Fucosylated glycosaminoglycan (FG) from sea cucumbers has been reported to have anticoagulant effects via targeting intrinsic tenase. However, FG from natural source also potentially poses risks due to its FXIIa activation and platelet aggregating effects. Here, we found that the effect of FG on human platelet aggregation depended on both the sulfation pattern and chain length. FGs with higher content of Fuc2S4S and larger molecular weight (≥14 kD) had stronger activity. Both platelet aggregation and P-selectin expression induced by TaFG (an FG from Thelenota ananas) were decreased as the molecular weight reduced. Ticagrelor, aspirin and wortmannin completely blocked the secretion (ADP) but only partially blocked the aggregation induced by TaFG. Tirofiban an αIIbβ3 antagonist however potently inhibited both the secretion and aggregation, with IC50 of 6.01 ± 1.1.97 nM. Furthermore, TaFG could bind to human αIIbβ3 with high affinity, and the affinities of two FGs were paralleled with their activity in platelet aggregation or activation. These results indicated that αIIbβ3 played an important role in TaFG-induced platelet aggregation which was mediated by PI3K, and that platelet secretion was required for the amplification of aggregation.
Pulmonary ischemia/reperfusion (IR) injury is the leading cause of acute lung injury, which is mainly attributed to reactive oxygen species (ROS) induced cell injuries and apoptosis. Since rosmarinic acid (RA) has been identified as an antioxidant natural ester, this natural compound might protect against pulmonary IR injury. In this study, the mice were given RA daily (50, 75, or 100 mg/kg) by gavage for 7 days before the pulmonary IR injury. We found that hypoxemia, pulmonary edema, and serum inflammation cytokines were aggravated in pulmonary IR injury. RA pretreatment (75 and 100 mg/kg) effectively reversed these parameters, while 50 mg/kg RA pretreatment was less pronounced. Our data also indicated RA pretreatment mitigated the upregulation of pro-oxidant NADPH oxidases (NOX2 and NOX4) and the downregulation of anti-oxidant superoxide dismutases (SOD1 and SOD2) upon IR injury. In vitro studies showed RA preserved the viability of anoxia/reoxygenation (AR)-treated A549 cells (a human lung epithelial cell line), and the results showed the protective effect of RA started at 5 μM concentration, reached its maximum at 15 μM, and gradually decreased at 20-25 μM. Besides, RA pretreatment (15 μM) greatly reduced the lactate dehydrogenase release levels subjected to AR treatment. Moreover, the results of our research revealed that RA eliminated ROS production and reduced alveolar epithelial cell apoptosis through activating the phosphatidylinositol 3 kinase (PI3K)/protein kinase B (Akt) signaling pathway, which was supported by using wortmannin, because in the presence of wortmannin, the RA-mediated protection was blocked. Meanwhile, wortmannin also reversed the protective effects of RA in mice. Together, our results demonstrate the beneficial role of RA in pulmonary IR injury via PI3K/Akt-mediated anti-oxidation and anti-apoptosis, which could be a promising therapeutic intervention for pulmonary IR injury.
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