CN101626682B - 用于外科植入的生物组织 - Google Patents

用于外科植入的生物组织 Download PDF

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CN101626682B
CN101626682B CN200780039836.0A CN200780039836A CN101626682B CN 101626682 B CN101626682 B CN 101626682B CN 200780039836 A CN200780039836 A CN 200780039836A CN 101626682 B CN101626682 B CN 101626682B
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田斌
J·戴维森
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Abstract

公开了处理生物组织的方法，其能够干燥保存所述组织。在一个实施方式中，所述方法包括用包含多元醇和C1-C3醇的非水性处理溶液接触所述生物组织和从溶液处理过的生物组织去除部分处理溶液。也公开了应用上述方法准备的生物组织和用这种组织制备的假体装置。

Description

用于外科植入的生物组织
技术领域
本发明涉及用于外科植入的生物组织的准备或贮存。本发明也涉及生物组织和包含这种组织的生物假体装置(bioprosthetic devices)。
背景技术
通常用于可植入生物假体装置中或用于修复患者中的受损组织的动物组织一般使用化学试剂如甲醛和/或戊二醛贮存或用所述化学试剂预先处理而提供给医疗人员。这种化学处理有助于防止植入的生物假体装置被受者排斥，提供进行灭菌和有助于稳定组织中的蛋白质，从而使组织或包含这种组织的装置更抵抗机械疲劳和蛋白水解酶引起的降解。例如，甲醛和戊二醛通过与组织中的胶原和其它蛋白形成交联而稳定组织。另外，组织通常保存在包含戊二醛(和/或甲醛)的稀释水溶液中，以便将组织成分保持在水合态和保持无微生物环境。然而，包含这些试剂的组织必须在外科植入前进行充分洗涤。而且，在选择用于植入的生物组织时，一个重要的考虑事项是，医疗人员应当能够容易以尽可能接近即用的形式得到组织，即，在手术前进行最少的准备。这降低了误差机率，也减少了植入时间。
已经尝试开发预处理生物组织和包含生物组织的生物假体装置的方法，以便这种组织和装置可以基本不含甲醛或戊二醛而提供给医疗人员，并且以即用于植入或再水合的基本干燥形式提供生物组织。解决这一问题的一个尝试包括将生物组织浸没在C1-C3醇水溶液中，例如，乙醇水溶液，参见Duran等的美国专利号6,277,555和6,630,001。被溶液处理的组织然后被浸没在甘油水溶液或低分子量(＜1000 D)聚乙二醇中。此后，组织被简短浸没在肝素水溶液中、冷冻和冻干。
美国专利号4,300,243描述了用于外科植入的生物胶原组织的准备。该组织的准备包含使组织与新鲜量的水混溶性有机溶剂接触，所述溶剂选自甲醇、乙醇、丙醇、异丙醇、丙酮、甲基·乙基酮、和它们的混合物。然后，将过量的醇在0.1巴至1.0巴之间的压力下去除。然而，已知这种脱水过程显著减少组织的总尺寸，即，组织收缩。另外，经历这种脱水过程的生物组织不能成功地再水合并基本上回到其原始尺寸。结果是，包含经历这种脱水过程的组织成分的生物假体装置不是进行植入的良好候选物。参见，美国专利号6,534,004，第1栏，第53-65行。
美国专利号6,534,004描述提供包含生物组织的生物假体装置的方法，用于干燥贮存。该方法包括用具有多个羟基的有机化合物(例如，多元醇、聚乙二醇或水溶性碳水化合物)水溶液处理生物组织。更优选的处理溶液之一是甘油或甘油衍生物水溶液。生物组织被浸没在这种溶液中，移出并使其风干。然后对组织进行灭菌和包装贮存。
