Change Trend of Viablity, Apoptosis and ROS Production of Neutrophil in vitro Culture

Objective: To explore the change trend of cell viability, apoptosis and reactive oxygen species (ROS) production of peripheral blood neutrophils in in vitro culture.

Methods: Neutrophils were isolated by density gradient centrifugation and cultured in medium containing 10% fetal bovine serum for diffentrent times in a fall humidified atmosphere with 5% CO2 at 37 °C. Cell viability, spontaneous apoptosis and ROS level were detected by flow cytometry at 0, 24, 48, 72 and 96 hours.

Results: The cell viability of neutrophils could be maintained above 90% at 24 h in culture in vitro, and there was no significant difference between culture in vitro at 0 h and 24 h (P>0.05), but after culture in vitro at 72 h, the cell viability of neutrophils obviously droped, the count of apoptotic cells increased, and after culture in vitro at 96 h the neutrophils almostly all died and the production of ROS at 12 h significantly decreased as compared with that at 0 h.

Conclusions: The cell viability of neutrophils at 24 h cultures in vitro can be maintained relative stable with cell viability above 90% and unobvious change of count of apoptotic cells, but the neutrophil function test of ROS should be carried out immediately after cell separation.