Biomomentum多轴机械测试仪Mach-1应用：立体光刻3D生物打印方法制作人角膜基质等效物。

生物医学工程学年鉴，生物材料-工程细胞行为，已出版：2020年6月5日3D生物打印技术是用于角膜基质组织再生的一种有前途的方法。在这项研究中，与角膜基质细胞混合的甲基丙烯酸明胶（GelMA）被用作生物墨水。基于可见光的立体光刻（SLA）3D生物打印方法用于打印人类角膜基质的解剖学相似的圆顶形结构。测试了两种不同浓度的GelMA大分子单体（7.5和12.5％）的角膜基质生物印记。由于高聚物浓度高，12.5％的GelMA比7.5％的GelMA坚硬，这使其更易于处理。就生物印刷支架的水含量和透光率而言，我们观察到具有12.5％GelMA浓度的支架更接近于天然角膜基质组织。回到顶端|提供反馈信息的人随后，细胞增殖，研究了在生物打印支架中封装的人角膜基质细胞的基因和蛋白质表达。在第1天和第7天，观察到在12.5％GelMA支架中的细胞相容性分别为81.86和156.11％，显着高于7.5％GelMA支架中的细胞相容性。7天后在12.5％的GelMA样品中观察到角膜基质细胞伸长，表明细胞附着，生长和在支架内的整合。与在塑料组织培养板上培养的细胞相比，在12.5％GelMA支架中培养的细胞的I型胶原蛋白，卢米肯和硫酸角质素的基因表达随时间增加。28天后，还使用免疫组织化学法观察了I型胶原蛋白和lumican的表达。使用垂直轴的微机械测试仪（测定交联时间MACH-c，Biomomentum，加拿大）。

Abstract

3D bioprinting technology is a promising approach for corneal stromal tissue regeneration. In this study, gelatin methacrylate (GelMA) mixed with corneal stromal cells was used as a bioink. The visible light-based stereolithography (SLA) 3D bioprinting method was utilized to print the anatomically similar dome-shaped structure of the human corneal stroma. Two different concentrations of GelMA macromer (7.5 and 12.5%) were tested for corneal stroma bioprinting. Due to high macromer concentrations, 12.5% GelMA was stiffer than 7.5% GelMA, which made it easier to handle. In terms of water content and optical transmittance of the bioprinted scaffolds, we observed that scaffold with 12.5% GelMA concentration was closer to the native corneal stroma tissue. Subsequently, cell proliferation, gene and protein expression of human corneal stromal cells encapsulated in the bioprinted scaffolds were investigated. Cytocompatibility in 12.5% GelMA scaffolds was observed to be 81.86 and 156.11% at day 1 and 7, respectively, which were significantly higher than those in 7.5% GelMA scaffolds. Elongated corneal stromal cells were observed in 12.5% GelMA samples after 7 days, indicating the cell attachment, growth, and integration within the scaffold. The gene expression of collagen type I, lumican and keratan sulfate increased over time for the cells cultured in 12.5% GelMA scaffolds as compared to those cultured on the plastic tissue culture plate. The expression of collagen type I and lumican were also visualized using immunohistochemistry after 28 days. These findings imply that the SLA 3D bioprinting method with GelMA hydrogel bioinks is a promising approach for corneal stroma tissue biofabrication.

The crosslinking time was determined using a vertical axis micromechanical tester (Mach-1 v500c, Biomomentum Inc., Montreal, QC, Canada) by recording in situ gelation kinetics.