Automation of Pressure Control Improves Whole Porcine Heart Decellularization
Automation of Pressure Control Improves Whole Porcine Heart Decellularization
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DOI：
10.1089/ten.tec.2014.0709
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发表时间：
2015-11-01
影响因子：
3
通讯作者：
Cook, Alonzo D.
中科院分区：
文献类型：
--
作者：
Momtahan, Nima;Poornejad, Nafiseh;Cook, Alonzo D.
全心脏去细胞化结合患者特异性细胞可能被证明是一种非常有价值的方法来设计新的心脏。温和的去污剂通常用于去细胞化过程中，但已知会使关键蛋白质和生长因子变性和溶解，因此在去细胞化过程中可能对细胞外基质（ECM）具有破坏性。在本研究中，猪心脏的脱细胞化在24小时内完成，仅用十二烷基硫酸钠暴露6小时，DNA去除率为98%。在去细胞化过程中自动控制压力减少了去污剂暴露时间，同时仍然完全去除免疫原性细胞碎片。当将天然组织样品与经处理的ECM进行比较时，巨噬细胞的刺激大大降低。通过DNA含量分析确认细胞完全去除。证明了一般胶原蛋白和弹性蛋白的保留。糖胺聚糖和胶原定量均显示脱细胞后含量无显著差异。脱细胞后ECM的压缩弹性模量在低应变下低于天然ECM，但在高应变下没有显著差异。天然和脱细胞心脏的血管的聚氨酯铸件表明，脱细胞后微血管网络得到保留。还开发了使用牛血的静态血液血栓形成测定。最后，通过将心脏成纤维细胞和内皮细胞重新接种在心肌和内皮细胞样品上来证明ECM样品的再细胞化潜力。
Whole heart decellularization combined with patient-specific cells may prove to be an extremely valuable approach to engineer new hearts. Mild detergents are commonly used in the decellularization process, but are known to denature and solubilize key proteins and growth factors and can therefore be destructive to the extracellular matrix (ECM) during the decellularization process. In this study, the decellularization of porcine hearts was accomplished in 24h with only 6h of sodium dodecyl sulfate exposure and 98% DNA removal. Automatically controlling the pressure during decellularization reduced the detergent exposure time while still completely removing immunogenic cell debris. Stimulation of macrophages was greatly reduced when comparing native tissue samples to the processed ECM. Complete cell removal was confirmed by analysis of DNA content. General collagen and elastin preservation was demonstrated. Glycosaminoglycans and collagen quantification both showed no significant differences in content after decellularization. The compression elastic modulus of the ECM after decellularization was lower than native at low strains, but there was no significant difference at high strains. Polyurethane casts of the vasculature of native and decellularized hearts demonstrated that the microvasculature network was preserved after decellularization. A static blood thrombosis assay using bovine blood was also developed. Finally, the recellularization potential of the ECM samples was demonstrated by reseeding cardiac fibroblasts and endothelial cells on the myocardium and endocardium samples.