CN113304327A - 一种抗凝血的密网支架及其制备方法 - Google Patents
一种抗凝血的密网支架及其制备方法 Download PDFInfo
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- CN113304327A CN113304327A CN202110578068.8A CN202110578068A CN113304327A CN 113304327 A CN113304327 A CN 113304327A CN 202110578068 A CN202110578068 A CN 202110578068A CN 113304327 A CN113304327 A CN 113304327A
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- Media Introduction/Drainage Providing Device (AREA)
Abstract
本发明公开了一种抗凝血的密网支架及其制备方法，支架包括密网支架基底以及键接于密网支架基底表面的有机小分子涂层，有机小分子涂层的厚度为5～500nm。在制备该支架时，先对密网支架基底进行清洗，然后采用处理液对密网支架基底进行处理，在其表面形成一层二氧化钛基层，再用紫外线对密网支架基底进行照射，最后将经过紫外照射的密网支架基底浸没于有机小分子酸溶液中，反应一定时间后即得。本发明针对密网支架的抗凝要求构建了二氧化钛‑柠檬酸涂层，该涂层能使密网支架在体内服役期间持续抗凝，从而减少患者术后抗凝治疗剂量，降低其经济压力与心理负担。
Description
技术领域
本发明属于医疗器械植入物技术领域，具体涉及一种抗凝血的密网支架及其制备方法。
背景技术
动脉瘤是由于动脉壁的病变或损伤而导致的一种膨胀性疾病，是一种常见的血管疾病。若没有及时治疗，动脉瘤随时都可能会破裂，威胁到患者的生命健康。目前，针对动脉瘤的治疗方法主要有三种：传统的外科夹闭手术、弹簧圈栓塞治疗、病变处植入密网支架或覆膜支架。外科手术存在一定的难度，创伤大且伴有并发症，一般不为患者和医生选择。弹簧圈栓塞术对大动脉瘤、宽口动脉瘤、假性动脉瘤、复发性动脉瘤的治疗风险高、效果差。通过密网支架的植入对病变动脉瘤处的血管重建，能够引导血液沿正常路径流动，改变瘤内的血流动力学，大大降低血管瘤破裂的风险；同时，密网支架能够减少血管瘤内的血供，血管瘤会逐渐萎缩变小，回到一个正常的水平。相比于外科夹闭手术与弹簧圈栓塞术具有安全性更高、适用范围更广、操作更简单等优势。
然而，目前使用的密网支架也存在缺点和不足：1.支架植入后对血管壁的挤压易导致血管内膜受损，进而诱发血栓造成再狭窄甚至闭塞；2.支架与血液的大面积接触导致表面蛋白等的粘附，造成血小板的粘附激活，形成血栓；3.支架长期放置导致血管内皮化延缓以及产生炎症等并发症。目前的解决方案主要是通过长期服用抗凝药物进行治疗，但这不仅会增加大出血的风险，而且对患者而言是巨大的心理压力和经济负担。
发明内容
针对上述现有技术，本发明提供一种抗凝血的密网支架及其制备方法，以解决现有密网支架植入后对血管壁的挤压易导致血管内膜受损，进而诱发血栓造成再狭窄甚至闭塞的问题。
为了达到上述目的，本发明所采用的技术方案是：提供一种抗凝血的密网支架及其制备方法，本发明中的密网支架包括密网支架基底以及键接于密网支架基底表面并具有抗凝血功能的有机小分子酸，有机小分子酸在密网支架基底表面形成厚度为5～500nm的涂层。
在上述技术方案的基础上，本发明还可以做如下改进。
进一步，密网支架基底为镍钛合金密网支架。
进一步，有机小分子酸为柠檬酸、单宁酸、乙酰水杨酸或没食子酸。
本发明中的抗凝血的密网支架经过以下步骤制得：
S1：对密网支架基底进行清洗并干燥；
