title: Discordant expression of tissue factor antigen and procoagulant activity on human monocytes activated with LPS and low dose cycloheximide.
abstract: The mechanisms underlying the superinduction of procoagulant activity by cycloheximide (CHX) on LPS-activated human monocytes have been investigated. Tissue factor (TF) activity of intact, viable cells was quantitated with a plasma recalcification assay and assays using chromogenic substrates specific for thrombin and factor Xa (FXa). TF antigen was measured simultaneously by immunocytochemical staining and immunoblotting with an anti-TF monoclonal antibody (MAb). Peripheral blood mononuclear cells (PBMC) activated with LPS in the presence of low dose CHX expressed more TF activity (approx. 100% increase) than cells activated with LPS alone. However, TF antigen levels were decreased approximately 70% by CHX. This discordant relationship was due primarily to differences in rates of activation of factor X (FX); LPS/CHX-treated PBMC activated nearly twice as much FX as LPS-treated cells (2.19 +/- 0.37 versus 1.10 +/- 0.21 ng FXa/10(6) PBMC/min, respectively). These studies indicate that TF cofactor activity on LPS/CHX-treated monocytes was approximately 7 times greater than that present on LPS-treated cells. Increased TF functional activity may be due to CHX-induced alterations in the type and content of phospholipids (PL) in the cell membrane. Results showed that exogenous mixed PL markedly increased TF activity on LPS-activated monocytes, but not on LPS/CHX-activated cells, without increasing TF antigen levels or altering cell viability. Membrane alterations may occur on monocytes in certain pathological or iatrogenic conditions resulting in a highly active form of TF in vivo.
authors.affiliation: Heart Research Institute, Camperdown, N.S.W., Australia.
authors.forename: J D
authors.initials: JD
authors.lastname: Walsh
authors.forename: C L
authors.initials: CL
authors.lastname: Geczy
mesh_terms: D000911:Antibodies, Monoclonal; D001779:Blood Coagulation Factors / Q000032:analysis / Q000096:biosynthesis*; D002478:Cells, Cultured; D003513:Cycloheximide / Q000494:pharmacology*; D004591:Electrophoresis, Polyacrylamide Gel; D005170:Factor X / Q000378:metabolism; D006801:Humans; D015151:Immunoblotting; D007150:Immunohistochemistry; D007700:Kinetics; D007963:Leukocytes, Mononuclear / Q000187:drug effects / Q000502:physiology*; D008070:Lipopolysaccharides; D008213:Lymphocyte Activation; D010743:Phospholipids / Q000494:pharmacology; D013925:Thromboplastin / Q000032:analysis / Q000096:biosynthesis*
chemical_list: D000911:Antibodies, Monoclonal; D001779:Blood Coagulation Factors; D008070:Lipopolysaccharides; D010743:Phospholipids; C045071:leukocyte procoagulant activity; D005170:Factor X; D013925:Thromboplastin; D003513:Cycloheximide