000. It was equipped with the Affymetrix GeneChip(tm) system, which provides an unbiased "snapshot" of differential gene expression in many animals, including the mouse and rat, as well as humans.  This approach enables biologists not only to test expected gene changes but also to look at unexpected, potentially important differences that would otherwise go undetected.

Since its inception, member laboratories have routinely accessed the Microarry Core for myriad and experiments. They have explored a variety of questions, ranging from how various treatments affect recovery from a spinal cord injury to how endogenous cell signaling may be interfering with regeneration. The use of a single, dedicated technician has insured that Reeve Foundation studies produced consistent data that is of the highest quality. In addition to its involvement in individual and inter-laboratory experiments, the Reeve Foundation Microarray Core played a central role in the latest Consortium-wide experiment.  That study characterized the changes in gene expression at the site of, as well as above and below, a moderate contusion injury in rats. The project involved 108 GeneChips and looked at four time points, spanning from three hours after injury to a more “chronic” state 35 days later. Analyzing the data produced a spatial and temporal profile of spinal cord injury and also identified several promising avenues for new clinical treatments. The study, “Spatial and temporal gene expression profiling of the contused rat spinal cord,” was published in ExpNeurol 189 (2004), pp204-221. (raw data and analyzed files available at http://genechip.salk.edu/genechip).

The Core also has improved the accuracy of its microarray experiments by consulting frequently with Affymetrix personnel and upgrading its equipment. The analysis of Affymetrix data is no trivial task, and Core personnel have continued to refine this process.  For instance, the laboratory developed a novel statistical method to isolate important gene changes, an accomplishment they described in the Journal of Neuroscience Methods (Aimone, J.F., Gage, F.H. 2004 135, 27-33).

The Injury Core (University of California, Irvine)

Any promising approach to treating spinal cord injuries must undergo thorough testing in animal models before it can move into human clinical trials. Yet spinal cord experiments on animals are technically challenging, labor intensive, and expensive. Among the hurdles are performing delicate, precise surgery on animals to produce a variety of standardized injury models; caring for the animals before, during, and after experiments; and producing objective measurements of the results. To make it easier, faster, and more economical for Consortium laboratories to move exciting bench research into animal experiments, Reeve Foundation set up the Injury Core in March 2001.

The Injury Core, under the direction of Aileen J. Anderson, PhD., has supported more than 60 Consortium projects. Equally important, it has been an invaluable training ground. It helped to prepare new Associates as they began their assignments in Consortium laboratories and supported those Associates who moved on to establish independent careers as spinal cord investigators. Educational activities for Associates have included workshops on these topics: locomotor training in humans and animal models; new animal models of spinal cord injury; use of the BMS locomotor rating scale for mouse models, and methods of GeneChip analysis. This support has been crucial in assuring that these young researchers will maintain a spinal cord focus. Moreover, with the new skills they have acquired, they are better equipped to complete the type of preliminary studies that will enable them to win larger, more long-term grants.

Recently, the scope of the Injury Core laboratory has expanded to include the development of resources and new knowledge for the spinal cord field. The Core has produced new behavioral measures to assess the effects of experimental treatments on animals. These tests will help to move successful animal studies into clinical trials. Also being evaluated in mouse models are a ladder beam test of functional recovery, in which mice walk over a horizontal ladder while researchers watch how many rungs the animals miss, and the use of Magnetic Resonance imaging (MRI) to determine the volume of spinal cord lesions. Moreover, in the last two years, the Core has been opened to investigators with individual CRF grants so they can test their research in animal models.  And in July 2005, Reeve Foundation launched a Core Pilot Data Program to give investigators the opportunity to participate in pilot studies of potential therapeutic interventions for spinal cord injury. The Core has also provided tissue samples, training, and other resources to other, non-CRF-funded scientists working in the spinal cord field.

The Vector Core (Salk Institute for Biological Studies)

Gene therapy is an exciting new approach that one day could limit the secondary wave of destruction from spinal cord injuries, prevent the formation of a cavity and scar tissue at the site of a lesion, and promote the repair of damaged spinal circuits. Doctors would deliver therapeutic genes via so-called vectors, viruses that have been genetically engineered so that they can infect cells without causing illnesses.  Instead, vectors deposit their genetic payloads into the nuclei of target cells, where the new genes begin to influence the production of proteins that, in turn, influence cell behavior. Scientists use different families of viral vectors, depending on the type of cell they are targeting.