S2：将经过S1处理后的密网支架基底浸没于温度为45～55℃的处理液中，并用氢氟酸将处理液的pH调节至3.5～4.0，浸泡0.5～4h后取出密网支架基底；所述处理液由氟钛酸铵溶液和硼酸溶液混合而成，其中氟钛酸铵的浓度为0.05～0.2M，硼酸的浓度为0.2～0.5M；
S3：将经过S2处理后的密网支架基底置于紫外下照射2～6h；
S4：将经过紫外照射后的密网支架基底浸没于浓度为1～3mg/mL有机小分子酸溶液中，于37～45℃下反应15～60min，取出，清洗并干燥，即得。
本发明的机理是：通过化学转化法对镍钛合金密网支架进行处理，在支架表面得到一层二氧化钛基层，再通过紫外照射增加其表面活性羟基，以利于后续柠檬酸等有机小分子的组装和沉积。基于自组装和共价接枝的原理，柠檬酸等有机小分子和二氧化钛表面的活性羟基进行共价接枝，形成具有纳米尺度的有机小分子功能涂层。
抗凝血的密网支架的制备方法在上述技术方案的基础上还可以做如下进一步的改进。
进一步，将密网支架基底置于去离子水中，超声清洗三次，每次5min；然后置于无水乙醇中，超声清洗三次，每次5min；再置于丙酮中，超声清洗三次，每次5min；最后真空干燥箱干燥。
进一步，处理液的温度为50℃，pH为3.88，密网支架基底在处理液中的浸泡时间为1h。
进一步，处理液中氟钛酸铵的浓度为0.1M，硼酸的浓度为0.3M。
进一步，S3中紫外照射时间为4h。
进一步，有机小分子酸为柠檬酸、单宁酸、乙酰水杨酸或没食子酸。
进一步，S4中反应温度为40℃，反应时间为30min。
本发明的有益效果是：
1.本发明构筑了纳米尺度的改性功能层，包含应用广泛的二氧化钛，天然有机小分子，并基于自组装和共价接枝的原理，将两种组分结合在一起，形成了纳米尺度涂层；并且，区别于传统载药涂层体系，本发明中通过共价接枝固定在表面的有机小分子涂层更加稳定牢固，可以实现表面长效功能化，并不涉及药物释放动力学，因此更加简单有效。
2.本发明中采用柠檬酸等有机小分子酸在密网支架基底表面形成涂层，不仅能够抑制血小板的粘附和激活，具有良好的抗凝效果，而且小分子涂层相较于高聚物涂层分子量更小，不会降解产生有害的低聚物等，再者，小分子形成的涂层更薄，也更容易对厚度进行控制。
附图说明
图1为镍钛合金密网支架基底键接柠檬酸涂层前后的扫描电镜形貌图；
图2为镍钛合金密网支架基底键接柠檬酸涂层后的红外光谱图；
图3为镍钛金属片及镍钛合金密网支架基底键接柠檬酸涂层前后亲水性演示结果；
图4为镍钛金属片及镍钛合金密网支架基底键接柠檬酸涂层前后内皮细胞粘附荧光染色图；
图5为镍钛合金密网支架基底键接柠檬酸涂层前后内皮细胞迁移结果。
图6为镍钛合金密网支架基底键接柠檬酸涂层前后溶血结果。
具体实施方式
下面结合实施例对本发明的具体实施方式做详细的说明。
实施例1
一种抗凝血的密网支架，包括镍钛合金密网支架基底以及键接于镍钛合金密网支架基底表面的柠檬酸涂层，柠檬酸涂层的厚度为300nm左右。
本实施例中的促内皮化的密网支架经过以下步骤制得：
S1：将密网支架基底置于去离子水中，超声清洗三次，每次5min；然后置于无水乙醇中，超声清洗三次，每次5min；再置于丙酮中，超声清洗三次，每次5min；最后真空干燥箱干燥；
S2：将经过S1处理后的镍钛合金密网支架基底浸没于温度为50℃的处理液中，并用氢氟酸将处理液的pH调节至3.88，浸泡2h后取出密网支架基底；处理液由氟钛酸铵溶液和硼酸溶液混合而成，其中氟钛酸铵的浓度为0.1M，硼酸的浓度为0.3M；
S3：将经过S2处理后的密网支架基底置于紫外下照射4h；
S4：将经过紫外照射后的密网支架基底浸没于浓度为2mg/mL柠檬酸溶液中，于40℃下反应30min，取出，清洗并干燥，即得。
实施例2
一种抗凝血的密网支架，包括镍钛合金密网支架基底以及键接于镍钛合金密网支架基底表面的乙酰水杨酸涂层，乙酰水杨酸涂层的厚度为500nm左右。
本实施例中的促内皮化的密网支架经过以下步骤制得：
S1：将密网支架基底置于去离子水中，超声清洗三次，每次5min；然后置于无水乙醇中，超声清洗三次，每次5min；再置于丙酮中，超声清洗三次，每次5min；最后真空干燥箱干燥；
