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Expression of fibronectin and laminin by different types of mouse glial cells cultured in a serum-free medium

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Abstract

The expression of fibronectin and laminin by cultured glial cells was studied. The glial culture from neonatal mouse cerebra maintained in a chemically defined, serum-free medium consisted of type-1 astrocytes, oligodendrocyte-type-2 astrocyte (O-2A) progenitor cells, oligodendrocytes and type-2 astrocytes. Double-labelling immunofluorescent experiments performed using the mixed glial culture indicated that fibronectin and laminin are expressed in different patterns among the glial subtypes. The staining intensities with anti-fibronectin or anti-laminin antibodies decreased in the order: type-1 astrocytes, O-2A progenitor cells and type-2 astrocytes. Both molecules were deposited in a fibrillar matrix underneath type-1 astrocytes, whereas only intracytoplasmic localization of these molecules was observed with O-2A progenitor cells and type-2 astrocytes. Western blot analysis showed that glial fibronectin has a slightly higher molecular weight than mouse plasma fibronectin (230 kDa) and that glial laminin is a variant with a 220 kDa B chain present and the 400 kDa A chain missing. Using enzyme-linked immunosorbent assays (ELISA), these molecules were detected in the glial extracellular matrix at the concentration of 4 ng/106 cells. A large amount of fibronectin (82 ng/106 cells) was secreted into the culture medium, while secretion of laminin was not detected.

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References

  • Akers RM, Mosher DF and Lilien JE (1981) Promotion of retinal neurite outgrowth by substratum-bound fibronectin. Dev. Biol. 86: 179–188.

    Google Scholar 

  • Amrani DL, Falk MJ and Mosesson MW (1985) Studies of fibronectin synthesized by cultured chick hepatocytes. Exp. Cell Res. 160: 171–183.

    Google Scholar 

  • Baum BJ, McDonald JA and Crystal RG (1977) Metabolic fate of the major cell surface protein of normal human fibroblasts. Biochem. Biophys. Res. Commun. 79: 8–15.

    Google Scholar 

  • Bögler O, Wren D, Barnett SC, Land H and Noble M (1990) Cooperation between growth factors promotes extended self-renewal and inhibits differentiation of oligodendrocyte-type-2 astrocyte (O-2A) progenitor cells. Proc. Natl. Acad. Sci. USA 87: 6368–6372.

    Google Scholar 

  • Chiu AY, delos Monteros AE, Cole RA, Loera S and de Vellis J (1991) Laminin and s-laminin are produced and released by astrocytes, schwann cells, and schwannomas in culture. Glia 4: 11–24.

    Google Scholar 

  • ffrench-Constant C and Raff MC (1986) The oligodendrocyte-type 2 astrocyte cell lineage is specialized for myelination. Nature (London) 323: 335–338.

    Google Scholar 

  • Gallo V and Bertolotto A (1990) extracellular matrix of cultured glial cells: Selective expression of chondroitin 4-sulfate by type-2 astrocytes and their progenitors. Exp. Cell. Res. 187: 211–223.

    Google Scholar 

  • Hatten ME, Furie MB and Rifkin DB (1982) Binding of developing mouse cerebellar cells to fibronectin: a possible mechanism for the formation of the external granular layer. J. Neurosci. 2: 1195–1206.

    Google Scholar 

  • Hayman EG and Ruoslahti E (1979) Distribution of fetal bovine serum fibronectin and endogenous rat cell fibronectin in extracellular matrix. J. Cell Biol. 83: 255–259.

    Google Scholar 

  • Hynes RO (1990) Fibronectins. Springer-Verlag, New York.

    Google Scholar 

  • Ignotz RA and Massague J (1986) Transforming growth factor-ß stimulates the expression of fibronectin and collagen and their incorporation into the extracellular matrix. J. Biol. Chem. 261: 4337–4345.

    Google Scholar 

  • Ishikawa E, Imagawa M, Hashida S, Yoshitake S, Hamaguchi Y and Ueno T (1983) Enzyme-labelling of antibodies and their fragments for enzyme immunoassay and immunohistochemical staining. J. Immunoassay 4: 209–327.

    Google Scholar 

  • Janzer RC and Raff MC (1987) Astrocytes induce blood-brain barrier properties in endothelial cells. Nature (London) 325: 253–257.

    Google Scholar 

  • Laemmli UK (1970) Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature (London) 227: 680–685.

    Google Scholar 

  • Liesi P (1985) Do neurons in the vertebrate CNS migrate on laminin? EMBO J. 4: 1163–1170.

    Google Scholar 

  • Liesi P, Dahl D and Vaheri A (1983) Laminin is produced by early rat astrocytes in primary culture. J. Cell Biol. 96: 920–924.

    Google Scholar 

  • Liesi P, Kirkwood T and Vaheri A (1986) Fibronectin is expressed by astrocytes cultured from embryonic and early postnatal rat brain. Exp. Cell Res. 163: 175–185.

    Google Scholar 

  • Liesi P, Kaakkola S, Dahl D and Vaheri A (1984) Laminin is induced in astrocytes of adult brain by injury. EMBO J. 3: 683–686.

