Vacuum-assisted decellularization: an accelerated protocol to generate tissue-engineered human tracheal scaffolds.
Vacuum-assisted decellularization: an accelerated protocol to generate tissue-engineered human tracheal scaffolds.
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DOI：
10.1016/j.biomaterials.2017.02.001
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发表时间：
2017-04
期刊：
影响因子：
14
通讯作者：
Janes SM
中科院分区：
文献类型：
--
作者：
Butler CR;Hynds RE;Crowley C;Gowers KH;Partington L;Hamilton NJ;Carvalho C;Platé M;Samuel ER;Burns AJ;Urbani L;Birchall MA;Lowdell MW;De Coppi P;Janes SM
不适合常规内镜或开放手术的大气管病变患者可能需要气管置换术，但目前对如何实现这一目标尚无共识。脱细胞或合成气管支架的组织工程为气道重建提供了新的途径。脱细胞人供体气管支架已应用于同情使用的临床病例，但自然来源的细胞外基质（ECM）支架需要漫长的准备时间。在这里，我们比较了临床应用的洗涤剂-酶法（DEM）和加速真空辅助脱细胞（VAD）方案。我们用这些技术检测了人供体气管脱细胞后的组织学外观、DNA含量和细胞外基质组成。此外，我们进行了扫描电子显微镜（SEM）和生物力学测试来分析脱细胞性能。为了评估用VAD制备的支架的生物相容性，我们在体外用原代人气道上皮细胞播种支架，并在鸡体内进行绒毛尿囊膜（CAM）和皮下植入试验。DEM和VAD两种方案均产生了脱细胞良好的气管支架，没有不良的机械效应，并且支架保留了体外和体内细胞整合的能力。我们得出的结论是，与DEM相比，使用VAD生产支架所需的时间大幅减少（大约9天vs. 3-8周），但并不影响生产的人体气管支架的质量。这些发现可能会为临床脱细胞技术提供信息，因为VAD可以加速支架的生产并降低相关成本。
Patients with large tracheal lesions unsuitable for conventional endoscopic or open operations may require a tracheal replacement but there is no present consensus of how this may be achieved. Tissue engineering using decellularized or synthetic tracheal scaffolds offers a new avenue for airway reconstruction. Decellularized human donor tracheal scaffolds have been applied in compassionate-use clinical cases but naturally derived extracellular matrix (ECM) scaffolds demand lengthy preparation times. Here, we compare a clinically applied detergent-enzymatic method (DEM) with an accelerated vacuum-assisted decellularization (VAD) protocol. We examined the histological appearance, DNA content and extracellular matrix composition of human donor tracheae decellularized using these techniques. Further, we performed scanning electron microscopy (SEM) and biomechanical testing to analyze decellularization performance. To assess the biocompatibility of scaffolds generated using VAD, we seeded scaffolds with primary human airway epithelial cells in vitro and performed in vivo chick chorioallantoic membrane (CAM) and subcutaneous implantation assays. Both DEM and VAD protocols produced well-decellularized tracheal scaffolds with no adverse mechanical effects and scaffolds retained the capacity for in vitro and in vivo cellular integration. We conclude that the substantial reduction in time required to produce scaffolds using VAD compared to DEM (approximately 9 days vs. 3–8 weeks) does not compromise the quality of human tracheal scaffold generated. These findings might inform clinical decellularization techniques as VAD offers accelerated scaffold production and reduces the associated costs.