Transplantation of an oligodendrocyte cell line leading to extensive myelination.

Abstract

Oligodendrocytes, the myelin-forming cells of the central nervous system, can be generated from progenitor cell lines and assayed for their myelinating properties after transplantation. A growth-factor-dependent cell line of rat oligodendrocyte progenitors (CG4) was carried through 31-48 passages before being transplanted into normal newborn rat brain or the spinal cord of newborn myelin-deficient (md) rats. In md rat spinal cord, CG4 oligodendrocyte progenitors migrated up to 7 mm along the dorsal columns, where they divided and myelinated numerous axons 2 weeks after grafting. CG4 cells were transfected with the bacterial lacZ gene and selected for high -galactosidase expression. The cell migration and fate of these LacZ+ cells were analyzed after transplantation. In normal newborn brain, LacZ+ oligodendrocyte progenitors migrated along axonal tracts from the site of injection and integrated in the forming white matter. In md rats, extensive migration (up to 12 mm) was revealed by staining for 3-galacsdase activity of the intact spinal cord where many grafted cells had moved into the posterior columns. Similar . 4-7). As myelin facilitates rapid impulse conduction along most nerve tracts, the lack or loss of myelin can result in important neurological dysfunction (8,9).Although the development of the oligodendrocyte lineage has been studied extensively in vitro (4-7), the molecular mechanisms controlling the growth and migration of oligodendrocyte progenitors (OPs) in vivo, the recognition of axons, the coordinate synthesis of multiple myelin internodes, the regulation of myelin protein genes, and the death of superfluous oligodendrocytes are largely unknown. An understanding of these molecular mechanisms could be approached by using a growth-factor-dependent cell line in which gene transfer or targeting could be performed and tested in a transplantation paradigm. In addition, such myelin-forming cell lines could be used to explore ways to experimentally correct genetic defects identified in several inherited dysmyelinating diseases and leukodystrophies of animals and humans (10, 11).Transplanted rodent oligodendrocytes and their precursor cells as well as stem cells can myelinate axons in myelindeficient or demyelinated animals (12)(13)(14)(15)(16)(17)(18)(19). Recent studies have shown that transplantation ofearly passages of OPs (20) expanded in the presence of growth factors or immortalized with a conditional oncogene (21), resulted in myelination of naked axons within an irradiated and chemically induced spinal cord lesion in adult rats. However, the myelinating potential ofthese grafted cells decreased considerably (20) (24,25).MATERIALS AND METHODS Cells and Animals. Newborn Sprague-Dawley rats were purchased from Taconic Farms and the breeding colony of md rats was maintained as described (23,24). CG4 cells were kindly provided by J.-C. Louis (presently at Amgen Biologicals). Cells were expanded in T25 or T75 flasks in serum-free DMEM (GIBCO/BRL) supplemented with N1, biotin, and 30% B104 ...

Oligodendroglial Progenitor Cell Therapy Limits Central Neurological Deficits in Mice with Metachromatic Leukodystrophy

“…When OLPs were transplanted in the unmyelinated brain of MLD newborns, almost all of surviving cells became MBPϩ PLPϩ OLs. These results are consistent with other reports using similar preparations of OLPs in models of myelin disorders (Tontsch et al, 1994;Franklin and Blakemore, 1995;Learish et al 1999;Espinosa de los Monteros et al 2001;Crang et al, 2004).…”

The Myelin Mutants as Models to Study Myelin Repair in the Leukodystrophies

“…2). In the md rat, in some cases almost the entire dorsal column can be myelinated at the site of engraftment [22,23,77]. In both the shi and md rat, myelin made by transplanted cells is characteristically thin, similar to that seen in endogenous remyelination.…”

“…This rat, therefore, can model those cases of severe connatal PMD associated with a duplication in the plp1 gene [20]. It will be important to determine whether the milieu of the CNS in PLP over-expression, hence the majority of PMD patients, is receptive to transplanted cells as it is in the missense mutations (md rat and the shaking [sh] pup) [21][22][23].…”

“…In the case of lacZ, the spinal cord or brain at the site of transplantation is incubated overnight in X-gal solution [23]. In 24 hours, a blue reaction product is seen at the site, which will frequently overlap with a white patch seen grossly prior to X-gal staining [23].…”

“…In the case of lacZ, the spinal cord or brain at the site of transplantation is incubated overnight in X-gal solution [23]. In 24 hours, a blue reaction product is seen at the site, which will frequently overlap with a white patch seen grossly prior to X-gal staining [23]. More recently, GFP has become the standard method of tracking cells and has some advantages over lacZ, as cell tracking is performed by shining a fluorescent light on the transplanted tissue and positive areas can be selected for further studies (Fig.…”