Plasma-Free Conditions and Long Term Storage

In vitro generation of artificial red blood cells (RBCs) is very important to overcome insufficient and unsafe blood supply. Despite recent progresses in RBCs engineering from several stem cell sources, none of them could succeed in generation of functional RBCs in the absence of serum/plasma and feeder cells. Without the elimination of serum and plasma, human RBC engineering in a large scale is impossible, especially for the future bioreactor system. Using an appropriate combination of cost-effective and safe reagents, the present study demonstrated the terminal maturation of hematopoietic stem cells into enucleated RBCs, which were functional comparable to donated human RBCs. Surprisingly, the viability of erythroid cells was higher in our serum-and feeder-free culture condition than in the previous serum-added condition. This was possible by supplementation with vitamin C in media and hypothermic conditions. Also, our report firstly presents the storability of artificial RBCs, which possibility is essential for clinical application. In summary, our report demonstrates engineering of human applicable RBCs with a dramatically enhanced viability and shelf-life in both serum-and stroma-free conditions. This innovative culture technology could contribute to the realization of the large-scale pharming of human RBCs using bioreactor systems.We thank So-Young Lee and Jung Nyu Chun for technical support. This work was supported by a grant from the National Research Foundation of Korea (2009-0070154, 2010-0011200) and Korean Health Industry Development Institute (A101782)