Prolonged shearing of insect cells in a couette bioreactor

Abstract

Viscometers have had a prominent role in the study of hydrodynamic damage to cell cultures. A Couette bioreactor overcomes stringent time limits of previous viscometric research. At low shear levels, the vessel supported robust growth of Spodoptera frugiperda cells (>92% viability) at 0.64±0.09 day −1 to a maximum cell density of 6.1×10 6 cells ml −1. The intrinsic rate of necrosis was 30+ times slower than the specific growth rate. Cell death was bimodal, confirming recent reports of apoptosis in uninfected insect cells. Population dynamics suggests that early apoptotic cell formation accelerated 100-fold over a month, surpassing the rate of necrosis by day 4. Early apoptosis was the rate-limiting step in the apoptotic pathway and particularly sensitive to culture conditions. Accepted methods of estimating shear exposure were revised to account for the pseudoplasticity of Couette cultures: power-law parameters m=0.660±0.054 dyne s 0.22 cm −2 and n=0.222±0.047. The maximum shear stress of 0.84 dynes cm −2 was 70+ times the value predicted for a Newtonian fluid (1.0 cp viscosity). Prolonged shearing in a Couette bioreactor will enable investigation of cumulative stress effects and the shear response of cell processes with a long half-life.