S2：将经过S1处理后的镍钛合金密网支架基底浸没于温度为45℃的处理液中，并用氢氟酸将处理液的pH调节至3.5，浸泡4h后取出密网支架基底；处理液由氟钛酸铵溶液和硼酸溶液混合而成，其中氟钛酸铵的浓度为0.05M，硼酸的浓度为0.5M；
S3：将经过S2处理后的密网支架基底置于紫外下照射6h；
S4：将经过紫外照射后的密网支架基底浸没于浓度为1mg/mL柠檬酸溶液中，于37℃下反应60min，取出，清洗并干燥，即得。
实施例3
一种抗凝血的密网支架，包括镍钛合金密网支架基底以及键接于镍钛合金密网支架基底表面的单宁酸涂层，单宁酸涂层的厚度为100nm左右。
本实施例中的促内皮化的密网支架经过以下步骤制得：
S1：将密网支架基底置于去离子水中，超声清洗三次，每次5min；然后置于无水乙醇中，超声清洗三次，每次5min；再置于丙酮中，超声清洗三次，每次5min；最后真空干燥箱干燥；
S2：将经过S1处理后的镍钛合金密网支架基底浸没于温度为55℃的处理液中，并用氢氟酸将处理液的pH调节至4.0，浸泡0.5h后取出密网支架基底；处理液由氟钛酸铵溶液和硼酸溶液混合而成，其中氟钛酸铵的浓度为0.2M，硼酸的浓度为0.2M；
S3：将经过S2处理后的密网支架基底置于紫外下照射2h；
S4：将经过紫外照射后的密网支架基底浸没于浓度为3mg/mL抗坏血酸溶液中，于45℃下反应15min，取出，清洗并干燥，即得。
结果分析
以实施例1制得的抗凝血的密网支架为例，对采用本发明方案制得的密网支架的性能进行详细说明。
采用扫描电镜对实施例1制得的抗凝血的密网支架的微观结构进行观察，结果如图1所示。从扫描结果可以看出，未改性的镍钛密网支架表明光滑平整，仅存在微小的加工导致的划痕，而改性之后的支架表面呈致密均匀分布的纳米颗粒状，涂层的整体覆盖完整，说明了涂层成功的构建在了镍钛密网支架的表面。
采用FTIR对实施例制得的抗凝血小分子功能改性层的化学组分进行分析，结果如图2所示。从FTIR谱图的结果可知，改性后的支架表面出现了Ti-O键的特征峰，说明了二氧化钛基底层成功的构建在了密网支架表面，出现的C＝O、C-OH、C-H以及-OH特征峰充分表明了柠檬酸小分子成功的接枝到了二氧化钛的表面。
通过UP水滴对实施例制得的抗凝血小分子功能改性层的表面亲疏水性进行演示，结果如图3所示。通过照片可知，改性之前的镍钛金属片以及镍钛金属支架都是疏水，而改性之后的样品表面的水滴能够均匀铺展开，充分说明柠檬酸小分子涂层提高了支架的亲水性。
另外，考察实施例1制得的抗凝血的密网支架的体外血小板粘附和激活情况。
1.使用的血液为无名志愿者捐献。将新鲜血液与3.8％的柠檬酸钠按照10:1混合，在离心机上离心15min，转速为1500rpm，取上清液，即为富板浆(PRP)；
2.准备好的样品加入置于24孔板中，每个样品表面滴加100μL的富板浆，置于37℃恒温孵箱中孵化1h；
3.将孵化的样品取出用0.9％的氯化钠清洗三遍，并用2.5％的戊二醛固定4h。
本发明利用免疫荧光染色和扫面电镜对血小板的粘附形态和激活情况进行观察，免疫荧光染色步骤为：
(1)用0.9％的氯化钠清洗3次固定后的样品；
(2)在每个样品表面滴加70μL罗丹明(Rhodamine-phalloidin)溶液，避光条件放置15min；
(3)用0.9％的氯化钠清洗染色样品3次并吹干，在荧光显微镜下观察。
扫描观察血小板步骤：
(1)上述样品进行脱水处理：将样品依次放于50％，75％，90％和100％的酒精溶液中，每次15min；
(2)脱醇处理：将样品依次置于50％，75％，90％和100％的乙酸异戊酯溶液中，每次15min；
(3)临界点干燥，并进行喷金处理，通过扫描电镜进行观察。
本实施例制得的样品表面血小板粘附荧光染色如图4所示，可以看出，对照组不锈钢的表面粘附了大量的血小板，并且部分成激活状。在镍钛金属板和镍钛密网支架表面也粘附了不少的血小板，而改性后的样品表面几乎没有血小板的粘附，这充分说明了柠檬酸小分子层具有良好的抗凝效果。
