Acellular Cardiac Extracellular Matrix as a Scaffold for Tissue Engineering: In Vitro Cell Support, Remodeling, and Biocompatibility
Acellular Cardiac Extracellular Matrix as a Scaffold for Tissue Engineering: In Vitro Cell Support, Remodeling, and Biocompatibility
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DOI：
10.1089/ten.tec.2009.0111
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发表时间：
2010-08-01
影响因子：
3
通讯作者：
Machluf, Marcelle
中科院分区：
文献类型：
--
作者：
Eitan, Yuval;Sarig, Udi;Machluf, Marcelle
我们开发了一种有效的脱细胞过程，用于从天然心脏组织中分离细胞外基质（ECM）。分离的ECM在弹性、强度和耐久性方面表现出理想的机械性能，这是用于心脏组织修复的支架所需的性能。本研究进一步探讨了这种支架在心脏组织工程中与种子细胞相互作用和生物相容性方面的潜在用途。我们使用了通常研究的成纤维细胞、心肌细胞和间充质干细胞，这些细胞被分离并接种到支架上。通过3，3 ′-双十八烷基氧碳菁高氯酸盐染色跟踪细胞密度和分布，并通过AlamarBlue（R）测定和荧光素-二乙酸盐/碘化丙啶染色评估它们的增殖和存活力。成纤维细胞接种的支架缩小到1-2 mm（3）球状体，其糖胺聚糖显著增加了23%。ECM重塑相关的胶原蛋白I和III，基质金属蛋白酶2，和1型金属蛋白酶的组织抑制剂的mRNA的表达进行了定量的实时聚合酶链反应，并发现显着升高成纤维细胞接种支架，与对照细胞板上。如机械测试所示，与无细胞支架相比，成纤维细胞接种的支架失去了一些灵活性，但获得了强度。接种心肌细胞的支架在接种后几天开始一致搏动，并且心肌细胞表达典型的功能性心脏标记物，如α-辅肌动蛋白、肌钙蛋白I和连接蛋白43。免疫荧光染色和扫描电镜观察显示细胞排列呈长条状。间充质干细胞接种的支架在培养物中保持活力超过24天。这些发现进一步加强了脱细胞心脏ECM作为心脏再生生物材料的潜在用途。
We have developed an efficient decellularization process for the isolation of extracellular matrix (ECM) from native cardiac tissue. The isolated ECM exhibited desirable mechanical properties in terms of elasticity, strength and durability-properties required from scaffolds used for cardiac tissue repair. This study further investigates the potential use of this scaffold for cardiac tissue engineering in terms of interactions with seeded cells and biocompatibility. We used the commonly studied fibroblasts, cardiomyocytes, and mesenchymal stem cells, which were isolated and seeded onto the scaffold. Cell density and distribution were followed by 3,3'-dioctadecyloxacarbocyanine perchlorate staining, and their proliferation and viability were assessed by AlamarBlue(R) assay and fluorecein-diacetate/propidium iodide staining. Fibroblast-seeded scaffolds shrank to 1-2 mm(3) spheroids, and their glycosaminoglycans significantly increased by 23%. The expression of ECM remodeling-related mRNAs of collagens I and III, matrix metalloproteinase 2, and type 1 tissue inhibitor of metalloproteinases was quantified by real-time polymerase chain reaction, and was found significantly elevated in fibroblast-seeded scaffold, compared with the control cells on plates. Fibroblast-seeded scaffolds lost some flexibility, yet gained strength compared with the acellular scaffolds, as shown by mechanical testing. Scaffold seeded with cardiomyocyte began to beat in concert few days after seeding, and the myocytes expressed typical functional cardiac markers such as a-actinin, troponin I, and connexin43. The cells revealed aligned elongated morphology, as presented by immunofluorescent staining and scanning electron microscopy. Mesenchymal stem cell-seeded scaffolds maintained viability over 24 days in culture. These findings further strengthen the potential use of acellular cardiac ECM as a biomaterial for heart regeneration.