Mechanisms Of Eosinophils Promote Hematopoietic Stem Cells Mobilization

IntroductionAsthma is a chronic airway inflammation disease induced by allergen and characterized by several kinds of cells and cellular components. Cells involved in this pathological process include eosinophil, mast cell and T cell and so on. The clinical features of asthma are airway hyperresponsiveness and reversible flow limitation. Eosinophils appear to be the key effector cells in asthma, and there is a positive correlation between increased numbers and activation of eosinophils and the severity of asthma. According to Global Asthma Report 2014, the most recent revised global estimate of asthma suggests that as many as 334 million people have suffered this disease among which 40 million patients are in China. Asthma imposes high social and economic costs. The current effective treatment is inhaled corticosteroids (ICS) which is not a clinical cure. Pathological cellular mechanisms and control measures are very important in this research field. Accordingly, developing the roles of eosniophil and anti-eosinophil therapy is still the key point in asthma research.Eosinophil is an end-stage blood cell which is characterized by aligned acidophil granules full of cytoplasm. It derives from the bone marrow and circulates at low levels in the peripheral blood in healthy individuals. The normal level of eosinophil is 0.5%-5%of total white blood cell. Despite of very low count in blood, it plays a very important role, especially in the pathological process of asthma. There are many researches about the pathophysiological roles in asthmatic inflammation such as: Eosinophils are recruited to the local airway, releasing specific granule proteins like MBP-1 and ECP, which contribute to tissue damage, especially to the epithelial cells; Secretion of IL-13 and IL-4 from eosinophils engages in and promotes the airway inflammation, goblet cell hyperplasia, mucin hyper-secretion as well as AHR; Eosinophils also secrete TGF-β which mediates smooth muscle cells hyperplasia and collagen deposition which promote airway remodeling.In addition to these, eosinophils have important roles in other sides. For example, it promotes host defence against helminthes through destroying the cell wall of herminth with MBP-1 and secreting toxic anti-helminth antibody; Eosinophils can perform numerous immune functions including antigen presentation, activating mast cells and regulating T cell polarization and function; Eosinophils can also recruit CD8+ T cells to the tumor location and suppress tumor growth.Eosinophil is differentiated from hematopoietic stem cells (HSC) under a complex regulation by several cytokines and transcription factors. The relationship between these two cell types have not been reported yet. In our previous study, we found that stem cells were activated to be LSK (lineage-Sca-l+c-kit+), and mobilized to the peripheral spleen in the asthmatic model. However, in eosinophil-deficiency mice (Phil), the mobilization was decreased. This suggested that eosinophil might have a regulation role on its superior stem cells. Based on this hypothesis, we designed a series of bone marrow transplantation and functional experiment as well as co-culture system with Phil and IL-5 transgenic (NJ.1638) mice to explore the relationship between eosinophil and stem cells, trying to detect the underlying mechanisms.ObjectiveDetect that epsinophils promote stem cell mobilization and develop the mechanisms.Methods1. Animal modelEstablish a classical asthma model on 6-10 weeks old WT and Phil mice with ovabulmin (OVA). Analyze the activated stem cells LSK of bone marrow and spleen with a multiparameter flow cytometry method. The generation of NJ.1638 and Phil were used to do the LSK analysis. Establish a bone marrow mobilization model with G-CSF in WT and Phil mice to detect the LSK level with or without eosinophils.2. Bone marrow transplantation (BMT)Total bone marrow cells from WT and NJ.1638 mice were used as donor cells to establish a non-competitive transplant experiment. LSK sorted form WT and NJ.1638 mice were used as donor cells to establish a competitive LSK transplant experiment. Homing ability of LSK from WT and NJ.1638 mice was detected at 16 h after co-transplant. Reactive Oxygen Species (ROS) in LSK was marked by a probe DAFH DA with a green fluorescence under detection. An in vitro eo-culture system was established to develop the relationship of eosinophils and LSK.Results1. In eosinophil-deficiency Phil mice, stem cell mobilization to the spleen was decreased.2. LSK count in bone marrow of NJ.1638 mice was significantly decreased while increased in spleen. The ROS level of NJ.1638 LSK was extremely higher than that in WT mice.3. The stem cell mobilization in generation of NJ.1638 and Phil crossed mice which performed high IL-5 and deleted eosinophil was eliminated, which indicated that the number of eosinophils is very important in the stem cell mobilization. 4. Difficiency of eosinophils did not affect the G-CSF-bone marrow mobilization.5. Bone marrow transplant revealed that the number of eosinophil is closed related to the stem cell mobilization, and LSK from NJ mice displayed decreased differentiation ability.6. ROS level of NJ.1838 LSK was high while the homing ability was not affected.7. Eosinophil can stimulate higher ROS level in LSK in vitro.ConclusionIncreased number of eosinophils promoted stem cell mobilization which might through ROS accumulation.