Soluble Thrombomodulin Antigen in Conditioned Medium Is Increased by Damage of Endothelial Cells

Abstract

SummaryThrombomodulin (TM) exists not only in endothelial cells but also in circulating plasma as soluble heterogeneous fragments. A release mechanism of soluble TM antigen from endothelial cells was investigated. Cultured human umbilical vein endothelial cells released about 0.6% of total cellular TM antigen into conditioned medium during 24 h. The release of TM antigen was not influenced by addition of various concentrations (0.01-5.0 μM) of monensin, which inhibits intracellular transport of secretory proteins, though the secretion of plasminogen activator inhibitor-1 from the cells was inhibited. The release of TM antigen was not increased when total cellular TM level increased 1.3- or 1.4-fold relative to control cells after stimulation with 0.1-1.0 U/ml thrombin or 3 mM dibutyryl cAMP, respectively. Exposure of endothelial cells for 6 h to mixture of 1 μM N-formyl-methionyl-leucyl-phenylalanine (FMLP) and 100 ng/ml lipopolysaccharide (LPS) decreased cellular TM level by 30% relative to control cells without increase in the TM release. The FMLP and LPS-stimulated leukocyte treatment of the cells increased the release of TM antigens into the medium in a time-dependent manner and the increased release of TM antigen paralleled the extent of cell damage as measured by 51Cr release. Hydrogen peroxide treatment of the cells increased the release of TM antigens into the medium in a time- and concentration-dependent manner. The increased release of TM antigen by hydrogen peroxide also paralleled the extent of cell damage. Soluble TM antigen in conditioned medium from the untreated control cells was composed of six heterogeneous fragments of 105, 56, 48, 33, 31 and 28 kDa observed on Western blots after sodium dodecyl sulfatepolyacrylamide gel electrophoresis under reducing conditions, and the soluble TM antigen from the hydrogen peroxide-treat-ment cells was composed of five heterogeneous fragments of 80, 56, 33, 31 and 28 kDa. These compositions were quite similar to that observed for soluble TM fragments in human plasma. It is suggested that soluble TM antigen is not secreted from endothelial cells, but results from cellular damage.

Polymyalgia Rheumatica and Type 2 Diabetes Mellitus Complicated with Electromyographic Abnormalities That Responded Well to Corticosteroid Therapy.

“…They suggested that the pathophysiology of neuropathy might not be due to systemic vasculitis, but rather to microvasculitis, causing ischemia in the vasa nervorum around peripheral nerves. Thrombomodulinis thought to be a good indicator of vascular injury (9,10). In this case, elevated levels of platelet and thrombomodulin before treatment with PSLwere also found.…”

Diagnostic value of hemostatic parameters in bone marrow transplant-associated thrombotic microangiopathy

“…TM is a high-affinity thrombin receptor on the surface of vascular endothelial cells and a cofactor in the activation of protein C. 23 There is a good correlation between TM levels and clinical complications related to endothelial cell injury in septicemia, VOD and GVHD. 16 Increased TM levels in patients with BMT-TMA have been reported by Zeigler et al 15 They tested a ratio of TM:creatinine to provide a measure of the renal contribution to the TM elevation.…”

Change in von Willebrand factor and carotid intima-media thickness in hypothyroid patients with normal thyroid function after levothyroxine replacement therapy

“…vWF was measured as described previously (28,29). Serum TM was determined by a one-step sandwich enzyme immunoassay, the reported sensitivity of which is 1 mg/l for soluble TM (30,31). Total cholesterol, triglyceride and high-density lipoprotein (HDL)-cholesterol were determined using an autoanalyzer.…”