准备用于干燥贮存的组织的所有前述现有技术方法都需要使用水溶液。然而，水溶液的使用使得组织易于受微生物污染。因此，期望从方法中完全消除水，并且期望因而使干燥组织不易受微生物影响。本发明提供这种方法。其提供准备生物组织或包含这种组织的生物假体装置的新方法，适于灭菌干燥贮存，并且医疗人员可以以尽可能接近即用的形式得到。
发明内容
公开了处理生物组织的方法，其能够干燥贮存所述组织。在一个实施方式中，所述方法包括用包含多元醇和C1-C3醇的非水性处理溶液接触所述生物组织和从被溶液处理的生物组织去除部分处理溶液。多元醇浓度可以是按体积计40-95％并且C1-C3醇浓度可以是按体积计5-60％。组织和处理溶液间的接触时间可以变化，以便控制处理过程。例如，在一些实施方式中，用处理溶液接触生物组织的步骤进行30分钟以上的时间。
在一些实施方式中，多元醇是甘油并且C1-C3醇选自乙醇、正丙醇、2-丙醇或它们的混合物。
待处理生物组织可以是哺乳动物组织，例如，选自心包、主动脉根和主动脉瓣以及肺动脉根和肺动脉瓣、腱、韧带、皮肤、硬膜、腹膜、血管、胸膜、隔膜(diameter)、二尖瓣和三尖瓣的组织。
在一个实施方式中，处理方法进一步包括将即用于灭菌盛放的生物组织置于包装形式，其中所述包装形式是预先灭菌的，或在将组织置于该包装形式和密封该包装形式后进行灭菌。
本文也公开了包含用于外科植入到人中的灭菌溶液处理的生物组织的密封包装，其中所述溶液处理的生物组织通过上述处理方法的实施方式中的一种进行准备。进一步地，公开了用于外科植入到人中的灭菌溶液处理的生物组织，其中所述灭菌溶液处理的生物组织由上述处理方法的实施方式中的一种进行准备。也公开了心脏瓣膜假体，其包括可收缩的弹性瓣膜构件，其中所述瓣膜构件包含利用上述处理方法的实施方式中的一种进行溶液处理的生物组织，所述瓣膜构件安装于其上的弹性支架构件，所述支架构件具有内表面和外表面，以及连于瓣膜构件并位于瓣膜构件和支架构件之间的支撑物，其中支架构件形成连续表面并包括提供足够硬以防止翻转的结构的支柱件，并且支撑物从支架构件内表面延伸到支架构件外表面，并且其中支架构件具有足够的径向和纵向刚度，以经得住植入所需的径向力，抵抗主动脉反冲力，和为瓣膜假体提供长期支撑。在一个实施方式中，心脏瓣膜假体被提供为灭菌形式并且即用于外科植入。
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具体实施方式
本发明涉及处理哺乳动物生物组织进行准备或贮存的方法。理想地，所述方法提供生物组织或包含这种组织的生物假体装置，即用于外科植入或再水合，例如，用生理盐水再水合。本发明的一个实施方式利用多元醇和C1-C3醇的非水性组合。方法包括用按体积计为35％以上的多元醇和余量为C1-C3醇的处理溶液接触生物组织。处理溶液不含水，即，其为非水性的。对于贮存在灭菌贮存液例如固定液中的组织，生物组织与处理溶液的接触——例如通过将组织浸没在溶液中，其保持时间足以用处理溶液交换组织间隙容量中的大部分预贮存液，所述预贮存液可以是水性的。所述方法也包括从溶液处理过的组织去除部分处理溶液，以提供即用于灭菌盛放的生物组织。
如本文所用，″多元醇″是包含多个碳原子和两个或更多个羟基的有机分子，其中羟基连于碳原子。示范性多元醇包括甘油、乙二醇、聚乙二醇、丙二醇、丁二醇和甘油衍生物。所述方法中使用的更优选多元醇中的两种是甘油和丙二醇，和甘油或丙二醇的衍生物。当然，混合物形式的一种或更多种多元醇可以用在本方法中。因此，应理解，术语″多元醇″包含两种或更多种多元醇的混合物。例如，甘油和丙二醇的混合物可以用在本方法中。