本实施例制得的样品表面血小板粘附扫描电镜图如图5所示，镍钛金属支架表面粘附了大量的血小板，改性后的样品表面光滑平整，没有血小板的粘附，结果与荧光染色一致。上述结果表明柠檬酸小分子改性层具有优秀的抗凝能力。
考察实施例制得的抗凝血小分子功能改性层的体外溶血情况。
1.本实施例使用的血液为无名志愿者捐献。将新鲜血液与0.9％的氯化钠以4:5的比例混合；
2.将样品分别装于试管内，每组样品至少3个。另取阴性对照试管加入10mL的0.9％的氯化钠溶液，阳性对照试管加入10mL的蒸馏水。将所有样本置于37℃恒温孵箱中孵化30min；
3.孵化30min后每个试管加入200μL稀释的新鲜血液，于37℃恒温孵箱中继续孵化60min；
4.取出孵化后的血液置于EP管中并标注名称，于离心机中离心15min，转速3000rpm。离心后进行拍照观察溶血情况并取上清液在540nm的条件下测定吸光度，按照下式计算溶血率：
R＝(A-C1)/(C2-C1)×100％
其中R-溶血率(％)，A-样品吸光度，C1-阴性对照样吸光度，C2-阳性对照样吸光度。
溶血实验结果如图6所示，从照片可以看出，除了阳性对照之外，其余试样都是澄清的上清液，从溶血率的计算结果来看，所有试样的溶血率都在0.6％以下，但是改性之后的样品溶血率低于不锈钢和镍钛密网支架，在0.24％左右，表明改性后的材料血液相容性很好。
虽然结合实施例对本发明的具体实施方式进行了详细地描述，但不应理解为对本专利的保护范围的限定。在权利要求书所描述的范围内，本领域技术人员不经创造性劳动即可作出的各种修改和变形仍属本专利的保护范围。
Claims (10)
1.一种抗凝血的密网支架，其特征在于：包括密网支架基底以及键接于所述密网支架基底表面并具有抗凝血功能的有机小分子酸，所述有机小分子酸在密网支架基底表面形成厚度为5～500nm的涂层。
2.根据权利要求1所述的抗凝血的密网支架，其特征在于：所述密网支架基底为镍钛合金密网支架。
3.根据权利要求1所述的抗凝血的密网支架，其特征在于：所述有机小分子酸为柠檬酸、单宁酸、乙酰水杨酸或没食子酸。
4.如权利要求1～3任一项所述的抗凝血的密网支架的制备方法，其特征在于，包括以下步骤：
S1：对密网支架基底进行清洗并干燥；
S2：将经过S1处理后的密网支架基底浸没于温度为45～55℃的处理液中，并用氢氟酸将处理液的pH调节至3.5～4.0，浸泡0.5～4h后取出密网支架基底；所述处理液由氟钛酸铵溶液和硼酸溶液混合而成，其中氟钛酸铵的浓度为0.05～0.2M，硼酸的浓度为0.2～0.5M；
S3：将经过S2处理后的密网支架基底置于紫外下照射2～6h；
S4：将经过紫外照射后的密网支架基底浸没于浓度为1～3mg/mL有机小分子酸溶液中，于37～45℃下反应15～60min，取出，清洗并干燥，即得。
5.根据权利要求4所述的制备方法，其特征在于，S1中密网支架基底的清洗包括以下步骤：将密网支架基底置于去离子水中，超声清洗三次，每次5min；然后置于无水乙醇中，超声清洗三次，每次5min；再置于丙酮中，超声清洗三次，每次5min；最后真空干燥箱干燥。
6.根据权利要求4所述的制备方法，其特征在于：所述处理液的温度为50℃，pH为3.88，密网支架基底在处理液中的浸泡时间为2h。
7.根据权利要求4所述的制备方法，其特征在于：所述处理液中氟钛酸铵的浓度为0.1M，硼酸的浓度为0.3M。
8.根据权利要求4所述的制备方法，其特征在于：S3中紫外照射时间为4h。
9.根据权利要求4所述的制备方法，其特征在于：所述有机小分子酸为柠檬酸、单宁酸、乙酰水杨酸或没食子酸。
10.根据权利要求4所述的制备方法，其特征在于：S4中反应温度为40℃，反应时间为30min。
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