Establishment of neutrophil-lineage stem cells from C57BL/6 mice.

Neutrophils are key effector player of the innate immune system that are rapidly recruited to infected tissues to clear pathogens. Neutrophils also have the capacity to engulf damaged tissue cells to clear, although at the same time neutrophil granules including myeloperioxidase, elastase and ROS may damage tissues. We have shown previously that neutrophils help to repair wounded tissue. However, neutrophils are already differentiated cells, which cannot divide. For future personalized stem cells therapy, it is important to establish neutrophil-precursor cells, which will differentiate to mature neutrophils to engulf pathogens or damaged cells by the transplantation in vivo. First we characterized 32Dcl3 cell line, which has been established previously by WEHI-3 conditional medium. These cell lines differentiated to neutrophils by the stimulation of G-CSF. Then we established similar clone from C57BL/6 mice by culturing bone marrow cells in WEHI-3 conditional medium. We could obtain a neutrophil-precursor cell line (B6NPC), which grows more than six months in WEHI-3 conditional medium. By FACS analysis, we found that both 32Dcl3 and B6NPC had GR-1, CD11b and F4/80 cell surface markers, which are myeloid-lineage markers. However, these cells also have early T cell markers, which fit the hypothesis of myeloid-based model of hematopoietic cell differentiation. By the transplantation of B6NPC to syngeneic mice at the same time of wounding, we found early recovery from wound healing.