1 Division of Pediatric Surgery, Department of Surgery, Massachusetts General Hospital , Boston, Massachusetts.
2 Department of Chemical Engineering, Northeastern University , Boston, Massachusetts.
Tissue Eng Part C Methods. 2017 Sep;23(9):565-573. doi: 10.1089/ten.TEC.2017.0269.
The native extracellular matrix (ECM) serves as a unique platform for tissue engineering because it provides an organ-specific scaffold in terms of both matrix composition and tissue architecture. However, efficacious cell-seeding techniques for recellularizing the ECM constructs with appropriate cell types to restore biological function remain under development. In this study, the impact of spraying as a seeding technique for repopulation of decellularized small intestine was investigated. In a series of experiments, CaCo-2 cells were first used to investigate the effect of spray device type and pressure on cell viability and to optimize parameters for seeding intestinal epithelial cells. High cell viability and a homogeneous cell distribution were obtained when cell suspensions were sprayed through an airbrush at low pressure. Next, the effect of seeding method and spray pressure on the size and dispersal of intestinal organoids, a more complex and clinically relevant intestinal stem cell population, was evaluated. The feasibility of seeding intestinal epithelial cells onto decellularized scaffolds was next studied using sprayed CaCo-2 cells, which survived the spray-seeding process and formed a monolayer on the scaffold. Finally, airbrush seeding was used to spray intestinal organoids onto the scaffolds, with cell survival and tissue architecture evaluated after 1 week of culture. Organoids seeded through pipetting onto the decellularized scaffold survived, but demonstrated aggregation, with cells organized around multiple small lumens. In contrast, organoids airbrush spray seeded at 0.35 bar onto the decellularized scaffold not only engrafted but also demonstrated formation of an epithelial monolayer that resembled the absorptive surface found on intestinal villi. The results suggest that seeding cells through airbrush spraying holds great potential for use in tissue engineering, especially for large-scale tubular organ recellularization.
天然细胞外基质 (ECM) 是组织工程的独特平台，因为它在基质组成和组织架构方面提供了具有器官特异性的支架。然而，用适当的细胞类型有效接种 ECM 构建体以恢复生物功能的细胞接种技术仍在开发中。在这项研究中，研究了喷雾作为一种再细胞化脱细胞小肠的接种技术的影响。在一系列实验中，首先使用 CaCo-2 细胞来研究喷雾装置类型和压力对细胞活力的影响，并优化肠上皮细胞接种的参数。当通过低压空气喷枪喷射细胞悬浮液时，获得了高细胞活力和均匀的细胞分布。接下来，评估了接种方法和喷雾压力对肠类器官（更复杂和更具临床相关性的肠干细胞群体）的大小和分散的影响。接下来，使用喷涂的 CaCo-2 细胞研究了将肠上皮细胞接种到脱细胞支架上的可行性，喷涂 CaCo-2 细胞在喷雾接种过程中存活并在支架上形成单层。最后，使用空气喷枪将肠类器官喷射到支架上，并在培养 1 周后评估细胞存活和组织架构。通过吸管接种到脱细胞支架上的类器官存活下来，但表现出聚集，细胞围绕多个小腔组织。相比之下，以 0.35 巴的压力通过空气喷枪将类器官喷射到脱细胞支架上不仅可以植入，而且还表现出形成类似于肠绒毛上吸收表面的上皮单层。结果表明，通过空气喷枪喷射细胞具有在组织工程中应用的巨大潜力，特别是用于大规模管状器官的再细胞化。
