 
         
         Summary
         
         Phosphatidylserine (PhtdSer) is an anionic aminophospholipid necessary for the development
            of optimal tissue factor (TF) activity at the cell surface. This study investigates
            the implication of a restricted lipid environment with respect to PhtdSer availability
            on TF expression and activity. K562 cells, showing a reduced ability to externalize
            PhtdSer, were transfected with human TF cDNA. PhtdSer exposure and TF activity were
            examined in transfected cells and compared to monocytic THP-1 cells expressing constitutive
            and inducible TF or megakaryocytic HEL cells showing a high PhtdSer externalization
            potency. TF expression was evidenced by flow cytometry and its activity measured using
            functional assays. PhtdSer exposure was monitored by enzymatic prothrombinase assay.
            One clone (DC9) expressed a stable amount of TF antigen without global modification
            of its membrane status. Despite a noticeable TF expression level, clone DC9 presented
            only a weak TF activity even after ionophore stimulation. The apparent Km
               , relative to factor X (FX) activation by TF-factor VIIa (FVIIa) complex, was 335
            nM versus 70 nM for THP-1 cells. The velocity of the reaction was found 3-fold slower in DC9
            than THP-1 cells. Ionophore treatment resulting in slightly enhanced amounts of available
            PhtdSer abolished this difference. The DC9 clone appears suitable for further investigations
            on the biology of TF expressed at the surface of cells where the contribution of PhtdSer
            is significantly attenuated. Such cells should enable further assessment of the role
            of TF as a receptor coupled to intracellular signaling pathways and its fate during
            apoptotic cell death.
         
         
            Abbreviations: TF: tissue factor, FX: factor X, FVII(a): factor VII(a), PhtdSer: phosphatidylserine,
            PhtdCho: phosphatidylcholine, PhtdEth: phosphatidylethanolamine, BSA: bovine serum
            albumin, dcTF: des-cyto tissue factor (TF1-244: membrane-anchored TF lacking the cytoplasmic tail)
         
         Keywords
Tissue factor - phosphatidylserine - K562 cells - procoagulant activity - microparticle
            release