ABSTRACT
Insulin-like growth factor II (IGF-II) plays significant roles in the growth and development of mammals through the regulation of mitogenesis and cell survival. Previously, IGF-II mRNA transcripts within the CNS were detected in the choroid plexus and leptomeninges (DeChiara et al., 1991). The objective of this study was to determine the expression pattern of IGF-II mRNA in different cell types of the cerebellum during development. We report here that the IGF-II gene is transcribed in granule and glial cells within the cerebellar parenchyma at various times during the early postnatal period in mice. IGF-II gene expression is further regulated by parent-specific imprinting such that only the paternal IGF-II allele is expressed in granule cells. In contrast, choroid plexus and leptomeninges express IGF-II mRNAs biallelically, indicating that cell type-specific regulation of genomic imprinting occurs within the mammalian CNS.
Subject(s)
Cerebellum/metabolism , Gene Expression Regulation/physiology , Genomic Imprinting , Insulin-Like Growth Factor II/genetics , Animals , Animals, Newborn , Cells, Cultured , Cerebellum/cytology , Choroid Plexus/metabolism , Mice , Mice, Inbred C57BL , Neurons/metabolism , Polymerase Chain Reaction/methods , RNA, Messenger/analysis , Transcription, GeneticABSTRACT
We have successfully established mixed glial cell primary cultures prepared from individual fetal human brains (15-18 weeks' gestation in age). Cultures were maintained for as long as 3 months in either 10% fetal calf serum (FCS) or serum-free chemically defined medium (CDM). By morphological and immunohistochemical criteria, the precursor cell for human oligodendrocytes (O-2A cell) was identified. This cell exhibited the bipolar morphology and A2B5-positive (A2B5+) immunoreactivity typical of the O-2A precursor cell. With time in culture, cells possessing a stellate morphology appeared, some of which stained with the O4 antibody, indicative of cell differentiation in the oligodendroglial lineage. At yet older culture age, arborized cells bearing the O1 (galactocerebroside, GC) immunohistochemical marker and displaying the morphological characteristics typical of more mature oligodendrocytes were found, confirming their oligodendroglial identity. Oligodendroglial differentiation was supported best by CDM rather than FCS. To complement these observations, double immunofluorescent studies were performed on parietal sections from human fetal brains at 20 to 22 weeks of gestation. Bipolar A2B5+, multipolar A2B5+/O4+, and arborized A2B5-/O1+ cells were found, thus confirming the presence of oligodendrocytes in human fetal brain at this stage of prenatal development and consistent with the observations made in cell culture.
Subject(s)
Brain/embryology , Oligodendroglia/cytology , Culture Media , Culture Techniques , Fluorescent Antibody Technique , Humans , Immunoenzyme Techniques , Stem Cells/cytologyABSTRACT
In summary, our studies show that IGFs are potent regulators of oligodendrocyte development and myelination in vitro and in vivo. IGFs act at several levels: by promoting proliferation of oligodendrocytes and oligodendrocyte precursors, by inducing immature oligodendrocyte precursors to develop into oligodendrocytes, and by regulating myelin gene expression and the amount of myelin produced per oligodendrocyte. Our findings indicate that IGFs play a crucial role in normal oligodendrocyte development and myelination, and suggest that IGFs may have applications for the promotion of remyelination in myelin disorders such as MS.
Subject(s)
Brain/cytology , Insulin-Like Growth Factor II/pharmacology , Insulin-Like Growth Factor I/biosynthesis , Insulin-Like Growth Factor I/pharmacology , Oligodendroglia/cytology , Animals , Cell Differentiation/drug effects , Cell Division , Cells, Cultured , Gene Expression , Mice , Mice, Transgenic , Myelin Proteins/biosynthesis , Nerve Regeneration/drug effects , Oligodendroglia/drug effects , Oligodendroglia/physiology , RNA, Messenger/biosynthesis , RatsABSTRACT
Insulin-like growth factor I (IGF-I) and high concentrations of insulin have been shown to stimulate an increase in the number of oligodendrocytes that appear in developing monolayer cultures of rat brain cells (McMorris et al., Proc Natl Acad Sci USA 83: 822-826, 1986; McMorris et al., Ann NY Acad Sci 605:101-109, 1990; McMorris and Dubois-Dalcq, J Neurosci Res 21:199-209, 1988). In the present study, we investigated whether IGF-I or insulin treatment induces a corresponding increase in the synthesis and accumulation of myelin. Aggregate cultures, established from 16-day-old fetal rat brains, were treated with either 100 ng/ml IGF-I or 5,000 ng/ml insulin and analyzed for the number of oligodendrocytes, activity of 2',3'-cyclic nucleotide 3'-phosphohydrolase (CNP), total amount of myelin, and synthesis rate of myelin proteins. Cultures treated with IGF-I beginning on day 2 after explantation contained 35-80% more oligodendrocytes and had 60-160% higher CNP activity than controls when tested on day 13, 20, or 27. By day 27, treated cultures had 35-90% more myelin than controls. Similar results were observed in response to 5,000 ng/ml insulin, a concentration at which insulin binds to IGF receptors and acts as an analogue of IGF-I. The synthesis rate of myelin proteins was measured in experiments using 5,000 ng/ml insulin. When treatment was begun at day 20 rather than day 2, cultures did not exhibit an increased number of oligodendrocytes over control during the following 4-6 days.(ABSTRACT TRUNCATED AT 250 WORDS)
