Sekiya Akiko, Morishita Eriko, Maruyama Keiko, Torishima Hiroki, Ohtake Shigeki
Department of Clinical Laboratory Science, Kanazawa University Graduate School of Medical Science.
J Atheroscler Thromb. 2015 Jul 23;22(7):660-8. doi: 10.5551/jat.28175. Epub 2015 Jan 16.
3-Hydroxy-3-methylglutaryl coenzyme A reductase inhibitors (statins) are cholesterol-lowering drugs with a variety of pleiotropic effects including antithrombotic properties. Tissue factor pathway inhibitor (TFPI), which is produced predominantly in endothelial cells and platelets, inhibits the initiating phase of clot formation. We investigated the effect of fluvastatin on TFPI expression in cultured endothelial cells.
Human umbilical vein endothelial cells (HUVECs) were treated with fluvastatin (0-10μM). The expression of TFPI mRNA and antigen were detected by RT-PCR and western blotting, respectively. The effects of mevalonate intermediates, small GTP-binding inhibitors, and signal transduction inhibitors were also evaluated to identify which pathway was involved. A luciferase reporter assay was performed to evaluate the effect of fluvastatin on TFPI transcription. The stability of TFPI mRNA was estimated by quantitating its levels after actinomycin D treatment.
Fluvastatin increased TFPI mRNA expression and antigen in HUVECs. Fluvastatin-induced TFPI expression was reversed by co-treatment with mevalonate or geranylgeranylpyrophosphate (GGPP). NSC23766 and Y-27632 had no effect on TFPI expression. SB203580, GF109203, and LY294002 reduced fluvastatin-induced TFPI upregulation. Moreover, fluvastatin did not significantly affect TFPI promoter activity. TFPI mRNA degradation in the presence of actinomycin D was delayed by fluvastatin treatment.
Fluvastatin increases endothelial TFPI expression through inhibition of mevalonate-, GGPP-, and Cdc42-dependent signaling pathways, and activation of the p38 MAPK, PI3K, and PKC pathways. This study revealed unknown mechanisms of the anticoagulant effect of statins and gave a new insight to its therapeutic potential for the prevention of thrombotic diseases.
3-羟基-3-甲基戊二酰辅酶A还原酶抑制剂（他汀类药物）是具有多种多效性作用（包括抗血栓形成特性）的降胆固醇药物。组织因子途径抑制剂（TFPI）主要在内皮细胞和血小板中产生，可抑制凝血形成的起始阶段。我们研究了氟伐他汀对培养的内皮细胞中TFPI表达的影响。
用人脐静脉内皮细胞（HUVECs）用氟伐他汀（0 - 10μM）处理。分别通过RT-PCR和蛋白质印迹法检测TFPI mRNA和抗原的表达。还评估了甲羟戊酸中间体、小GTP结合抑制剂和信号转导抑制剂的作用，以确定涉及哪条途径。进行荧光素酶报告基因测定以评估氟伐他汀对TFPI转录的影响。通过在放线菌素D处理后定量其水平来估计TFPI mRNA的稳定性。
氟伐他汀增加了HUVECs中TFPI mRNA的表达和抗原。与甲羟戊酸或香叶基香叶基焦磷酸（GGPP）共同处理可逆转氟伐他汀诱导的TFPI表达。NSC23766和Y-27632对TFPI表达无影响。SB203580、GF109203和LY294002降低了氟伐他汀诱导的TFPI上调。此外，氟伐他汀对TFPI启动子活性没有显著影响。氟伐他汀处理可延迟放线菌素D存在下TFPI mRNA的降解。
氟伐他汀通过抑制甲羟戊酸、GGPP和Cdc42依赖性信号通路以及激活p38 MAPK、PI3K和PKC通路来增加内皮TFPI表达。本研究揭示了他汀类药物抗凝作用的未知机制，并为其预防血栓性疾病的治疗潜力提供了新的见解。
