Summary
Tissue-plasminogen activator (t-PA) has been identified in human megakaryocytes isolated from human rib fragments or from culture cells. The presence of t-PA antigen in the endoplasmic reticulum of mature cells was demonstrated by ultrastructural immunochemistry. These results suggest that t-PA can be synthesized by megakaryocytes as well as by vascular endothelial cells. In order to confirm this possibility, in situ hybridization using antisense RNA biotinylated probe was used in megakaryocyte culture at different steps of maturation. The presence of t-PA-RNA was clearly demonstrated at an early step of differentiation. Biotin staining was enhanced in mature cells and this was correlated with the detection of t-PA antigen by immunocytochemistry.
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References
Anderson MLM, Young BD (1985) Quantitative filter hybridization. In: Hames BD, Higgins SJ (eds) Nucleic acid hybridization. IRL Press, Oxford-Washington DC, pp 73–112
Avner P, Arnaud D, Sultan Y, Maisonneuve P, Jeanneau C (1984) Monoclonal antibody against the human factor VIII/von Willebrand molecule: characterization and potential for screening of von Willebrand patients. Dev Biol Stand 57:69–76
Binder M, Tourmente S, Roth J, Renaud M, Gehring WJ (1986) In situ hybridization at the electron microscope level: localization of transcripts on ultrathin sections of Lowicryl K4M-embedded tissue using biotinylated probes and protein-A-gold complexes. J Cell Biol 102:1646–1653
Bloch B, Popouili T, Le Guellec D, Normand E, Chouham S, Guitteny AF, Bohlem P (1986) In situ hybridization histochemistry for the analysis of gene expression in the endocrine and central nervous system tissues: a 3-year experience. J Neurosci Res 16:183–200
Brigati DJ, Myerson D, Leary JJ, Spalholz B, Travis SZ, Fong CKY, Hsiung GD, Ward DC (1983) Detection of viral genomes in cultured cells and paraffin-embedded tissue sections using biotin-labeled hybridization probes. Virology 126:32–50
Brisson-Jeanneau C, Solberg LA, Sultan Y (1990) Presence of functionnally active tissue-plasminogen activator in human CFU-M derived megakaryocytes in vitro. Fibrinolysis 4:107–115
Chan VTW, Fleming KA, McKee JOD (1985) Detection of subpicogram quantities of specific DNA sequences on blot hybridization with biotinylated probes. Nucleic Acid Res 13:8083–8091
Jeanneau C, Nichols WL, Solberg LA, Sultan Y (1986) Fibrinolytic activity and immunolocalization of t-PA in human megakaryocytes. Blood 68:347 (Abstract)
Jeanneau C, Sultan Y (1988) Tissue plasminogen activator in human megakaryocytes and platelet: immunocytochemical localization, immunoblotting and zymographic analysis. Thromb Haemost 59:529–534
Koga Y, Caccian, Toyonaga B, Spolsky R, Yanagi Y, Yoshikai Y, Mark TN (1986) A human T cell specific cDNA clone (YT 16) encodes a protein with extensive homology to a family of protein-tyrosine kinases. Eur J Immunol 16:1643–1646
Langer PR, Walsrop AA, Ward DC (1981) Enzymatic synthesis of biotin-labeled polynucleotides: novel nucleic acid affinity probes. Proc Natl Acad Sci USA 78:6633–6637
Loskutoff DJ, Edgington T (1977) Synthesis of a fibrinolytic activator and inhibitor by endothelial cells. Proc Natl Acad Sci USA 74:3903–3907
Maniatis T, Fritsch EF, Sambrook J (1982) Molecular cloning. A laboratory manual. Cold Spring Harbor Laboratory
Marks GJ, Hart TK, Rush GF, Hoffstein ST, Fong KL, Bugelski PJ (1990) Internalization of recombinant tissue-type plasminogen activator by isolated rat hepatocytes is via coated pits. Thromb Haemost 63:251–258
Nelles L, Lijnen RH, Collen D, Holmes WE (1987) Characterization of a fusion protein consisting of amino acids 1 to 263 of tissue-type plasminogen activator and amino acids 144 to 411 of urokinase-type plasminogen activator. J Biol Chem 262:10855–10862
Rouer E, Van Huynh T, Lavareda de Souza S, Lang MC, Fischer S, Benarous R (1989) Structure of the human lck gene: differences in genomic organisation within src-related genes affect onlyN-terminal exons. Gene 84:105–113
Singer HR, Ward DC (1982) Actin gene expression visualized in chicken muscle tissue culture by using in situ hybridization with a biotinated nucleotide analog. Proc Natl Acad Sci USA 79:7331–7335
Solberg LA, Lamal N, Messner HA (1985) Characterization of human megakaryocytic colony formation in human plasma. J Cell Physiol 124:6–74
Veillette A, Foss FM, Sausville EA, Bolen JB, Rose NN (1987) Expression of the LCK tyrosine kinase gene in human colon carcinoma and other non lymphoid human tumor cell lines. Oncogene Res 1:357–374
Vinci G, Tabilio A, Deschamps JF, van Haecke D, Henri A, Guichard J, Terreroo P, Lansdorp PM, Hercend T, Vainchenker W, Breton-Gorius J (1984) Immunological study of in vitro maturation of human megakaryocytes. Br J Haematol 56:589–605
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Brisson-Jeanneau, C., Nelles, L., Rouer, E. et al. Tissue-plasminogen activator RNA detected in megakaryocytes by in situ hybridization and biotinylated probe. Histochemistry 95, 23–26 (1990). https://doi.org/10.1007/BF00737224
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DOI: https://doi.org/10.1007/BF00737224