In vitro myelination by oligodendrocyte precursor cells transfected with the neurotrophin-3 gene.
Oligodendrocyte precursor cells require exogenous neurotrophin-3 (NT-3) for differentiation into oligodendrocytes. We transfected precursor cells with the gene for NT-3 and looked for changes in their development into myelin-forming cells. The expression of NT-3 in transfected cells was demonstrated by reverse transcription followed by PCR as well as by Northern blots. Direct synthesis of the neurotrophin product and its release to the culture supernatants were also shown by specific ELISA. Transfection converts precursor cells into actively dividing cells that can incorporate 3H-thymidine into DNA. In the absence of growth factors, a parallel increase in the survival of the transfected cultures was also demonstrated by the MTT test. The final demonstration of biological changes in transfected versus untreated cells was a 10-fold increase in myelin basic protein production observed in Western blots and the direct observation by phase-contrast and electron microscopy of myelin membranes in cocultures with hippocampal neurons. We discuss the future use of this transfected cells in regeneration and functional recovery in experimental models of multiple sclerosis.
Rubio
Nazario
N
Instituto Cajal, Consejo Superior de Investigaciones Cientificas (CSIC), Madrid, Spain. nazario@cajal.csic.es
Rodriguez
Rodrigo
R
Arevalo
Maria Angeles
MA
D000818:Animals; D000831:Animals, Newborn; D002454:Cell Differentiation / Q000235:genetics*; D002455:Cell Division / Q000235:genetics; D002470:Cell Survival / Q000235:genetics; D002478:Cells, Cultured; D003711:Demyelinating Diseases / Q000235:genetics / Q000378:metabolism / Q000628:therapy; D005822:Genetic Vectors / Q000235:genetics; D008854:Microscopy, Electron; D004676:Myelin Basic Protein / Q000096:biosynthesis; D009186:Myelin Sheath / Q000235:genetics / Q000378:metabolism* / Q000648:ultrastructure; D009416:Nerve Regeneration / Q000235:genetics; D020933:Neurotrophin 3 / Q000096:biosynthesis / Q000235:genetics*; D009836:Oligodendroglia / Q000378:metabolism* / Q000648:ultrastructure; D012333:RNA, Messenger / Q000378:metabolism; D051381:Rats; D017208:Rats, Wistar; D013234:Stem Cells / Q000378:metabolism* / Q000648:ultrastructure; D014162:Transfection / Q000379:methods; D015854:Up-Regulation / Q000235:genetics
D004676:Myelin Basic Protein; D020933:Neurotrophin 3; D012333:RNA, Messenger