Under the direction of Fred H. Gage, Ph.D., the Vector Core laboratory is designing and manufacturing new vectors for collaborative projects in the Consortium. These vectors are non-toxic, long lasting, and able to produce sufficient amounts of the therapeutic genes to create the desired biologic changes. These vectors can be used in a variety of animals and can infect both dividing and non-dividing cells. The core can create:

• New versions of two of the most useful) types of vectors: recombinant adeno-associated virus (rAAV) vectors and human immunodeficiency virus (HIV)-based lentiviral vectors.
• New forms of rAAV and lentiviral vectors that enable scientists to adjust how much genetic material they produced after having been inserted either into tissue cultures or animals.  This regulation would be accomplished by administering, say, an antibiotic either orally or by injection.
• Vectors to manipulate gene sequences in vivo.

Applications to Spinal Cord Injury

A major obstacle in treating injuries and diseases in the brain and spinal cord is that many possible therapeutic substances consist of large molecules that cannot pass through the blood-brain barrier. Viral vectors can execute a kind of end run around this obstacle.  That is, doctors would administer to patients viral vectors packed with genes that would spur the body to produce its own medicine precisely in the spinal cord where it is needed.  The Vector Core will collaborate with other Consortium laboratories to test therapeutic genes in the injured spinal cords of animal models.  These genes have the potential to:

• Protect against cavitation
• Limit the death of motor neurons
• Spur axonal elongation
• Maintain the structural and functional integrity of the damaged cord

The Gage laboratory is highly qualified to collaborate on these experiments. Researchers there recently discovered that when AAV vectors that express therapeutic genes are injected intramuscularly, they move efficiently from muscle to the motor neurons that link muscles with the spinal cord. The Gage team recently utilized this novel transport system to test two treatments - glial derived neurotrophic factor (GDNF) and insulin like growth factor-1 (IGF-1) - on animal models of Amyotrophic Lateral Sclerosis, also known as Lou Gehrig’s disease or ALS. Researchers found that IGF-1 prolonged life and delayed disease progression, even after overt disease symptoms were present.  In addition, IGF-1 significantly slowed cell death in this disease model. Based on the positive results from these experiments, clinical trials are being designed to test this exciting gene therapy in people with ALS. This proposal seeks to translate the basic scientific discoveries made in the Gage laboratory into a new treatment for spinal cord injury.

Behavior Core (University of California, Los Angeles)

Animal studies of potential treatments for spinal cord injuries require increasingly specialized tools to evaluate the animals’ behavior before and after the experiment. The days of relying on visual assessments of, say, how well a rat can balance on two legs, have given way to objective, quantifiable methods that exploit cutting-edge biomedical technologies. Access to these technologies in a Core facility enables Consortium members to obtain discriminating analyses of animal models under well-controlled conditions.

The major goals of this Core are to provide rapid, quantitative measurements of posture and locomotion in animal models of spinal cord injury and to compare the effectiveness of experimental treatments. Such measurements range from electrophysiological tests that determine how well synapses are working to robotic and video-based evaluations of the dynamics of joint movements during locomotion. These assessments can be combined with electromyographic studies that monitor electrical activity in muscle fibers in acute and chronic rodent models of spinal cord injuries.

Directed by V. Reggie Edgerton, Ph.D., the Behavior Core is developing new ways to study motor behavior in mice, rats, and primates and improve current assessment methods. As these techniques are refined, they will be made available to all Consortium laboratories. Also, studies that include training mice or rats to step or to stand can be conducted with any Consortium member. The use of robotic devices to train animals with spinal cord injuries to step or to stand is a critical element of spinal cord research. These sophisticated devices can control the specific motor task that an animal is mastering as well as quantify how much learning has occurred. Consortium laboratories are encouraged to send personnel to participate in studies and to learn techniques related to neuromotor physiology.

Members can utilize the Behavior Core in several ways.  Consortium laboratories can administer an experimental treatment to animal models and then send them to the Core for assessment and termination within a day or two.  Animals can receive an initial treatment in a Consortium laboratory and then be sent to UCLA for training. Finally, an animal study can be studied exclusively at this Core. Personnel from CRF laboratories provide most of the day-to-day care and treatment of the animals while learning the Core techniques to provide quantitative evaluations of motor behavior.