3B's Research Group-Biomaterials, Biodegradables and Biomimetics, University of Minho, Headquarters of the European Institute of Excellence on Tissue Engineering and Regenerative Medicine, AvePark, 4806-909 Taipas, Guimarães, Portugal.
Integr Biol (Camb). 2012 Mar;4(3):318-27. doi: 10.1039/c2ib00170e. Epub 2012 Feb 2.
We report on the development of a new array-based screening flat platform with the potential to be used as a high-throughput device based on biomimetic polymeric substrates for combinatorial cell/3D biomaterials screening assays in the context of tissue engineering. Polystyrene was used to produce superhydrophobic surfaces based on the so-called lotus effect. Arrays of hydrophilic regions could be patterned in such surfaces using UV/ozone radiation, generating devices onto which combinatorial hydrogel spots were deposited. The biological performance of encapsulated cells in hydrogels could be tested in an in vitro 3D environment assuming that each site was isolated from the others due to the high contrast of wettability between the patterned spots and the superhydrophobic surroundings. Three different polymers-chitosan, collagen and hyaluronic acid-were combined with alginate in different proportions in order to obtain combinatorial binary alginate-based polymeric arrays. The effect of the addition of gelatin to the binary structures was also tested. The gels were chemically analyzed by FTIR microscopic mapping. Cell culture results varied according to the hydrogel composition and encapsulated cell types (L929 fibroblast cells and MC3T3-E1 pre-osteoblast cells). Cell viability and number could be assessed by conventional methods, such as MTS reduction test and dsDNA quantification. Non-destructive image analysis was performed using cytoskeleton and nuclei staining agents and the results were consistent with the ones obtained by conventional sample-destructive techniques. Briefly, L929 cells showed higher number and viability for higher alginate-content and collagen-containing hydrogels, while MC3T3-E1 showed higher cell viability and cell number in lower alginate-content and chitosan containing hydrogels. The addition of gelatin did not influence significantly cell metabolic activity or cell number in any of the encapsulated cell types.
我们报告了一种新的基于阵列的筛选平面平台的开发，该平台具有成为高通量设备的潜力，基于仿生聚合物基底，用于组织工程中细胞/3D 生物材料组合筛选分析。聚苯乙烯被用于生产基于所谓的“荷叶效应”的超疏水表面。亲水区阵列可以使用 UV/臭氧辐射在这些表面上进行图案化，生成可以沉积组合水凝胶点的器件。在体外 3D 环境中，可以测试水凝胶中包封细胞的生物学性能，假设由于图案化斑点和超疏水环境之间的润湿性高度对比，每个位点都与其他位点隔离。三种不同的聚合物-壳聚糖、胶原蛋白和透明质酸-与海藻酸钠以不同的比例组合，以获得组合的二元海藻酸钠基聚合物阵列。还测试了向二元结构中添加明胶的效果。通过 FTIR 微观映射对凝胶进行化学分析。细胞培养结果根据水凝胶组成和包封细胞类型（L929 成纤维细胞和 MC3T3-E1 前成骨细胞）而有所不同。细胞活力和数量可以通过常规方法评估，如 MTS 还原试验和 dsDNA 定量。使用细胞骨架和核染色剂进行非破坏性图像分析，结果与传统的破坏性样本技术获得的结果一致。简而言之，L929 细胞在含有更高海藻酸钠和胶原蛋白的水凝胶中表现出更高的细胞数量和活力，而 MC3T3-E1 细胞在含有更低海藻酸钠和壳聚糖的水凝胶中表现出更高的细胞活力和数量。明胶的添加并没有显著影响任何包封细胞类型的细胞代谢活性或细胞数量。