本发明多种实施方式中使用的生物组织可以是哺乳动物组织，包括人组织，如得自人供体或尸体的组织。也可以使用牛、羊、马和猪组织。这些实施方式中使用的组织类型与用于普通外科手术中的组织相同，并且包含心包、主动脉根和主动脉瓣以及肺动脉根和肺动脉瓣、腱、韧带、皮肤、腹膜、筋膜、胸膜、二尖瓣和三尖瓣。
本领域普通技术人员会理解，根据本发明某些实施方式的处理方法可以用固定组织或天然组织实施。在某些情况下，用于本方法的生物组织在先前已通过用固定液处理组织进行固定。如本文所用，″固定″组织是其中其蛋白质与天然组织中的蛋白质相比具有降低的溶解度、抗原性和生物降解性质的组织。一般地，组织通过组织蛋白质的胺基团与醛交联而固定。组织固定方法可以包含化学和/或酶脱细胞步骤。漂洗固定组织，以大大降低组织中未反应固定剂的量。另一情况下，在本申请中公开的本发明处理之前，生物组织临时贮存在缓冲盐水溶液中。
生物组织与处理溶液接触，例如，通过将组织浸没在溶液中。在一些实施方式中，处理溶液的体积是与该溶液接触的生物组织的体积的至少2倍、至少50倍或至少100倍。如果生物组织要连于生物假体装置进行植入，则可以在其连接于生物假体装置之前或之后，用处理溶液处理该组织。例如，牛心包可以在形成于心瓣膜、血管移植物、支架遮盖物或心包片(patch)中之前或牛心包形成于心瓣膜、血管移植物、支架遮盖物或心包片中之后时间用处理溶液处理。
C1-C3醇选自甲醇、乙醇、异丙醇和正丙醇。乙醇是本方法中使用的优选醇。另外，应该理解，术语″C1-C3醇″包含两种或更多种C1-C3醇的混合物。例如，乙醇和异丙醇的混合物可以用在本方法中。
发明人已发现，多元醇相对于C1-C3醇的浓度影响贮存期间生物组织的尺寸稳定性。有关组织的尺寸稳定性的一个另外的因素是组织本身的类型。对于某些生物组织，处理溶液包括按体积计70％至95％的多元醇和按体积计5％至30％的C1-C3醇。例如，申请人已确定，对于牛心包瓣膜，处理溶液优选包含按体积计70％至95％的多元醇和按体积计5％至30％的C1-C3醇。
对于贮存在贮存液或固定剂中的组织，通过使生物组织与非水性处理溶液接触，水从组织的间隙容量去除，使得组织基本干燥。
本领域技术人员会容易知道，生物组织可以在一个或更多个独立的接触步骤中与处理溶液接触。替代在一个处理步骤中用处理溶液接触组织，接触步骤可以用相同或类似浓度的多元醇和C1-C3醇的处理溶液重复一次或更多次。例如，可以安排包含同一处理溶液的两次浸没浴。包含预贮存液的生物组织浸没在第一浴中第一时间段。然后从第一浴移去组织并浸没在第二浴中第二时间段。当然，普通技术人员可以确定，两个浴中的每一个中的多元醇和C1-C3醇的各自浓度是不同的，但在陈述并要求保护的量之内。
生物组织和处理溶液间的接触时间在一定程度上依赖于组织厚度和类型。如可以预料的，对于相对厚的组织，通常需要更多的接触时间。此外，如本领域普通技术人员公知，方法变量如搅动、温度、压力、流速等会影响需要的接触时间。
多元醇和C1-C3醇混合物(例如，75％-25％混合物)的制备可以在搅拌下进行数小时。用该溶液处理组织需要的时间取决于上述许多因素，并且可以进行数小时。在许多情况下，方法包含用处理溶液保持生物组织8小时或更长。在某些情况下，暴露时间可以超过12、16或24小时。
一旦生物组织充分暴露于处理溶液，将组织从溶液移去并在标准室温和湿度下暴露于环境气体或惰性环境例如氮气，以便没有不利地影响组织性质(一般地，在约15℃至约25℃的温度下和优选约50％以下的相对湿度下)。优选地，在清洁室或层流净化台中、在环境房间条件下进行干燥约1-4小时。
