Hammond T G, Hammond J M
Nephrology Section, Tulane University Medical Center, Louisiana Veterans Research Corporation, and Veterans Affairs Medical Center, New Orleans 70112, USA.
Am J Physiol Renal Physiol. 2001 Jul;281(1):F12-25. doi: 10.1152/ajprenal.2001.281.1.F12.
Suspension culture remains a popular modality, which manipulates mechanical culture conditions to maintain the specialized features of cultured cells. The rotating-wall vessel is a suspension culture vessel optimized to produce laminar flow and minimize the mechanical stresses on cell aggregates in culture. This review summarizes the engineering principles, which allow optimal suspension culture conditions to be established, and the boundary conditions, which limit this process. We suggest that to minimize mechanical damage and optimize differentiation of cultured cells, suspension culture should be performed in a solid-body rotation Couette-flow, zero-headspace culture vessel such as the rotating-wall vessel. This provides fluid dynamic operating principles characterized by 1) solid body rotation about a horizontal axis, characterized by colocalization of cells and aggregates of different sedimentation rates, optimally reduced fluid shear and turbulence, and three-dimensional spatial freedom; and 2) oxygenation by diffusion. Optimization of suspension culture is achieved by applying three tradeoffs. First, terminal velocity should be minimized by choosing microcarrier beads and culture media as close in density as possible. Next, rotation in the rotating-wall vessel induces both Coriolis and centrifugal forces, directly dependent on terminal velocity and minimized as terminal velocity is minimized. Last, mass transport of nutrients to a cell in suspension culture depends on both terminal velocity and diffusion of nutrients. In the transduction of mechanical culture conditions into cellular effects, several lines of evidence support a role for multiple molecular mechanisms. These include effects of shear stress, changes in cell cycle and cell death pathways, and upstream regulation of secondary messengers such as protein kinase C. The discipline of suspension culture needs a systematic analysis of the relationship between mechanical culture conditions and biological effects, emphasizing cellular processes important for the industrial production of biological pharmaceuticals and devices.
悬浮培养仍然是一种流行的培养方式，它通过控制机械培养条件来维持培养细胞的特定特性。旋转壁式生物反应器是一种优化的悬浮培养容器，旨在产生层流并将培养中细胞聚集体上的机械应力降至最低。本综述总结了有助于建立最佳悬浮培养条件的工程原理以及限制该过程的边界条件。我们建议，为了将机械损伤降至最低并优化培养细胞的分化，悬浮培养应在诸如旋转壁式生物反应器之类的固-体旋转库埃特流、零顶空培养容器中进行。这提供了流体动力学操作原理，其特点是：1）绕水平轴的固体旋转，其特点是不同沉降速率的细胞和聚集体共定位、流体剪切和湍流最佳降低以及三维空间自由度；2）通过扩散进行氧合。悬浮培养的优化是通过应用三个权衡来实现的。首先，应通过选择密度尽可能接近的微载体珠和培养基来使终端速度最小化。其次，旋转壁式生物反应器中的旋转会产生科里奥利力和离心力，这两者直接取决于终端速度，并随着终端速度的最小化而最小化。最后，悬浮培养中营养物质向细胞的质量传输取决于终端速度和营养物质的扩散。在将机械培养条件转化为细胞效应的过程中，有几条证据支持多种分子机制发挥作用。这些机制包括剪切应力的影响、细胞周期和细胞死亡途径的变化以及诸如蛋白激酶C等二级信使的上游调节。悬浮培养学科需要对机械培养条件与生物学效应之间的关系进行系统分析，重点关注对生物制药和生物装置工业生产重要的细胞过程。
