Effect of culture pH on erythropoietin production by Chinese hamster ovary cells grown in suspension at 32.5 and 37.0°C

Abstract

To investigate the effect of culture pH in the range of 6.85-7.80 on cell growth and erythropoietin (EPO) production at 32.5 and 37.0 degrees C, serum-free suspension cultures of recombinant CHO cells (rCHO) were performed in a bioreactor with pH control. Lowering culture temperature from 37.0 to 32.5 degrees C suppressed cell growth, but cell viability remained high for a longer culture period. Regardless of culture temperature, the highest specific growth rate (mu) and maximum viable cell concentration were obtained at pH values of 7.00 and 7.20, respectively. Like mu, the specific consumption rates of glucose and glutamine decreased at 32.5 degrees C compared to 37.0 degrees C. In addition, they increased with increasing culture pH. Culture pH at 32.5 degrees C affected specific EPO productivity (q(EPO)) in a different fashion from that at 37 degrees C. At 37 degrees C, the q(EPO) was fairly constant in the pH range of 6.85-7.80, while at 32.5 degrees C, the q(EPO) was significantly influenced by culture pH. The highest q(EPO) was obtained at pH 7.00 and 32.5 degrees C, and its value was approximately 1.5-fold higher than that at pH 7.00 and 37.0 degrees C. The proportion of acidic EPO isoforms, which is a critical factor for high in vivo biological activity of EPO, was highest in the stationary phase of growth, regardless of culture temperature and pH. Although cell viability rapidly decreased in death phase at both 32.5 and 37.0 degrees C, the significant degradation of produced EPO, probably by the action of proteases released from lysed cells, was observed only at 37.0 degrees C. Taken together, through the optimization of culture temperature and pH, a 3-fold increase in maximum EPO concentration and a 1.4-fold increase in volumetric productivity were obtained at pH 7.00 and 32.5 degrees C when compared with those at 37.0 degrees C. These results demonstrate the importance of optimization of culture temperature and pH for enhancing EPO production in serum-free, suspension culture of rCHO cells.