Dvir Tal, Benishti Nessi, Shachar Michal, Cohen Smadar
Department of Biotechnology Engineering, Ben-Gurion University of Negev, Beer Sheva, Israel.
Tissue Eng. 2006 Oct;12(10):2843-52. doi: 10.1089/ten.2006.12.2843.
We developed a novel perfusion bioreactor that is capable of cultivating multiple 3-dimensional (3D) cellular constructs in one flow chamber with a total cross-section area of 20 cm(2). Two unique features integrated into the bioreactor provided a homogenous fluid flow along the bioreactor cross-section and maximal exposure of the cellular constructs to the perfusing medium. Mathematical modeling of the fluid flow regime in the perfusion bioreactor showed that integrating a flow-distributing mesh 1.5 cm upstream from the construct compartment imposed an equal medium flow and shear stress of 0.6 dynes /cm(2) along the entire cell construct cross-section area. The design of 95.8%open-pore-area fixing nets enabled the exposure of 99.88% of the cell construct volume to the perfusing medium. Cardiac cell constructs seeded with physiologically relevant cell density (0.7 x 10(8) cells/cm(3)) in alginate scaffolds developed into homogenous compacted cardiac tissue, as judged using cell staining with fluorescein diacetate and hematoxylin-eosin histology. The cell constructs maintained 80% viability for nearly 2 weeks, whereas in static-cultivated cell constructs, only 50% of the initial cells remained, as determined according to total DNA content and MTT viability assay. Medium perfusion resulted in better cell viability, presumably due to the convective-diffusive transport of oxygen, compared with oxygen diffusion within the static-cultivated cell constructs, as well as due to efficient removal of harmful cell secretions. It is envisioned that this bioreactor would be useful for 3D cultivation of different mammalian cells for purposes of tissue engineering or production of valuable biologicals.
我们开发了一种新型灌注生物反应器，它能够在一个总横截面积为20平方厘米的流动腔室中培养多个三维（3D）细胞构建体。该生物反应器集成的两个独特特征可使流体沿生物反应器横截面均匀流动，并使细胞构建体最大程度地接触灌注培养基。对灌注生物反应器中流体流动状态的数学建模表明，在构建体隔室上游1.5厘米处集成一个流量分配网，可使整个细胞构建体横截面积上的培养基流量和剪切应力均为0.6达因/平方厘米。开孔面积为95.8%的固定网设计可使99.88%的细胞构建体体积接触到灌注培养基。在藻酸盐支架中接种具有生理相关细胞密度（0.7×10⁸个细胞/立方厘米）的心脏细胞构建体，通过用二醋酸荧光素染色和苏木精-伊红组织学判断，其发育成了均匀致密的心脏组织。根据总DNA含量和MTT活力测定，细胞构建体在近2周内保持80%的活力，而在静态培养的细胞构建体中，仅50%的初始细胞存活。与静态培养的细胞构建体中的氧气扩散相比，培养基灌注可提高细胞活力，这可能是由于氧气的对流扩散运输，以及有效去除有害细胞分泌物。可以设想，这种生物反应器将有助于用于组织工程或生产有价值生物制品的不同哺乳动物细胞的3D培养。
