Transplantation of Cultured Premyelinating Oligodendrocytes into Normal and Myelin-Deficient Rat Brain

Zhang, Mao Sheng; Gordon, Marcia N.; Aymie, Michael; Vellis, Jean de

doi:10.1159/000111653

Dev Neurosci

1992

DOI: 10.1159/000111653

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Transplantation of Cultured Premyelinating Oligodendrocytes into Normal and Myelin-Deficient Rat Brain

Mao Sheng Zhang¹,

Marcia N. Gordon²,

Michael Aymie³

et al.

Abstract: Cultures of oligodendroglial cells at various stages of maturation, from progenitors to maturing oligodendrocytes, were prepared from neonatal rat brain primary cultures and then were prelabeled in the culture dish with the fluorescent dye, fast blue (FB). Single cell suspensions were grafted into normal or myelin-deficient rat brains. The normal as well as the myelin-deficient in vivo environment allowed cell survival, migration, and differentiation. The FB⁺ cells expressed the oligodendroglial mar… Show more

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Cited by 30 publications

(22 citation statements)

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“…The phenotypic differentiation of 02A cells was studied in tissue sections by single and double immunolabeling and examined by fluorescence microscopy to visualize grafted cells and their progeny. We have reported (9) that FB-labeled cultured glial cells transplanted into normal and myelin-deficient (md) Wistar rats remain identifiable in immunostained tissue sections up to 6 months after grafting (9). We now report that transplanted 02A progenitors in vivo develop into oligodendrocytes but not astrocytes.…”

mentioning

confidence: 87%

“…§1734 solely to indicate this fact. the fluorescent dye FB (Sigma) as described (9). Briefly, a sterile aqueous solution ofFB dye was prepared at 10 mg/ml.…”

mentioning

confidence: 99%

“…We have previously reported that single or double immunofluorescence of tissue sections allowed us to identify host FB-and grafted FBI cells (9). All immunostained cell cultures and tissue sections were analyzed with a Nikon FX FIG.…”

mentioning

confidence: 99%

“…We have shown (9) that FB is not cytotoxic and does not alter the differentiation of glial cells in vitro and in vivo after transplantation. In vivo-grafted cells expressed GC, glycerol phosphate dehydrogenase, and myelin basic protein (9). The dye does not pass into other cells and can be visualized by fluorescence microscopy for >6 months.…”

mentioning

confidence: 99%

“…The dye does not pass into other cells and can be visualized by fluorescence microscopy for >6 months. Although cell proliferation results in dilution of the dye, FB+ cells show intense labeling after six cell divisions in culture (9). Thus FB dye appears to be one of the best cell markers at the light microscopic level for tracking single cells and studying their differentiation and fate in the in vivo microenvironment.…”

mentioning

confidence: 99%

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O2A progenitor cells transplanted into the neonatal rat brain develop into oligodendrocytes but not astrocytes.

Monteros

Zhang

Vellis

1993

Proc. Natl. Acad. Sci. U.S.A.

124

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The differentiation of the bipotential 02Aprogenitor cell into an oligodendrocyte or a type 2 astrocyte has been well documented in cell cultures of various regions of the central nervous system. The appropriate tools to prove its existence in vivo have been lacking. We report on an in vitro-in vivo approach that combines stable labeling of an enriched population of cultured 02A progenitors by the fluorescent dye fast blue, followed by their transplantation into neonatal rat brains, which allowed us to study the influence of the brain microenvironment on their lineage decision. Recently, the existence of type 2 astrocytes in vivo has been questioned (6, 7). Raffand coworkers (5) have estimated the in vivo appearance of type 1 astrocytes, oligodendrocytes, and type 2 astrocytes in the rat optic nerve to be at postnatal day 0 (PO), P4, and P15, respectively. However, a light and electron microscopic immunocytochemical analysis of [3H]thymidine pulse-labeled developing rats has revealed only two periods of astrocyte and oligodendrocyte generations, the earliest being that of GFAP+ cells near PO followed by a wave of oligodendrocytes at P4-P8 (6, 7). A major obstacle to resolve this controversy has been the lack of appropriate markers to identify type 2 astrocytes in the brain (5). The antibody A2B5 is not a reliable marker in tissue sections because it stains unrelated cell types and intracellular structures (for review, see refs. 7 and 8).In this study, we propose an approach to compare the fate of 02A progenitors in the in vivo vs. in vitro environment. An enriched population of 02A progenitors in culture was labeled with the fluorescent dye fast blue (FB) and transplanted into neonatal rat brains. The phenotypic differentiation of 02A cells was studied in tissue sections by single and double immunolabeling and examined by fluorescence microscopy to visualize grafted cells and their progeny. We have reported (9) that FB-labeled cultured glial cells transplanted into normal and myelin-deficient (md) Wistar rats remain identifiable in immunostained tissue sections up to 6 months after grafting (9). We now report that transplanted 02A progenitors in vivo develop into oligodendrocytes but not astrocytes. MATERIALS AND METHODSAnimals. Normal Wistar and md Wistar rat pups were raised in our breeding colonies. Rats were housed in a restricted-access temperature-controlled vivarium on a 12-h light/12-h dark cycle, with free access to food and water. Fifty percent of pups born to previously identified female carriers of the md trait are genetically normal (+/y) and 50% carry the md trait (md/y). Males ofboth genotypes were used at P5 as recipients for cell transplants.Cell Culture. Primary cultures were prepared from newborn rat brains as described (10). These cultures were maintained in Waymouth medium supplemented with 10% (vol/ vol) fetal calf serum. After 9 days in culture, 02A cells growing on top ofthe astrocyte monolayer (type 1 astrocytes) were removed by a modification of the method of McCarthy and de Vel...

