- Progenitor cells caught by time-lapse imaging
Study may offer insight into multiple sclerosis
A transgenic zebra fish expressing enhanced GFP has helped bring a broader understanding of how nerve cells obtain their myelin sheath, the membrane that acts as an electrical insulator. The research may eventually play a role in developing therapies to treat multiple sclerosis, a degenerative nerve disease characterized by erosions in the myelin sheath.
Investigators at Vanderbilt University in Nashville, Tenn., and at the University of Bath in the United Kingdom created the transgenic zebra fish to express fluorescent proteins in oligodendrocyte progenitor cells. During embryo development, these cells divide and migrate throughout the central nervous system and give rise to oligodendrocytes, which then grow and enwrap the axons to create the myelin sheath. According to team leader Bruce Appel, this process is essentially the same as in humans.
Oligodendrocyte progenitor cells develop in certain parts of the nervous system and migrate to their target axons. The number and distribution of the progenitor cells and oligodendrocytes must be matched to the number and distribution of target axons. The scientists demonstrated progenitor cell migration in an intact organism, and their results provide hints about the mechanisms that regulate migration and division, according to Appel.
Studying this process is important because the human nervous system can partially remyelinate after multiple sclerosis or other demyelinating disease or injury. According to Appel, adult humans have many oligodendrocyte progenitor cells and, following demyelination, those near the lesion divide, migrate into the lesion, and partially wrap and myelinate axons.
However, remyelination generally is incomplete. “Our premise is that mechanisms that regulate myelination during development also operate during remyelination. Consequently, understanding the former might facilitate design of therapies to promote remyelination,” he said.
The investigators conducted time-lapse imaging employing a Zeiss microscope equipped with a spinning disk confocal setup from PerkinElmer. They collected images starting at 36 hours post-fertilization and ending at 72 hours postfertilization when axon wrapping begins. The work is detailed in the December 2006 issue of Nature Neuroscience.
Using time-lapse photography and transgenic zebra fish labeled with enhanced GFP, researchers have determined that oligodendrocyte progenitor cells appear to explore their surroundings by extending filopodialike processes as they migrate and begin to enwrap axons. In these lateral views of the trunk region of the zebra fish spinal cord starting at 48 hours postfertilization, oligodendrocyte progenitor cells are marked by +, %, # and $. Arrows point to areas where processes from two neighboring cells have begun to enwrap axons. Anterior is toward the left, and ventral is toward the bottom.
The images highlighted the movement of the highly branched filopodialike processes that are extensions of the oligodendrocyte progenitor cell walls. The filopodia have been observed in other studies, but their role in myelination was unknown. The researchers determined that the filopodia appear to be actively exploring their environments by extending and retracting the members as they move. This exploration is “highly dynamic” during the development phase when the progenitor cells are migrating.
To explore this process further, they collected images at 2-min intervals to measure changes in the lengths of the filopodia. Filopodia extension and retraction ranged from approximately 1 to 13 μm every 2 min, averaging between 4.2 and 7.4 μm.
In addition, the cells could move up to 253 μm within the 36-hour imaging period. Interestingly, when oligodendrocyte progenitor cells met, they changed migratory paths. To further investigate, the scientists marked different oligodendrocyte progenitor cells with various colors of fluorescent protein and watched as the cells encountered each other in the developing embryo. As the cells met, they continued to remodel their filopodia and gradually withdrew from each other.
Because myelination is important for a functioning nervous system, and its destruction plays a key role in multiple sclerosis and other diseases, the researchers examined how the oligodendrocyte progenitor cells responded to injury.
They hypothesized that, if the cells use the extension and retraction of the filopodia to recognize other nearby oligodendrocyte progenitor cells, then other cells would fill the void if oligodendrocyte progenitor cells were destroyed. To test this, the scientists ablated specific oligodendrocyte progenitor cells with a Photonic Micropoint laser system using 5.0-s pulses of 440-nm light.
Within three hours of ablation, oligodendrocyte progenitor cells began to migrate into the unoccupied areas. As early as two hours after ablation, nearby oligodendrocyte progenitor cells began remodeling their processes to extend into the ablated area, and the cells filled the area in five hours. In zebra fish with ablated regions, 44 percent of the cells divided, compared with only 16 percent of the cells in control embryos. In addition, the researchers found that replacement cells came only from oligodendrocyte progenitor cells and not from other types of neural precursors.
Appel said that his group is identifying genes tied to myelination that might be useful for therapies and testing their roles with their system.
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