然后，将处理和干燥的组织或包含处理和干燥的组织的生物假体装置包装在基本不含液体的容器或包装中，用于随后的再水合或植入。″基本不含液体″的容器或包装指非流体环境，其中水或其它物质的存在被限制为环境气体中这种物质的大致含量(更精确地受到相对湿度的限定)。组织或包含该组织的生物假体装置被放在微生物抗性容器或包装中。一个优选方法是应用真空于包装以便最小化O2水平，并可以另外利用惰性气体如氮气回填。这种灭菌包装方法是本领域技术人员已知的。然后，包装的处理组织可以通过气体灭菌方法或通过暴露于离子辐射——包括γ和电子束方法——而灭菌。为了确保在灭菌后腔室(chamber)保持灭菌状态，包装件由不透微生物如细菌和真菌的材料制成。组织或包含该组织的生物假体装置放在室中和/或灭菌后，室被密封。
通过暴露于离子辐射或灭菌气体进行灭菌，特别是通过暴露于环氧乙烷灭菌，是本领域中的技术。在优选实施方式中，组织通过暴露组织于γ辐射而灭菌。通过暴露于环氧乙烷灭菌的常规方法的实例包括，在38℃的温度下，在8至10psig的室压下，暴露于10％环氧乙烷和90％氯氟烃(hydrochlorofluorocarbon)24小时或在54-57℃的温度下暴露130分钟。
得到的产品是基本灭菌和密封的以基本干燥形式存在的可植入组织或包含这种组织的生物假体装置。它特别非常适于外科植入人类患者中，用于治疗诸多疾病或病症。外科植入前，该生物组织或包含该组织的生物假体装置从包装中移去，并且任选地组织成分通过暴露于水溶液、优选灭菌水溶液被再水合。组织可以通过多次浸泡在灭菌溶液如生理盐水中而再水合。组织中的甘油可以由盐水容易地洗去。大多数甘油通过在盐水中洗涤组织约3-5分钟而洗脱出来。
所述处理生物组织和随后包装该组织或包含这种组织的装置的方法消除了由医疗人员在外科植入前对组织或生物假体装置灭菌的需要。所述方法在准备用于外科植入的组织和生物假体装置中提供较大的制造控制。换言之，准备用于植入的组织或假体装置的方法不需要在医院或手术室中进行。另外，医疗人员可能需要的唯一步骤是用生理盐水或其它适当的再水合剂再水合组织或生物假体装置上的组织。
所述方法提供生物组织或包含这种组织的生物假体装置，其具有基本即用于外科植入到患者中的尺寸稳定性。在干燥贮存约24小时和在生理盐水中再水合约5分钟后，根据本申请所述和所要求保护的方法处理的生物组织一般会恢复其原始水合尺寸的至少大约97％，更优选至少大约99％。因此，根据所述和所要求保护的方法准备的生物组织非常适用于可植入生物假体装置中，并且特别适用于植入患者心血管系统的接触血液的生物假体装置中。
本发明实施方式涉及生物假体装置，其包含本申请所述方法处理的心包组织心瓣膜。如所述，心包组织在连于装置之前或之后可以根据所述方法处理。心瓣膜具有在瓣膜接合处(固定器的一部分)在瓣膜边缘连在一起的多个瓣叶，它们一般轴向排列并均匀布置在瓣膜轴周围。瓣膜接合处各自布置在相邻的弯曲瓣膜尖之间，沿着瓣膜边缘。生物假体心瓣膜可以进一步包括固定器，其包括在轴周围排列以通常沿瓣膜尖端接触心瓣膜的多个尖端支撑物。连于固定器的心包瓣膜用本申请所述方法准备和处理。生物假体心瓣膜的更完整描述描述于美国专利号5,928,281中，其转让给Edwards Lifesciences(爱德华兹生命科学公司)，其全部内容并入本申请作为参考。
实施例1
在戊二醛存在下固定的牛心包被切割，以去除松弛组织和脂肪，并且被切片为具有表1和2所述厚度和直径的片(discs)。处理前测定每一片的尺寸，并且每一片被浸没在表1和2所述的各种处理溶液组合物中。处理溶液在使用溶液前约16小时制备。组织浸没在各种溶液组合物的每一个中，溶液：组织体积比是100∶1。组织保持与各种处理溶液接触约16小时，随后从溶液小心移去组织，使过量液体从组织排出并且风干组织。再次测量每一片的尺寸。处于干燥形式16小时后，每一片在常规盐水中再水合。再次测定再水合尺寸。表1
Figure G2007800398360D00081
表2
Figure G2007800398360D00082