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mentioning

confidence: 87%

“…§1734 solely to indicate this fact. the fluorescent dye FB (Sigma) as described (9). Briefly, a sterile aqueous solution ofFB dye was prepared at 10 mg/ml.…”

mentioning

confidence: 99%

mentioning

confidence: 99%

mentioning

confidence: 99%

mentioning

confidence: 99%

See 3 more Smart Citations

O2A progenitor cells transplanted into the neonatal rat brain develop into oligodendrocytes but not astrocytes.

Monteros

Zhang

Vellis

1993

Proc. Natl. Acad. Sci. U.S.A.

124

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Transferrin is an early marker of hepatic differentiation, and its expression correlates with the postnatal development of oligodendrocytes in mice

et al. 1997

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Transferrin (Tf), the iron transport protein, is essential for the growth and differentiation of cells. Therefore, it provides an excellent model to analyze the regulatory mechanisms controlling the expression of a eukaryotic gene in different cell types and during fetal and adult life. In this study, the tissue-specific and developmental regulation of the Tf gene in vivo were analyzed. Human Tf mRNA was detected mainly in fetal and adult liver. A weaker expression was observed in adult and fetal brain and in fetal spleen. By in situ hybridization the presence of mouse Tf mRNA was detected in the hepatic primordia. This is the first observation pointing out Tf as an early marker of hepatic differentiation, prior to the formation of the liver. Thus, TF may be an important tool to follow the hepatic specification of the gut endoderm. Mouse Tf mRNA was also detected in the liver bud and subsequently in the liver throughout fetal life, and in newborn and adult animals. No expression of the Tf gene was observed in the mouse fetal central nervous system (CNS). In contrast, Tf mRNA was detected from the 5th day after birth in the derivatives of the caudal part of the neural tube and subsequently in the derivatives of the rhomboencephalon and that of the prosencephalon. These results indicate that Tf gene expression correlates with the postnatal development of oligodendrocytes in the mouse CNS. To test whether the control elements of the human gene previously found in ex vivo experiments were also active in vivo during fetal and adult life, we fused the -4000/+395' flanking region of the human gene to the coding region of the lacZ gene and generated transgenic mice. The expression of the reporter gene during development was analyzed.

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Transplantation of CG4 oligodendrocyte progenitor cells in the myelin-deficient rat brain results in myelination of axons and enhanced oligodendroglial markers

et al. 1997

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Transplantation of oligodendrocyte (Ol) progenitor cells into the central nervous system is a promisingapproach for the treatment of myelin disorders. This approach requires providing adequate numbers of healthy cells with myelinating potential. We recently showed the successful transplantation of Ol progenitors into the myelin-deficient (md) rat brain. In the present work, CG4 cells, a cell line with properties of Ol progenitors, were labeled with fast blue and grafted into P3-P5 pups born to carrier mothers. Examination of host brains 2 weeks posttransplant indicated that CG4 cells display a much more extensive migration capacity than their wild-type counterparts. These cells synthesized myelin components. In addition, ultrastructural analysis showed myelin formation along axons of md hosts in various brain regions, including corpus callosum, cerebellum, and brainstem. Furthermore, in situ hybridization studies performed on sagittal sections revealed extensive expression of transferrin-mRNA within the md host parenchyma. The high survival and functional features displayed by CG4 cells after transplantation, together with their striking wide distribution within the host parenchyma, as assessed by the presence of myelinated fibers in mutant hosts, emphasizes the importance of using highly motile and proliferative Ol progenitor cells. Strategies to improve the condition and life span of md rat pups are currently under investigation.

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Transplantation of Cultured Premyelinating Oligodendrocytes into Normal and Myelin-Deficient Rat Brain

Cited by 30 publications

References 24 publications

O2A progenitor cells transplanted into the neonatal rat brain develop into oligodendrocytes but not astrocytes.

O2A progenitor cells transplanted into the neonatal rat brain develop into oligodendrocytes but not astrocytes.

Transferrin is an early marker of hepatic differentiation, and its expression correlates with the postnatal development of oligodendrocytes in mice

Transplantation of CG4 oligodendrocyte progenitor cells in the myelin-deficient rat brain results in myelination of axons and enhanced oligodendroglial markers

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