The Role And Their Proliferation Mechanism Of Regulatory B Cells It Neonatal Sepsis

Research backgroundNeonatal sepsis,especially premature sepsis is a major cause of neonatal death,it is also one of the main challenges facing the global public health.More and more attention has been paid to the role of immune regulation in the pathogenesis of sepsis.In recent years,a group of B lymphocyte subsets regulatory B lymphocytes（Breg cells）have been discovered,which play a key role in the occurrence and development of autoimmune disease,allergic disease,organ transplantation infection,tumor and other diseases,but their role in neonatal sepsis is not clear.Therefore,in-depth study of the characteristics and regulatory mechanism of Breg cells in neonatal sepsis is expected to provide a more effective strategy for reducing the incidence and mortality of neonatal sepsis.Materials and methodsPeripheral venous blood was collected from the infants with sepsis（40 cases）at the time of onset,14 days and 21 days after the onset,and from the control group（40 cases）.Flow cytometry and ELISA were used to detect immune cells and their subgroups as well as plasma related cytokines.Meanwhile,the clinical severity of sepsis was assessed by neonate PELOD-2 score and the correlation with immune cells and subgroups was analyzed.The mRNA of peripheral blood was detected by RNA-seq,and the enrichment pathways of different genes were analyzed by GO and KEGG after qRT-PCR verification,and the mechanism of proliferation and differentiation of CD19⁺CD24^hiCD38^hi Breg cells was preliminarily explored.Finally,the mechanism involved in the proliferation and differentiation of CD19⁺CD24^hi CD38^hi Breg cells was further explored by isolating umbilical cord blood mononuclear cells and conducting induction culture in vitro.Results1.Peripheral blood B cells,especially CD19⁺CD24^hiCD38thi Breg cells subgroup,were significantly increased 7 days after the onset of sepsis and were negatively correlated with clinical severity;It was also found that the levels of IL-10 in plasma and CD19⁺CD24^hiCD38^hi Breg cells were significantly increased 7 days after the onset of sepsis.2.At the transcriptional level,significant differences were found in sepsis at different time points and the control group,while significant differences were found in sepsis at different time points,but no statistical significance was found in the control group.Through GO and KEGG enrichment pathways analysis,we found it to be closely related to the immune response and mTOR metabolic pathways.3.Compared with the control group,the level of BAFF in peripheral blood of sepsis patients was significantly increased at the onset and 7 days after the onset.The levels of BAFF-R had a tendency to increase in peripheral blood.Significantly,both the umbilical cord blood mononuclear cells and purified B cells were significantly increased after LPS and CpG stimulation.4.After LPS and CpG stimulation,B cells purified from umbilical cord blood mononuclear cells can significantly induce transformation into CD19⁺CD24^hiCD38^hi Breg cells.The BAFF combined with LPS and CpG stimulation can significantly improve the conversion rate of B cells to CD19⁺CD24^hi CD38^hi Breg cells.5.After BAFF combined with LPS and CpG stimulation,the improve conversion rate of B cells to CD19⁺CD24^hiCD38^hi Breg was related to the activation of PI3K-mTOR pathway and the enhancement of p70S6K and p4E-BP1 phosphorylation.ConclusionIn this study,it was found that peripheral blood CD19⁺CD24^hi CD38^hi Breg cells were significantly increased 7 days after the onset of sepsis in sepsis patients,and their ability to produce IL-10 in the cells was significantly enhanced.At the same time,the plasma IL-10 level was also significantly increased in the children with sepsis 7 days after the onset.Significantly,this study also found that the increased CD19⁺CD24^hi CD38^hi Breg cells were negatively correlated with the severity of the disease.Transcriptional and protein levels showed that the increased BAFF level in peripheral blood was related to the proliferation and differentiation of CD19⁺CD24^hi CD38^hi Breg cells by activating the PI3K-mTOR signaling pathway p70S6K and 4E-BP1.Therefore,this study not only revealed the changes and proliferative differentiation mechanism of CD19⁺CD24^hi CD38^hi Breg cells in the pathogenesis of sepsis,but also provided a new idea for the monitoring and immunotherapy of sepsis in neonates.