处理和干燥的组织是柔韧的并且不会由于物理操作而破裂或破坏。其通过在环境温度下浸没在例如生理缓冲盐水中约5分钟而再水合。再水合后，组织在外观上与原始(未处理)固定组织不可区分。如表1和2的数据所示，对所评价的所有组分范围，组织保持尺寸稳定性。然而，对于更严格的尺寸考虑，例如生物假体心瓣膜，范围约50至95％甘油和余量为醇的处理溶液组成范围是优选的。从再水合后组织厚度的测定值得到类似结果。在此情况下，对于用甘油∶乙醇体积比为80∶20至50∶50的溶液处理的那些片而言，没有观察到组织厚度差异。虽然本发明的多种优选实施方式为示范目的已经公开，但本领域技术人员会理解，各种修改、添加和/或替换是可能的，这不背离本发明的范围和精神，如权利要求书中所公开的。

Claims (11)

1.处理牛心包组织的方法，所述方法包括：
使牛心包组织与包含甘油和C1-C3醇的非水性处理溶液接触，所述C1-C3醇选自乙醇、甲醇、正丙醇、2-丙醇或其混合物，其中所述甘油是按体积计70-95％；和
从溶液处理过的牛心包组织去除部分所述处理溶液，
其中所述方法用于非治疗目的。
2.权利要求1的方法，其中所述甘油是按体积计70-80％，并且所述C1-C3醇是按体积计20-30％。
3.权利要求2的方法，其中所述处理溶液包括按体积计75％的甘油和按体积计25％的C1-C3醇。
4.权利要求1的方法，其中使所述牛心包组织与所述处理溶液接触的步骤进行30分钟以上的时间。
5.权利要求1的方法，进一步包括将即用于灭菌盛放的所述牛心包组织置于包装形式，其中所述包装形式是预先灭菌的，或在将所述组织置于所述包装形式和密封所述包装形式后进行灭菌。
6.包含用于外科植入到人中的灭菌的溶液处理的牛心包组织的密封包装，所述溶液处理的牛心包组织通过包括下列步骤的方法进行准备：
使牛心包组织与包含甘油和C1-C3醇的非水性处理溶液接触，所述C1-C3醇选自乙醇、甲醇、正丙醇、2-丙醇或其混合物，其中所述甘油是按体积计70-95％；和
从溶液处理过的组织去除部分所述处理溶液，以提供溶液处理过的牛心包组织。
7.权利要求6的密封包装，其中所述处理溶液包括按体积计70％至80％的甘油和按体积计20％至30％的C1-C3醇。
8.用于外科植入到人中的灭菌的溶液处理的牛心包组织，所述灭菌的溶液处理的牛心包组织通过包括下列步骤的方法进行准备：
使牛心包组织与包含按体积计70％至90％的甘油和按体积计10％至30％的C1-C3醇的处理溶液接触，所述C1-C3醇选自乙醇、甲醇、正丙醇、2-丙醇或其混合物；和
从溶液处理过的组织去除部分所述处理溶液，以提供即用于灭菌盛放的牛心包组织。
9.权利要求8的牛心包组织，其中所述处理溶液包括按体积计70％至80％的甘油；和按体积计20％至30％的C1-C3醇。
10.心瓣膜假体，其包括：
可收缩的弹性瓣膜构件，其中所述瓣膜构件包括溶液处理的牛心包组织，所述溶液处理包括：
使所述牛心包组织与包含按体积计70％至90％的甘油和按体积计10％至30％的C1-C3醇的处理溶液接触，所述C1-C3醇选自乙醇、甲醇、正丙醇、2-丙醇或其混合物；和
从溶液处理过的组织去除部分所述处理溶液，以提供即用于灭菌盛放的牛心包组织；
弹性支架构件，其中安装所述瓣膜构件，所述支架构件具有内表面和外表面；和
支撑物，其连接于所述瓣膜构件并位于所述瓣膜构件和所述支架构件之间，其中所述支架构件形成连续表面并包括提供足够硬以防止翻转的结构的支柱件，并且所述支撑物从所述支架构件内表面延伸到所述支架构件外表面，并且其中所述支架构件具有足够的径向和纵向刚度，以经得住植入所需的径向力、抵抗主动脉反冲力和为所述瓣膜假体提供长期支撑。
11.权利要求10的心瓣膜假体，其以灭菌形式提供并且即用于外科植入。
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