管状材料对血小板激活作用的体外评价

In vitro evaluation of platelet activation by tubular biomaterials

背景:ISO10993-4及GB/T16886.4中将血液相容性的评价分为5个方面:血栓形成、凝血、血小板、补体、血液学。目前国内较为确定和成熟的体外血液相容体外评价方法有溶血试验、凝血试验及血小板黏附试验,而对血小板激活及补体系统激活方面的研究很少。目的:评价聚乙烯、聚氯乙烯及聚甲基乙烯基硅氧烷3种基础材料管在体外对血小板的激活作用,初步建立一种体外评价管状材料对血小板激活作用的方法。方法:制备聚氯乙烯管、聚乙烯管、硅橡胶管的内径3.7mm,外径5mm,长35cm。每管1mL血液注入聚乙烯、聚氯乙烯、硅橡胶管,管的两端用两通连接,置于恒温培养振荡器中,30°倾斜,接口向上,37℃,140r/min,振荡3.5h。放射免疫法检测材料与血液接触后贫血小板血浆中血小板α颗粒蛋白水平,流式细胞仪检测材料与血液接触后血液中血小板α颗粒蛋白阳性血小板百分率、活化的gpⅡb/Ⅲa复合物阳性血小板百分率。结果与结论:放射免疫法检测结果显示聚乙烯、聚氯乙烯管与血液接触后贫血小板血浆中血小板α颗粒蛋白水平大于硅橡胶管(P<0.05)。聚乙烯和聚氯乙烯管与血液接触后贫血小板血浆中血小板α颗粒蛋白水平差异无显著性意义(P>0.05)。流式细胞术检测结果显示聚乙烯、聚氯乙烯管与血液接触后血液中血小板α颗粒蛋白阳性血小板百分率大于硅橡胶管(P<0.05),聚乙烯管与血液接触后血液中血小板α颗粒蛋白阳性血小板百分率大于聚氯乙烯管(P<0.05)。3种材料与血液接触后血液中活化的gpⅡb/Ⅲa复合物阳性血小板百分率差异无显著性意义(P>0.05)。实验初步建立一种管材料与血液较为合理的接触方式,并可以考虑血浆血小板α颗粒蛋白是较好的反映血小板激活程度的评价指标。用流式细胞术检测血浆血小板α颗粒蛋白阳性血小板百分率更为敏感。

BACKGROUND：In according to ISO-10993-4 and GB/T 16886.4,the in vitro hemo-compatibility evaluation on biomaterials includes thrombosis,coagulation factors,platelets and platelet functions,hematology and complement system.However,in the case of China,the in vitro hemo-compatibility evaluations were performed only thrombosis,coagulation factors and platelet attachment,the investigation on evaluation of platelet and complement activations is less reported.OBJECTIVE：To evaluate the effect of polyethylene,polyvinyl chloride and polymethylvinylsiloxane tubes on platelet activation,and establish a useful method to evaluate the effect of tubular materials on platelet activation.METHODS：Tubes of polyethylene,polyvinyl chlorid and silastic were established by 3.7 mm inner diameter,3.5 mm external diameter,and 35 cm length,respectively.1 mL blood was injected into the tube of polyethylene,polyvinyl chlorid and silastic,respectively.The tubes were connected using a two-way tube,shaken at 140 r/min by 30° sloping for 3.5 hours at 37 ℃.Radioimmunoassay was employed to detect α-granules protein level of platelet poor plasma,while flow cytometry was used to detect the percentage of positive platelet of α-granules protein and that of activated gpⅡb/Ⅲa composite.RESULTS AND CONCLUSION：Radioimmunoassay showed that α-granules protein level of platelet poor plasma in the polyethylene and polyvinyl chlorid tubes was significantly greater than that in the silastic tube（P 0.05）.There were no significant differences in α-granules protein between polyethylene and polyvinyl chlorid（P 0.05）.Flow cytometry indicated that percentage of positive platelet of α-granules protein in the polyethylene and polyvinyl chlorid tubes was significantly greater than that in the silastic tube（P 0.05）;the percentage in the polyethylene tube was significantly greater than that in the polyvinyl chlorid tube（P 0.05）.There was no significant differences in the percentage of positive platelet of activated gpⅡb/Ⅲa composite between the three materials（P 0.05）.A useful blood-material contact model was established,and it was considered that α-granules protein is an available parameter for evaluating platelet activation.The percentage of positive platelet of α-granules protein determined by flow cytometry was a more sensitive parameter for evaluating platelet activation.