Multidrug resistance protein 4 (MRP4/ABCC4) regulates thrombus formation in vitro and in vivo
Multidrug resistance protein 4 (MRP4/ABCC4) regulates thrombus formation in vitro and in vivo
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DOI：
10.1016/j.ejphar.2014.05.001
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发表时间：
2014-08-15
影响因子：
5
通讯作者：
Lin, Kuan-Hung
中科院分区：
文献类型：
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作者：
Lien, Li-Ming;Chen, Zhih-Cherng;Lin, Kuan-Hung
多药耐药蛋白4（multidrug resistance protein 4，MRP 4）是三磷酸腺苷结合盒转运蛋白ABCC亚家族的成员之一，其功能是从血小板上清除环核苷酸并将ADP摄取到血小板致密颗粒中。然而，MRP 4是否直接参与血小板活化仍不清楚。因此，我们研究的目的是确定血小板活化中MRP 4调节的详细机制。我们的研究结果显示，MRP 4抑制剂MK 571抑制胶原诱导的血小板聚集，PKA抑制剂H89部分逆转了这种聚集，但腺苷酸环化酶（AC）抑制剂SQ 22536和鸟苷酸环化酶（GC）抑制剂ODQ不能逆转这种聚集，这表明MK 571可以通过不依赖于环核苷酸产生的途径来防止胶原诱导的聚集。在本研究中，我们发现MK 571抑制胶原诱导的ATP释放和钙动员。MK 571还能抑制蛋白激酶C、JNK和AkL的磷酸化，电子自旋共振实验表明MK 571能显著减少羟自由基的形成。此外，MK 571延迟PEA-100装置的体外血小板栓形成，并延迟荧光素钠照射的小鼠肠系膜微静脉中的血栓形成。然而，先前的研究报告MK 571也阻断MRP 1和白三烯D4（LTD 1）受体。因此，MK 571是否通过MRP 1或LTD 4受体抑制血小板活化需要考虑和进一步确定。总之，除了阻断环核苷酸的转运外，MRP 4抑制还可以防止体外和体内血栓形成。我们的研究结果还支持这样的想法，即MRP 4可能代表用于治疗血栓栓塞性疾病的开发或新型治疗干预的潜在靶点。(C)2014 Elsevier By. All rights reserved.
The multidrug resistance protein 4 (MRP4) is a member of the ABCC subfamily of the adenosine triphosphate-binding cassette transporters that remove cyclic nucleotides from platelets and uptake ADP into dense granule in platelets. However, whether MRP4 directly involves platelet activation remains unclear. Thus, the aim of our study was to determine the detailed mechanisms underlying the regulation at MRP4 in platelet activation. Our results revealed that the MRP4 inhibitor MK571 inhibited collagen induced platelet aggregation which was partially reversed by the PKA inhibitor H89, but not by the adenylyl cyclase (AC) inhibitor SQ22536 and the guanylyl cyclase (GC) inhibitor ODQ suggesting that MK571 can prevent collagen -induced aggregation via a route independent of cyclic nucleotide production. In the present study, we found that MK571 inhibited collagen induced ATP release and calcium mobilization. The phosphorylaLion of protein kinase C, JNK, and AkL was also inhibited by MK571, and electron spin resonance experiment showed that MK571 significantly reduced hydroxyl radical formation. Moreover, MK571 delayed platelet plug formation in vitro by a PEA -100 device, and delayed thrombus formation in mesenteric venules of mice irradiated by fluorescein sodium. However, previous studies have reported that MK571 also blocks MRP1 and leukorriene D4 (LTD1) receptor. Therefore, whether MK571 inhibits platelet activation through MRP1 or LTD4 receptor needs to be considered and further defined. In conclusion, in addition to blocking the transport of cyclic nucleotides, MRP4 inhibition may prevent thrombus formation in vitro and in vivo. Our findings also support the idea that MRP4 may represent a potential target for the development or novel therapeutic interventions for the treatment of thromboembolic disorders. (C) 2014 Elsevier By. All rights reserved.