Mitotic separation of daughter cells in human lymphoma B cell line Daudi involves L-selectin engagement and shedding

Background and Objective. A novel role for shedding of the surface molecule L-selectin has been proposed as an adjunctive phenomenon during cell detac hment from marrow stroma or vessel endothelium. We wished to examine whethe r variations in expression of L-selectin on a lymphoma B cell line were lin ked to shedding. Design and Methods. Mapping of L-selectin expression on the surface of Daud i lymphoma cells was performed by flow cytometry, fluorescence microscopy, and electron microscopy. Levels of shed L-selectin were evaluated by Wester n blotting of culture supernatants. Evaluation of cell cycle and proliferat ive activity was performed by flow cytometry. Results. Large Daudi cells in S+G(2)/M phases were L-selectin positive, whe reas small Daudi cells in G(0)/G(1) phase were L-selectin negative. During mitosis, L-selectin was distributed along the cleavage furrow, and graduall y lost. Electron microscopy revealed that separating Daudi cells were negat ive for L-selectin on the entire surface, except minute aggregates of L-sel ectin within the cleavage furrow. Addition of agents known to interfere wit h the ligand-binding portion of L-selectin (sulfatides, MoAbs: Lam1.3 and T Q1) results in loss of L-selectin. Removal of L-selectin by digestion with chymotrypsin inhibits Daudi proliferation. The MoAb FMC46 did not interfere with proliferation. Proliferating Daudi cells produced large quantities of shed L-selectin. Inhibition of Daudi proliferation resulted in levels of s hed L-selectin below the limit of detection. Interpretation and Conclusions. L-selectin is re-distributed on the cell su rface of Daudi cells during the last phase of mitosis, in which plasma memb rane invagination occurs between newly formed daughter cells. Shedding of L -selectin is involved in the cytokinesis of Daudi cells. (C) 1999, Ferrata Storti Foundation.