Division of Craniofacial Function Engineering, Tohoku University, Graduate School of Dentistry, 4-1 Seiryo-machi, Aoba-ku, Sendai 980-8575, Japan.
Biomaterials. 2012 Nov;33(33):8430-41. doi: 10.1016/j.biomaterials.2012.08.040. Epub 2012 Aug 30.
Since oxygen is one of the critical limiting factors for maintaining cell viability and function, a great deal of effort is being focused on improving the oxygen supply to three-dimensional (3D) cellular constructs. Here, we report a technique to construct spheroids utilizing 3D culture chips with a rapid and simple method for the replication of the surface structures of a polydimethylsiloxane (PDMS) mold. The resultant spheroid culture chip made it possible to rapidly yield high numbers of the spheroids at a time as well as to obtain uniform spheroids with a narrow size distribution and to collect the spheroids easily and noninvasively. The most important feature of this spheroid culture chip is that it enables direct oxygen supply to the cells because the chip is made with only gas-permeable PDMS. When human hepatoma HepG2 cells were grown on the oxygen-permeable chips as a model for liver cells, the cellular growth was remarkably enhanced, and the anaerobic glycolysis was significantly reduced compared to the non-oxygen-permeable chips. Furthermore, the oxygen-permeable chip improved the albumin secretion rates compared to the conventional spheroid culture system after 10 days. Histochemical and immunohistochemical analyses demonstrated that the oxygen-permeable chip dramatically prevented hypoxia in the core of the spheroids and subsequent central necrosis. Surprisingly, the diameters of approximately 400 and 600 μm were estimated to be the threshold of the hypoxic and survival size, respectively, for the HepG2 spheroids in the oxygenated chip. These results indicate that this chip is useful for engineering 3D cellular constructs with high viability and functionality for tissue engineering.
由于氧气是维持细胞活力和功能的关键限制因素之一，因此人们正在努力提高向三维（3D）细胞构建体供氧的能力。在这里，我们报告了一种利用 3D 培养芯片构建球体的技术，该技术采用快速简单的方法复制聚二甲基硅氧烷（PDMS）模具的表面结构。由此产生的球体培养芯片可一次快速产生大量球体，并获得具有较窄尺寸分布且易于非侵入性收集的均匀球体。这种球体培养芯片的最重要特征是它能够为细胞提供直接的氧气供应，因为该芯片仅由透气的 PDMS 制成。当用人肝癌 HepG2 细胞作为肝细胞模型在透气芯片上生长时，与非透气芯片相比，细胞生长显著增强，无氧糖酵解明显减少。此外，与传统的球体培养系统相比，透气芯片在 10 天后提高了白蛋白分泌率。组织化学和免疫组织化学分析表明，透气芯片可显著防止球体核心缺氧和随后的中心坏死。令人惊讶的是，在含氧芯片中，直径约为 400 和 600μm 的球体被估计分别为 HepG2 球体缺氧和存活的临界尺寸。这些结果表明，该芯片可用于构建具有高活力和功能的用于组织工程的 3D 细胞构建体。
