麒麟菜多肽对血小板聚集及角叉菜胶诱导的小鼠尾部血栓形成的影响

Effects of Eucheuma peptides on platelet aggregation and mouse tail thrombosis induced by carrageenan

目的探讨麒麟菜多肽抗血小板聚集的效果,以及其对角叉菜胶诱导的小鼠尾部血栓形成的影响。方法采用光学比浊法检测不同剂量的麒麟菜多肽对二磷酸腺苷(ADP)和凝血酶诱导的血小板聚集的影响,该体外实验分4组,采用生理盐水或麒麟菜多肽2.5、5.0、10.0μg/ml处理血小板,采用FLASH连续光谱荧光酶标仪动态监测血小板聚集情况,计算血小板最大聚集率(MAR)。体内实验以角叉菜胶腹腔注射复制小鼠尾部血栓模型,采用麒麟菜多肽15,30,60mg/kg和氯吡格雷100mg/kg灌胃处理,连续7d。于末次给药后1h采用角叉菜胶诱发血栓形成,24h后用游标卡尺测量小鼠尾部长度及尾部血栓形成长度,计算血栓长度占尾部总长度的百分比(%)。结果体外实验中,光学比浊法证实课题组研究提取的麒麟菜多肽可抑制ADP和凝血酶诱导的血小板聚集。ADP诱导的血小板MAR在生理盐水处理组为100%,在麒麟菜多肽低、中、高剂量组分别为61.84%、71.04%、60.95%,与生理盐水组比较差异有统计学意义(P<0.01);凝血酶诱导的血小板MAR在上述4组分别为100%、75.34%、69.96%、73.74%,与生理盐水组比较差异有统计学意义(P<0.01)。体内实验中,麒麟菜多肽低、中、高剂量组和氯吡格雷组对角叉菜胶诱导的小鼠尾部血栓形成的抑制率分别为36.89%、17.25%、22.99%、18.96%,与生理盐水组比较差异均有统计学意义(P<0.01)。结论麒麟菜多肽可抑制角叉菜胶诱发的小鼠尾部血栓形成,具有防治血栓形成的潜在功能。

Objective This study is designed to investigate the effect of the peptide from Eucheuma on platelet aggregation and mouse tail thrombosis induced by carrageenan. Methods The peptide was purified from Eucheuma using sephdexG-50 gel chromatography and HPLC. The inhibitory effects of peptide on platelet aggregation induced by adenosine diphosphate （ADP）/ thrombin were detected by light transmittance aggregometry. The platelet were treated with normal saline or Eucheuma peptides （2.5, 5.0, 10.0μg/ml）. The platelet aggregation was detected by FLASH continuous spectral fluorescence microplate reader, and the maximum platelet aggregation rate （MAR） was calculated. The peptide （15, 30, 60mg/kg） and 100mg/kg clopidogrel were intraperitoneal injected into mice for 7 days to confirm the anti-thrombosis activity of the peptide in carrageenan-induced mouse tail thrombosis model. Results The platelet aggregation can be inhibited by the peptide. Comparing the maximum platelet aggregation rate induced by ADP, the results showed that control group was 100%, 2.5μg/ml peptide group was 61.84%, 5.0μg/ml peptide group was 71.04% and 10.0p.g/ml peptide was 60.95%. The maximum platelet aggregation rate induced by thrombin was also tested. The results showed that control group was 100%, 2.5μg/ml peptide group was 75.34%, 5.0μg/ml peptide group was 69.96% and 10.0μg/ ml peptide group was 73.74%. The thrombosis rate （2.5μg/ml peptide group was 36.89%, 5.0μg/ml peptide group was 17.25%,10.0μg/ml peptide group was 22.99%, and 100mg/kg clopidogrel group was 18.96%, respectively） and the length ofthrombus in mice can be reduced by the peptide （15-60mg/kg）. Conclusions The carrageenan induced tail thrombosis in mice can be inhibited by the peptide of Eucheuma. It is suggested that the peptide of Eucheuma is a potential compound in anti-thrombosis.