Department of Mechanical and Nuclear Engineering, College of Engineering, Virginia Commonwealth University, Richmond, Virginia, United States of America.
Department of Biomedical Engineering, College of Engineering, Virginia Commonwealth University, Richmond, Virginia, United States of America.
PLoS One. 2022 Jun 16;17(6):e0269499. doi: 10.1371/journal.pone.0269499. eCollection 2022.
Tissue engineering commonly entails combining autologous cell sources with biocompatible scaffolds for the replacement of damaged tissues in the body. Scaffolds provide functional support while also providing an ideal environment for the growth of new tissues until host integration is complete. To expedite tissue development, cells need to be distributed evenly within the scaffold. For scaffolds with a small diameter tubular geometry, like those used for vascular tissue engineering, seeding cells evenly along the luminal surface can be especially challenging. Perfusion-based cell seeding methods have been shown to promote increased uniformity in initial cell distribution onto porous scaffolds for a variety of tissue engineering applications. We investigate the seeding efficiency of a custom-designed perfusion-based seed-and-culture bioreactor through comparisons to a static injection counterpart method and a more traditional drip seeding method. Murine vascular smooth muscle cells were seeded onto porous tubular electrospun polycaprolactone scaffolds, 2 mm in diameter and 30 mm in length, using the three methods, and allowed to rest for 24 hours. Once harvested, scaffolds were evaluated longitudinally and circumferentially to assess the presence of viable cells using alamarBlue and live/dead cell assays and their distribution with immunohistochemistry and scanning electron microscopy. On average, bioreactor-mediated perfusion seeding achieved 35% more luminal surface coverage when compared to static methods. Viability assessment demonstrated that the total number of viable cells achieved across methods was comparable with slight advantage to the bioreactor-mediated perfusion-seeding method. The method described is a simple, low-cost method to consistently obtain even distribution of seeded cells onto the luminal surfaces of small diameter tubular scaffolds.
组织工程通常需要将自体细胞源与生物相容性支架结合使用，以替代体内受损组织。支架提供功能支持，同时为新组织的生长提供理想的环境，直到宿主完全整合。为了加速组织发育，需要将细胞均匀分布在支架内。对于直径较小的管状支架，如用于血管组织工程的支架，均匀地在腔表面播种细胞可能特别具有挑战性。基于灌注的细胞播种方法已被证明可促进各种组织工程应用中多孔支架初始细胞分布的均匀性增加。我们通过与静态注射对照方法和更传统的滴注播种方法进行比较，研究了定制的基于灌注的播种和培养生物反应器的播种效率。使用三种方法将小鼠血管平滑肌细胞播种到直径为 2 毫米、长度为 30 毫米的多孔管状静电纺聚己内酯支架上，并允许其静置 24 小时。收获后，从纵向和周向评估支架，使用 alamarBlue 和活/死细胞检测评估存在有活力的细胞，并通过免疫组织化学和扫描电子显微镜评估其分布。平均而言，与静态方法相比，生物反应器介导的灌注播种可实现 35%的更大腔面覆盖率。活力评估表明，跨方法获得的总活细胞数相当，生物反应器介导的灌注播种方法略有优势。所描述的方法是一种简单、低成本的方法，可始终如一地将播种的细胞均匀分布在小直径管状支架的腔表面上。
