In the present study, the signal pathways involved in NO formation and iNOS expression in RAW 264.7 macrophages stimulated by LTA were investigated. We also compared the relative inhibitory ...activities and mechanisms of PMC, a novel potent antioxidant of α-tocopherol derivatives, with those of YC-1, an sGC activator, on the induction of iNOS expression by LTA in cultured macrophages
in vitro and LTA-induced hypotension
in vivo. LTA induced concentration (0.1–50
μg/mL)- and time (4–24
hr)-dependent increases in nitrite (an indicator of NO biosynthesis) in macrophages. Both PMC (50
μM) and YC-1 (10
μM) inhibited NO production, iNOS protein, mRNA expression, and IκBα degradation upon stimulation by LTA (20
μg/mL) in macrophages. On the other hand, PMC (50
μM) almost completely suppressed JNK/SAPK activation, whereas YC-1 (10
μM) only partially inhibited its activation in LTA-stimulated macrophages. Moreover, PMC (10
mg/kg, i.v.) and YC-1 (5
mg/kg, i.v.) significantly inhibited the fall in MAP stimulated by LTA (10
mg/kg, i.v.) in rats. In conclusion, we demonstrate that YC-1 shows more-potent activity than PMC at abrogating the expression of iNOS in macrophages
in vitro and reversing delayed hypotension in rats with endotoxic shock stimulated by LTA. The inhibitory mechanisms of PMC may be due to its antioxidative properties, with a resulting influence on JNK/SAPK and NF-κB activations. YC-1 may be mediated by increasing cyclic GMP, followed by, at least partly, inhibition of JNK/SAPK and NF-κB activations, thereby leading to inhibition of iNOS expression.
The intracellular mechanisms underlying oxidized low density lipoprotein (oxLDL)‐signaling pathways in platelets remain obscure and findings have been controversial. Therefore, we examined the ...influence of oxLDL in washed human platelets. In this study oxLDL concentration‐dependently (20–100 μg/mL) inhibited platelet aggregation in human platelets stimulated by collagen (1 μg/mL) and arachidonic acid (60 μM), but not by thrombin (0.02 U/mL). The activity of oxLDL was greater at 24 h in inhibiting platelet aggregation than at 12 h. At 24 h, oxLDL concentration‐dependently inhibited intracellular Ca2+ mobilization and thromboxane B2 formation in human platelets stimulated by collagen. In addition, at 24 h oxLDL (40 and 80 μg/mL) significantly increased the formation of cyclic AMP, but not cyclic GMP or nitrate. In an ESR study, 24 h‐oxLDL (40 μg/mL) markedly reduced the ESR signal intensity of hydroxyl radicals (OH−) in both collagen (2 μg/mL)‐activated platelets and Fenton reaction (H2O2+Fe2+). The inhibitory effect of oxLDL may induce radical‐radical termination reactions by oxLDL‐derived lipid radical interactions with free radicals (such as hydroxyl radicals) released from activated platelets, with a resultant lowering of intracellular Ca2+ mobilization followed by inhibition of thromboxane A2 formation, thereby leading to increased cyclic AMP formation and finally inhibited platelet aggregation. This study provides new insights concerning the effect of oxLDL in platelet aggregation.
