Summary
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/106 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.
