Supplementation of Phosphatidylcholine Protects the Hybridoma Cells from Apoptosis Induced by Serum Withdrawal

Abstract

Serum withdrawal from the culture medium was reported to retard the cell growth and induce apoptosis. Recently, it was demonstrated that serum withdrawal caused an elevation in cellular ROS levels, and induced apoptosis cell death in human U397 cells. ROS such as$\mathop {\rm{O}}\nolimits_{\rm{2}}^{ \bullet - }$ and H₂O₂ are important mediators of intracellular signaling pathway. Phosphatidylcholine (PtdCho) is an important phospholipid of eukaryotic cell membrane; it also plays an important role in signal transduction. It is synthesized via the cytidine diphosphocholine (CDP-choline) pathway, and the PtdCho deficiency due to the inhibition of this pathway is one of the factors that triggers apoptosis in many cell lines. The presence of exogenous H₂O₂ was reported to strongly inhibit the PtdCho synthesis and thus caused apoptosis in a human microglial cell line. Hence, there may be a possible link between the serum withdrawal and PtdCho synthesis and transduction pathway. Thus, in this study, the effect of PtdCho supplementation on the apoptotic cell death induced by the serum withdrawal was investigated. In the present study, 100 μg/ml of PtdCho was supplemented to the hybridoma culture in the presence of serum and under serum withdrawal conditions. Under the serum withdrawal condition, the culture supplemented with PtdCho was able to maintain about 30% viability for over 6 days, while the viability of the culture without PtdCho supplementation has decreased to about 10%. In the serum supplemented culture, the presence of exogenous PtdCho was able to enhance the survivability of hybridoma at the death phase of the culture. The viability of the PtdCho culture was maintained at about 35% for over 8 days, while the viability of the control culture has dropped to about 2%. The majority of the dead cells in the control culture were identified as apoptotic cells. No effect on the specific growth rate, maximum cell density and monoclonal antibody productivity were observed in the culture supplemented with PtdCho. The results of the present study showed that PtdCho is able to protect the hybridoma cells from apoptosis triggered by serum withdrawal.