Extensive Neuronal Differentiation of Human Neural Stem Cell Grafts in Adult Rat Spinal Cord

Abstract

Background

Effective treatments for degenerative and traumatic diseases of the nervous system are not currently available. The support or replacement of injured neurons with neural grafts, already an established approach in experimental therapeutics, has been recently invigorated with the addition of neural and embryonic stem-derived precursors as inexhaustible, self-propagating alternatives to fetal tissues. The adult spinal cord, i.e., the site of common devastating injuries and motor neuron disease, has been an especially challenging target for stem cell therapies. In most cases, neural stem cell (NSC) transplants have shown either poor differentiation or a preferential choice of glial lineages.

Methods and Findings

In the present investigation, we grafted NSCs from human fetal spinal cord grown in monolayer into the lumbar cord of normal or injured adult nude rats and observed large-scale differentiation of these cells into neurons that formed axons and synapses and established extensive contacts with host motor neurons. Spinal cord microenvironment appeared to influence fate choice, with centrally located cells taking on a predominant neuronal path, and cells located under the pia membrane persisting as NSCs or presenting with astrocytic phenotypes. Slightly fewer than one-tenth of grafted neurons differentiated into oligodendrocytes. The presence of lesions increased the frequency of astrocytic phenotypes in the white matter.

Conclusions

NSC grafts can show substantial neuronal differentiation in the normal and injured adult spinal cord with good potential of integration into host neural circuits. In view of recent similar findings from other laboratories, the extent of neuronal differentiation observed here disputes the notion of a spinal cord that is constitutively unfavorable to neuronal repair. Restoration of spinal cord circuitry in traumatic and degenerative diseases may be more realistic than previously thought, although major challenges remain, especially with respect to the establishment of neuromuscular connections.

Abstract

When neural stem cells from human fetal spinal cord were grafted into the lumbar cord of normal or injured adult nude rats, substantial neuronal differentiation was found.

Editors' Summary

Background.

Every year, spinal cord injuries, many caused by road traffic accidents, paralyze about 11,000 people in the US. This paralysis occurs because the spinal cord is the main communication highway between the body and the brain. Information from the skin and other sensory organs is transmitted to the brain along the spinal cord by bundles of neurons, nervous system cells that transmit and receive messages. The brain then sends information back down the spinal cord to control movement, breathing, and other bodily functions. The bones of the spine normally protect the spinal cord but, if these are broken or dislocated, the spinal cord can be cut or compressed, which interrupts the information flow. Damage near the top of the spinal cord can paralyze the arms and legs (tetraplegia); damage lower down paralyzes the legs only (paraplegia). Spinal cord injuries also cause many other medical problems, including the loss of bowel and bladder control. Although the deleterious effects of spinal cord injuries can be minimized by quickly immobilizing the patient and using drugs to reduce inflammation, the damaged nerve fibers never regrow. Consequently, spinal cord injury is permanent.

Why Was This Study Done?

Scientists are currently searching for ways to reverse spinal cord damage. One potential approach is to replace the damaged neurons using neural stem cells (NSCs). These cells, which can be isolated from embryos and from some areas of the adult nervous system, are able to develop into all the specialized cells types of the nervous system. However, because most attempts to repair spinal cord damage with NSC transplants have been unsuccessful, many scientists believe that the environment of the spinal cord is unsuitable for nerve regeneration. In this study, the researchers have investigated what happens to NSCs derived from the spinal cord of a human fetus after transplantation into the spinal cord of adult rats.

What Did the Researchers Do and Find?

The researchers injected human NSCs that they had grown in dishes into the spinal cord of intact nude rats (animals that lack a functioning immune system and so do not destroy human cells) and into nude rats whose spinal cord had been damaged at the transplantation site. The survival and fate of the transplanted cells was assessed by staining thin slices of spinal cord with an antibody that binds to a human-specific protein and with antibodies that recognize proteins specific to NSCs, neurons, or other nervous system cells. The researchers report that the human cells survived well in the adult spinal cord of the injured and normal rats and migrated into the gray matter of the spinal cord (which contains neuronal cell bodies) and into the white matter (which contains the long extensions of nerve cells that carry nerve impulses). 75% and 60% of the human cells in the gray and white matter, respectively, contained a neuron-specific protein six months after transplantation but only 10% of those in the membrane surrounding the spinal cord became neurons; the rest developed into astrocytes (another nervous system cell type) or remained as stem cells. Finally, many of the human-derived neurons made the neurotransmitter GABA (one of the chemicals that transfers messages between neurons) and made contacts with host spinal cord neurons.

What Do These Findings Mean?

These findings suggest that human NSC grafts can, after all, develop into neurons (predominantly GABA-producing neurons) in normal and injured adult spinal cord and integrate into the existing spinal cord if the conditions are right. Although these animal experiments suggest that NSC transplants might help people with spinal injuries, they have some important limitations. For example, the spinal cord lesions used here are mild and unlike those seen in human patients. This and the use of nude rats might have reduced the scarring in the damaged spinal cord that is often a major barrier to nerve regeneration. Furthermore, the researchers did not test whether NSC transplants provide functional improvements after spinal cord injury. However, since other researchers have also recently reported that NSCs can grow and develop into neurons in injured adult spinal cord, these new results further strengthen hopes it might eventually be possible to use human NSCs to repair damaged spinal cords.

Additional Information.

Please access these Web sites via the online version of this summary at http://dx.doi.org/doi:10.1371/journal.pmed.0040039.

• The US National Institute of Neurological Disorders and Stroke provides information on spinal cord injury and current spinal cord research

• Spinal Research (a UK charity) offers information on spinal cord injury and repair

• The US National Spinal Cord Injury Association Web site contains factsheets on spinal cord injuries

• MedlinePlus encyclopedia has pages on spinal cord trauma and interactive tutorials on spinal cord injury

• The International Society for Stem Cell Research offers information on all sorts of stem cells including NSCs

• The US National Human Neural Stem Cell Resource provides information on human NSCs, including the current US government's stance on stem cell research