Fabrication of pulsatile cardiac tissue grafts using a novel 3-dimensional cell sheet manipulation technique and temperature-responsive cell culture surfaces
Fabrication of pulsatile cardiac tissue grafts using a novel 3-dimensional cell sheet manipulation technique and temperature-responsive cell culture surfaces
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DOI：
10.1161/hh0302.105722
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发表时间：
2002-02-22
影响因子：
20.1
通讯作者：
Okano, T
中科院分区：
文献类型：
--
作者：
Shimizu, T;Yamato, M;Okano, T
修复受损心脏的细胞移植疗法的最新进展鼓励人们进一步尝试利用培养的心肌细胞对 3 维 (3-D) 心脏组织进行生物工程改造。目前，心脏组织工程正在利用传统技术来制造 3D 可生物降解支架作为临时细胞外基质。相比之下，现在描述了使用细胞片层 3 维分层新技术制造搏动心脏移植物的新方法。我们应用了接枝有温度响应性聚合物聚（N-异丙基丙烯酰胺）（PIPAAm）的新型细胞培养表面，只需降低温度，无需任何酶处理，汇合细胞即可从表面分离为细胞片。从 PIPAAm 移植表面分离的新生大鼠心肌细胞片被覆盖以构建心脏移植物。分层的细胞片开始同时脉冲，片之间通过 connexin43 建立形态通讯。当 4 个片材分层时，宏观观察到工程结构会自发脉冲。在体内，将分层心肌细胞片移植到裸鼠的皮下组织中。移植后三周，检测来自移植物的表面电图并肉眼观察自发搏动。组织学研究显示心脏组织的特征结构和收缩组织内的多个新生血管形成。与移植到 8 周龄大鼠体内的构建体相比，移植到 3 周龄大鼠体内的构建体表现出更多的心肌细胞肥大和更少的结缔组织。搏动性心脏移植物的长期存活期已被证实长达 12 周。这些结果表明，通过分层心肌细胞片在体外和体内实现了电通信脉动 3-D 心脏结构。基于该技术的心脏组织工程可能对心脏模型制造和心血管组织修复有用。本文全文可在 http://www.circresaha.org 上获取。
Recent progress in cell transplantation therapy to repair impaired hearts has encouraged further attempts to bioengineer 3-dimensional (3-D) heart tissue from cultured cardiomyocytes. Cardiac tissue engineering is currently pursued utilizing conventional technology to fabricate 3-D biodegradable scaffolds as a temporary extracellular matrix. By contrast, new methods are now described to fabricate pulsatile cardiac grafts using new technology that layers cell sheets 3-dimensionally. We apply novel cell culture surfaces grafted with temperature-responsive polymer, poly(N-isopropylacrylamide) (PIPAAm), from which confluent cells detach as a cell sheet simply by reducing temperature without any enzymatic treatments. Neonatal rat cardiomyocyte sheets detached from PIPAAm-grafted surfaces were overlaid to construct cardiac grafts. Layered cell sheets began to pulse simultaneously and morphological communication via connexin43 was established between the sheets. When 4 sheets were layered, engineered constructs were macroscopically observed to pulse spontaneously. In vivo, layered cardiomyocyte sheets were transplanted into subcutaneous tissues of nude rats. Three weeks after transplantation, surface electrograms originating from transplanted grafts were detected and spontaneous beating was macroscopically observed. Histological studies showed characteristic structures of heart tissue and multiple neovascularization within contractile tissues. Constructs transplanted into 3-week-old rats exhibited more cardiomyocyte hypertrophy and less connective tissue than those placed into 8-week-old rats. Long-term survival of pulsatile cardiac grafts was confirmed up to 12 weeks. These results demonstrate that electrically communicative pulsatile 3-D cardiac constructs were achieved both in vitro and in vivo by layering cardiomyocyte sheets. Cardiac tissue engineering based on this technology may prove useful for heart model fabrication and cardiovascular tissue repair. The full text of this article is available at http://www.circresaha.org.