Molyneaux Kathleen, Wnek Maria D, Craig Sonya E L, Vincent Jason, Rucker Imani, Wnek Gary E, Brady-Kalnay Susann M
Department of Molecular Biology & Microbiology, Case Western Reserve University, Cleveland, Ohio, USA.
Department of Macromolecular Science and Engineering, Case Western Reserve University, Cleveland, Ohio, USA.
J Biomed Mater Res B Appl Biomater. 2021 Nov;109(11):1744-1753. doi: 10.1002/jbm.b.34832. Epub 2021 Apr 13.
We employed aqueous solutions of highly-hydrolyzed (>99+%) poly(vinyl alcohol), PVA, to coat plastic dishes as a method to efficiently induce three-dimensional (3D) culturing of cells. The coatings were prepared by simple evaporation of 3 wt/vol% solutions of PVA in water and require no additional processing steps after air drying under sterile conditions. The coating allows spheroids to form in solution. Spheroid formation is usually preferable to two-dimensional (2D) culturing as it creates a more realistic ex vivo model of some human tissues and tumors. Using PVA-coated cell culture plates, we demonstrated that we can grow reproducibly sized spheroids using several human glioma cell lines, including LN229, U87 MG, and Gli36, and the embryonic kidney cell line, 293T. Spheroids formed on PVA-coated plates grow as well as on other commercially-available, low-attachment plates, and have excellent optical imaging properties. As spheroids, LN229 cells express markers of cancer stem cells. Finally, we confirmed that spheroids generated on PVA-coated plates are sensitive to molecular perturbations, as increased expression of the cell adhesion molecule PTPμ significantly increased the size of spheroids. The PVA hydrogel layer is an effective tool for creating a more realistic ex vivo culture system than traditional 2D culture and can be used to generate cell spheroids for potential application in drug screening and personalized medicine for diseases such as cancer.
我们使用了高度水解（>99%）的聚乙烯醇（PVA）水溶液来包被塑料培养皿，以此作为一种有效诱导细胞三维（3D）培养的方法。通过简单蒸发3 wt/vol%的PVA水溶液来制备包被层，在无菌条件下空气干燥后无需额外的加工步骤。这种包被层能使球体在溶液中形成。与二维（2D）培养相比，球体形成通常更可取，因为它能创建更接近某些人体组织和肿瘤的体外模型。使用PVA包被的细胞培养板，我们证明了可以使用几种人类胶质瘤细胞系，包括LN229、U87 MG和Gli36，以及胚胎肾细胞系293T，可重复性地培养出大小一致的球体。在PVA包被板上形成的球体生长情况与其他市售的低附着板上的球体一样好，并且具有出色的光学成像特性。作为球体，LN229细胞表达癌症干细胞标志物。最后，我们证实了在PVA包被板上生成的球体对分子扰动敏感，因为细胞粘附分子PTPμ的表达增加显著增大了球体的尺寸。PVA水凝胶层是一种有效的工具，可用于创建比传统二维培养更接近真实情况的体外培养系统，并且可用于生成细胞球体，有望应用于药物筛选和癌症等疾病的个性化医疗。
