Chien-Huang Lin

Reduction in Lipopolysaccharide-Induced Thrombocytopenia by Triflavin in a Rat Model of Septicemia

BACKGROUNDnThrombocytopenia frequently occurs early in the course of Gram-negative bacterial infections. Triflavin, an Arg-Gly-Asp-containing disintegrin, has been suggested to interfere with the interaction of fibrinogen with the glycoprotein IIb/IIIa complex. The present study was undertaken to determine whether triflavin could prevent thrombocytopenia in lipopolysaccharide (LPS)-treated rats.nnnMETHODS AND RESULTSnIn this study, 51Cr-labeled platelets were used to assess blood and tissue platelet accumulation after LPS challenge. The administration of LPS (4 mg/kg IV bolus) for 4 hours induced a reduction in radiolabeled platelets in blood and an obvious accumulation of platelets in liver. Triflavin (500 microg/kg) but not GRGDS (20 mg/kg) significantly prevented the alteration of radiolabeled platelet distribution in blood and liver when induced by LPS. Furthermore, triflavin but not GRGDS markedly suppressed the elevation in plasma thromboxane B2 concentration within the 4-hour period of LPS administration. In LPS-treated rats, the 5-hydroxytryptamine level was lower in the blood and higher in the liver compared with levels in normal saline-treated rats. Pretreatment with triflavin (500 microg/kg) significantly reversed the 5-hydroxytryptamine concentration in blood and liver of LPS-treated rats. In histological examinations and platelet adhesion assay, triflavin markedly inhibited the adhesion of platelets to subendothelial matrixes in vivo and in vitro.nnnCONCLUSIONSnThe results indicate that triflavin effectively prevents thrombocytopenia, possibly through the following 2 mechanisms: (1) Triflavin markedly inhibits platelet aggregation, resulting in decreased thromboxane A2 formation. (2) It inhibits the adhesion of platelets to subendothelial matrixes, thereby leading to a reversal in the distribution of platelets in blood and liver in LPS-treated rats.

Induction of cyclooxygenase‐2 protein by lipoteichoic acid from Staphylococcus aureus in human pulmonary epithelial cells: involvement of a nuclear factor‐κB‐dependent pathway

This study investigated the role of protein kinase C (PKC) and transcription factor nuclear factor‐κB (NF‐κB) in cyclooxygenase‐2 (COX‐2) expression caused by lipoteichoic acid (LTA), a cell wall component of the gram‐positive bacterium Staphylococcus aureus, in human pulmonary epithelial cell line (A549). LTA caused dose‐ and time‐dependent increases in COX‐2 expression and COX activity, and a dose‐dependent increase in PGE2 release in A549 cells. The LTA‐induced increases in COX‐2 expression and COX activity were markedly inhibited by dexamethasone, actinomycin D or cyclohexamide, but not by polymyxin B, which binds and inactivates endotoxin. The phosphatidylcholine‐phospholipase C (PC‐PLC) inhibitor (D‐609) and the phosphatidate phosphohydrolase inhibitor (propranolol) reduced the LTA‐induced increases in COX‐2 expression and COX activity, while phosphatidylinositol‐phospholipase C inhibitor (U‐73122) had no effect. The PKC inhibitors (Go 6976, Ro 31‐8220 and GF 109203X) and NF‐κB inhibitor, pyrrolidine dithiocarbamate (PDTC), also attenuated the LTA‐induced increases in COX‐2 expression and COX activity. Treatment of A549 cells with LTA caused an increase in PKC activity in the plasma membrane; this stimulatory effect was inhibited by D‐609, propranolol, or Go 6976, but not by U‐73122. Exposure of A549 cells to LTA caused a translocation of p65 NF‐κB from the cytosol to the nucleus and a degradation of IκB‐α in the cytosol. Treatment of A549 cells with LTA caused NF‐κB activation by detecting the formation of NF‐κB‐specific DNA‐protein complex in the nucleus; this effect was inhibited by dexamethasone, D‐609, propranolol, Go 6976, Ro 31‐8220, or PDTC. These results suggest that LTA might activate PC‐PLC and phosphatidylcholine‐phospholipase D to induce PKC activation, which in turn initiates NF‐κB activation, and finally induces COX‐2 expression and PGE2 release in human pulmonary epithelial cell line.

Lipoteichoic acid-induced cyclooxygenase-2 expression requires activations of p44/42 and p38 mitogen-activated protein kinase signal pathways

This study investigated the role of p44/42 and p38 mitogen-activated protein kinase (MAPK) in cyclooxygenase-2 expression caused by lipoteichoic acid in human pulmonary epithelial cell line (A549). Lipoteichoic acid-induced increases in cyclooxygenase activity and cyclooxygenase-2 expression were attenuated by tyrosine kinase inhibitors (genistein and tyrphostin AG126), a MAPK/extracellular signal-regulated protein kinase (MEK) inhibitor [2-amino-3-methoxyflavone] (PD 98059) and a p38 MAPK inhibitor [4-(4-fluorophenyl)-2-(4-methylsulfinylphenyl)-5-(4-pyridyl)1H-imidazole] (SB 203580). Lipoteichoic acid-induced p44/42 MAPK activation was inhibited by protein kinase C (PKC) inhibitors [12-(2-cyanoethyl)6,7,12,13-tetrahydro-13-methyl-5-oxo-5H-indolo(2,3-a)pyrrolo(3,4-c)-carbazole] (Go 6976) and [3-[1-[3-(amidinothio)propyl-1H-indol-3-yl]-3-(1-methyl-1H-indol-3-yl)maleimide]] (Ro 31-8220), genistein and PD 98059. Lipoteichoic acid-induced increase in p38 MAPK activity was inhibited by Go 6976, Ro 31-8220, genistein and SB 203580. Lipoteichoic acid-mediated formation of nuclear factor-kappa B (NF-kappa B)-specific DNA-protein complex was inhibited by genistein, tyrphostin AG126, PD 98059 and SB 203580. These results suggest that the activations of both p44/42 and p38 MAPK by lipoteichoic acid result in stimulation of NF-kappa B-specific DNA-protein binding and subsequent cyclooxygenase-2 expression in A549 cells. Both events required activation of upstream tyrosine kinase and PKC.

Inhibition of platelet thromboxane formation and phosphoinositides breakdown by diisoeugenol

Abstract— Diisoeugenol inhibited the platelet aggregation and ATP release of rabbit platelets caused by ADP, arachidonic acid, platelet‐activating factor (PAF), collagen and thrombin. Prolongation of the incubation time of platelets with diisoeugenol did not cause further inhibition and the aggregability of platelets could not be restored after washing. In human platelet‐rich plasma, diisoeugenol inhibited the biphasic aggregation and ATP release induced by adrenaline and ADP in a concentration‐dependent manner. Thromboxane B2 formation caused by arachidonic acid, collagen and thrombin was markedly inhibited by diisoeugenol in a concentration‐dependent manner. Diisoeugenol also inhibited the formation of inositol monophosphate caused by collagen, PAF and thrombin. The cAMP level of washed platelets was not changed by diisoeugenol. It is concluded that the antiplatelet effect of diisoeugenol is due to the inhibition of thromboxane formation and phosphoinositides breakdown.