Background Metabolic syndrome displays a significant prothrombotic state which renders patients highly susceptible to myocardial infarction, ischaemic stroke and peripheral vascular diseases. Although many clinical studies showed that the prothrombotic state is a key feature of metabolic syndrome, the pathogenesis and mechanisms involved are not completely understood. Metabolic syndrome is often characterised by obesity and subsequent high circulating concentrations of free fatty acids (FFAs). Thrombomodulin (TM)- protein C system is a key endogenous anticoagulant system. We therefore examined the effects of FFAs on the expression of TM- protein C system and the mechanisms involved.
Methods 8 to10 weeks of male wide-type mice were divided into two groups. One group of mice (n=15) were fed a chow diet, while the other group of mice (n=15) were fed a high-fat diet for 20 weeks. Tail-bleeding time and the occlusion time induced by FeCl3 were recorded. Circulating free fatty acids were measured. Primary human aortic endothelial cells (HAECs) were cultured and treated with different dose of palmitic acid (PA). TM expression and protein C activation were measured with western blot and RT-PCR. The effects of JNK, p38 stress pathways and transcriptional factor Foxo1 involved in PA-inhibited TM expression were elucidated with siRNA-induced gene silencing and DNA plasmids-induced gene over-expression.
Results Mice fed a high-fat diet showed notably higher blood level of FFAs (p<0.05), and a shortened tail-bleeding time and a shortened occlusion time induced by FeCl3 (p<0.01). Furthermore, TM was down-regulated in obese mice (p<0.001). PA significantly decreased the expression of TM and protein C activation in a dose-dependent fashion (p<0.01). Silencing JNK and p38 with specific siRNAs decreased PA-induced thrombomodulin suppression (p<0.01), while wide type of JNK plasmid can enhance PA's inhibitory effect on TM expression (p<0.05). When endothelial cells were transiently transfected with Foxo1–specific siRNAs, PA-induced inhibition of TM expression was significantly decreased (p<0.001). Furthermore, wide type and constitutively active of Foxo1 plasmids enhanced PA's inhibitory effect of TM expression (p<0.05).
Conclusion In summary, a high-fat diet induced high elevated circulating FFAs level, and the prothrombotic state in mice. PA inhibited the expression of TM and the activation of protein C in endothelial cells. JNK, p38/Foxo1 pathway mediated free fat acids' inhibitory effect of TM expression, which may be a new therapeutic target in treating and reducing cardiovascular and cerebrovascular complications of the metabolic syndrome.