Inner Autonomous Feedback Loop In Hematopoietic System Regulates Dormant And Active Status Of Hematopoietic Stem Cells

Traditional models for hematopoietic stem cell transplantation(HSCT) require myeloablation by whole body irradiation or chemical treatment. However,these ablation approaches disturb the functions of non-hematopoietic cells and even introduce potential tumorigenesis while achieving successful HSCT. It is unknown whether specific ablation of hematopoietic cells is a sufficient conditioning method for HSCT. Further, it remains elusive that whether an inner feedback loop exists in hematopoietic system that can balance the status of HSCs. For the scientific questions, we used a novel approach to produce hematopoietic cells with the Diphtheria Toxin Receptor (DTR)-GFP reporter, which can be specifically killed in the presence of DT in vivo, and allowing hematopoietic-specific ablation without killing non-DTR positive blood cells and non-hematopoietic tissue cells. By this model, we proved that the hematopoietic-specific ablationimmediately activated hematopoietic stem cells from dormancy to activate. Our myeloablationapproaches for mouse BMT can replace the traditional myeloablation by whole body irradiation or chemical treatment.Briefly, a Diphtheria toxin receptor (DTR)-GFP reporter element was targeted into the ROSA26 locus to produce DTR-GFP reporter mice (LSL-DTR), with a loxp-stop-loxp cassette. Then the LSL-GFP reporter mice were crossed to Vav-Cre mice to produce double transgenic mice (DTR-GFP mice). We then transplanted DTR-GFP+Bone Marrow (CD45.2+) into sublethal irradiated recipients (CD45.1+) to achieve 50-80% chimeras in hematopoietic system. When the hematopoiesis in the recipients reached homeostasis four months after transplantation, we injected DT to ablate the DTR-GFP+white blood cells, without killing CD45.1+hematopoietic cells and other non-hematopoietic cells. Cell cycle results showed that the WT hematopoietic stem cells were immediately driven into cell cycle due to the inner loss of hematopoietic components, which indicated that an inner autonomous feedback mechanism existed in hematopoietic system and the dormant HSCs were directly activated by the stress loss of blood cells. Further experiments demonstrate that allogeneic donor’s hematopoietic stem cells were successfullytransplanted into the DTR chimeras after DT treatment.In summary, we established a novel approach toachieve the specific ablation of hematopoietic cells without killing non-hematopoietic tissue cells, and found that the inner autonomous feedback loop in hematopoietic system could regulatedormant and active status of hematopoietic stem cells. We developed a new approach to study the regulation of HSCs in vivo, and separate the total body conditions from HSCT.