STAGE-SPECIFIC OLIGONUCLEOTIDE UPTAKE IN MURINE BONE-MARROW B-CELL PRECURSORS

Fluorescein isothiocyanate (FITC)-conjugated phosphodiester and phosph orothioate oligonucleotides were used in four-color flow cytometry wit h murine bone marrow cells stained with monoclonal antibody specific f or the differentiation markers B220, S7 (CD43), and BP-1 to show possi ble stage-specific oligonucleotide uptake. Relatively low uptake was o bserved among pre-Pro- and early Pro-B cells. Late Pro-B- and pre-B ce lls had increased oligonucleotide uptake, whereas B cells had a lower level. cell membrane binding of oligonucleotides varied during B-cell differentiation in parallel with internalization, which was documented by confocal microscopy. An FITC-conjugated polyanionic dextran sulfat e also showed differentiation-related B-cell association, suggesting t he presence of cell membrane binding sites specific for polyanions as opposed to a unique feature of the DNA backbone. Interpretation of ant isense experiments in murine bone marrow cells will need to account fo r the heterogeneous oligonucleotide uptake among differentiating B cel ls. (C) 1994 by The American Society of Hematology.