    Google Scholar 

  • Liesi P and Risteli L (1989) Glial cells of mammalian brain produce a variant form of laminin. Exp. Neurol. 105: 86–92.

    Google Scholar 

  • Manthorpe M, Engvall E, Ruoslahti E, Longo FM, Davis GE and Varon S (1983) Laminin promotes neuritic regeneration from cultured peripheral and central neurons. J. Cell Biol. 97: 1882–1890.

    Google Scholar 

  • Martin GR and Timpl R (1987) Laminin and other basement membrane components. Annu. Rev. Cell Biol. 3: 57–85.

    Google Scholar 

  • Mckinnon RD, Matsui T, Dubois-Dalcq M and Aaronson SA (1990) FGF modulates the PDGF-driven pathway of oligodendrocyte development. Neuron 5: 603–614.

    Google Scholar 

  • Michler-Stuke A, Wolff J and Bottenstein JE (1984) Factors influencing astrocyte growth and development in defined media. Int. J. Devl. Neurosci 2: 575–584.

    Google Scholar 

  • Norton WT, Farooq M, Fields KL and Raine CS (1983) The long-term culture of bulk-isolated bovine oligodendroglia from brain. Brain Res. 270: 295–310.

    Google Scholar 

  • Paetau A (1988) Glial fibrillary acidic protein, vimentin and fibronectin in primary cultures of human gliomas and fetal brain. Acta Neuropathl. 75: 448–455.

    Google Scholar 

  • Price J and Hynes RO (1985) Astrocytes in culture synthesize and secrete a varient form of fibronectin. J. Neurosci. 5: 2205–2211.

    Google Scholar 

  • Raff MC (1989) Glial cell diversification in the rat optic nerve. Science 243: 1450–1455.

    Google Scholar 

  • Raff MC, Abney ER, Cohen J, Lindsay R and Noble M (1983a) Two types of astrocytes in cultures of developing rat white matter: differences in morphology, surface gangliosides, and growth characteristics. J. Neurosci. 3: 1289–1300.

    Google Scholar 

  • Raff MC, Fields KL, Hakomori S, Mirsky R, Pruss RM and Winter J (1979) Cell-type-specific markers for distinguishing and studying neurons and the major classes of glial cells in culture. Brain Res. 174: 283–308.

    Google Scholar 

  • Raff MC, Miller RH and Noble M (1983b) A glial progenitor cell that develops in vitro into an astrocyte or an oligodendrocyte depending on culture medium. Nature (London) 303: 390–396.

    Google Scholar 

  • Raff MC, Mirsky R, Fields KL, Lisak RP, Dorfman SH, Silberberg DH, Gregson NA, Liebowitz S and Kennedy MC (1978) Galactocerebroside: a specific cell surface antigen marker for oligodendrocytes in culture. Nature (London) 274: 813–816.

    Google Scholar 

  • Richardson WD, Pringle N, Mosley MJ, Westermark B and Dubois-Dalcq M (1988) A role for platelet-derived growth factor in normal gliogenesis in the central nervous system. Cell 53: 309–319.

    Google Scholar 

  • Rogers SL, Letourneau PC, Palm SL, McCarthy J and Furcht LT (1983) Neurite extension by peripheral and central nervous system neurons in response to substratum-bound fibronectin and laminin. Dev. Biol. 98: 212–220.

    Google Scholar 

  • Schlaepfer W (1977) Immunological and ultrastructural studies of neurofilaments isolated from rat peripheral nerve. J. Cell Biol. 74: 226–240.

    Google Scholar 

  • Stewart GR and Pearlman AL (1987) Fibronectin-like immunoreactivity in the developing cerebral cortex. J. Neurosci. 7: 3325–3333.

    Google Scholar 

  • Towbin H, Staehelin T and Gordon J (1979) Electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets: procedure and some applications. Proc. Natl. Acad. Sci. USA 76: 4350–4354.

    Google Scholar 

  • Yamada KM, Yamada SS and Pastan I (1976) Cell surface protein partially restores morphology, adhesiveness, and contact inhibition of movement to transformed fibroblasts. Proc. Natl. Acad. Sci. USA 73: 1217–1221.

    Google Scholar 

  • Yoshida T and Takeuchi M (1991) Primary culture and cryopreservation of mouse astrocytes under serum-free conditions. Cytotechnology 5: 99–106.

    Google Scholar 

  • Wujek JR, Haleem-Smith H, Yamada Y, Lipsky R, Lan YT and Freese E (1990) Evidence that the B2 chain of laminin is responsible for neurite outgrowth-promoting activity of astrocyte extracelluar matrix. Dev. Brain Res. 55: 237–247.

    Google Scholar 

  • Zhou FC (1990) Four patterns of laminin-immunoreactive structure in developing rat brain. Dev. Brain Res. 55: 191–201.

    Google Scholar 

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Yoshida, T., Takeuchi, M. Expression of fibronectin and laminin by different types of mouse glial cells cultured in a serum-free medium. Cytotechnology 7, 187–196 (1991). https://doi.org/10.1007/BF00365930

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