The amyloid beta peptide (Abeta), a mediator of neuronal and vascular degeneration in the pathogenesis of Alzheimer's disease and cerebral amyloid angiopathy may have peripheral actions. Platelets ...are enriched with Abeta and have been shown to enhance platelet actions. However, the detailed signaling pathways through which Abeta activates platelets have not been previously explored. In this study, we examined the intra-platelet Abeta distribution using a gold labeling technique and noted that Abeta was predominantly localized in the cytoplasm of resting platelets. A marked increase in Abeta-gold labeling in an open canalicular system was observed in collagen-activated platelets. Exogenous Abeta (2-10 microM) stimulated platelet aggregation accompanied by phospholipase Cgamma2 (PLCgamma2) phosphorylation, phosphoinositide breakdown, and Ca(2+)i mobilization as well as protein kinase C (PKC) activation. Ro318220, an inhibitor of PKC, suppressed Abeta-induced platelet aggregation, PKC activation, and Ca(2+)i mobilization in platelets, suggesting that the PLCgamma2-PKC pathway is involved in Abeta-induced platelet aggregation. In the electron spin resonance study, Abeta (2 and 10 microM) markedly triggered hydroxyl radical formation in platelets. In an in vivo study, Abeta (2mg/kg) significantly shortened the latency for inducing platelet plug formation in the mesenteric venules of mice. In conclusion, we are the first to demonstrate (1) the distribution of Abeta in human platelets; and that (2) Abeta activation of platelets is mediated, at least partially, by the PLCgamma2-PKC pathway; and (3) Abeta triggers thrombus formation in vivo.
Seven children with spastic diplegic cerebral palsy and 14 age- and gender-matched nondisabled children participated in the present study for an investigation and comparison of their static standing ...balance under altered sensory environments. The type of visual input (full, occluded, or sway referenced vision) and the type of somatosensory input (fixed or compliant foot support) were varied factorially to give six sensory environments. Each participant was tested barefooted for 30 s under all six conditions. A force platform collected the ground reaction force, from which standing balance was calculated as the sway area of the center of pressure. The results showed that when somatosensory information was reliable (fixed foot support), there was no significant difference in stance stability between the children with spastic diplegic cerebral palsy and their matched controls, and both types of children were equally affected by the type of visual input. However, when somatosensory information was unreliable (compliant foot support), the difference in stance stability between the children with spastic diplegic cerebral palsy and their matched controls was significantly greater when the visual input was deprived (occluded) or unreliable (sway referenced) than when it was reliable. These results suggest that the children with spastic diplegic cerebral palsy may have difficulties in resolving intersensory conflicts for maintenance of standing balance, or the demands of motor control in sensory conflict conditions outweigh the motor ability of children with spastic diplegic cerebral palsy.
Lycopene is a natural carotenoid antioxidant that is present in tomatoes and tomato products. The pharmacologic function of lycopene in platelets is not yet understood. Therefore, in this study we ...sought to systematically examine the effects of lycopene in the prevention of platelet aggregation and thrombus formation. We found that lycopene concentration-dependently (2–12 μmol/L) inhibited platelet aggregation in human platelets stimulated by agonists. Lycopene (6 and 12 μmol/L) inhibited phosphoinositide breakdown in platelets labeled with tritiated inositol, intracellular Ca
+2 mobilization in Fura-2 AM–loaded platelets, and thromboxane B
2 formation stimulated by collagen. In addition, lycopene (6 and 12 μmol/L) significantly increased the formations of cyclic GMP and nitrate but not cyclic AMP in human platelets. Rapid phosphorylation of a protein of 47,000 Da (P47), a marker of protein kinase C activation, was triggered by PDBu (60 nmol/L). This phosphorylation was markedly inhibited by lycopene (12 μmol/L) in phosphorus-32–labeled platelets. In an in vivo study, thrombus formation was induced by irradiation of mesenteric venules in mice pretreated with fluorescein sodium. Lycopene (5, 10, and 20 mg/kg) significantly prolonged the latency period for the induction of platelet-plug formation in mesenteric venules. These results indicate that the antiplatelet activity of lycopene may involve the following pathways: (1) Lycopene may inhibit the activation of phospholipase C, followed by inhibition of phosphoinositide breakdown and thromboxane B
2 formation, thereby leading to inhibition of intracellular Ca
+2 mobilization. (2) Lycopene also activated the formations of cyclic GMP/nitrate in human platelets, resulting in the inhibition of platelet aggregation. The results may imply that tomato-based foods are especially beneficial in the prevention of platelet aggregation and thrombosis.
