Expression Of ProBDNF/p75NTR Signal In Antibody Secreting Cells Of SLE Patients And Mice

Background:Brain derived neurotrophic factor precursor（proBDNF）is a neurotrophic factor widely distributed in the central nervous system.It has different biological functions from mature BDNF.In recent years,studies have found that proBDNF and its high affinity receptor,neurotrophic receptor p75（p75NTR）are widely expressed in the immune system and have a certain immune regulation function.It plays an important role in inflammatory related diseases such as aortic dissection,sepsis,multiple sclerosis and depression.Systemic lupus erythematosus（SLE）is a typical chronic autoimmune disease.The main feature of the disease is the over activation of B cells and the production of a large number of antibody secreting cells（ASCs）,which leads to the secretion of a variety of autoantibodies and the damage of corresponding target organs.Therefore,exploring the inhibition target of ASCs is notable subjects of science research.Objective:To explore the expression of proBDNF/p75NTR signal in ASCs in SLE patients and animal models,and its correlation with the development of the SLE disease.Method:1)Blood samples were collected from SLE patients and their matched healthy donors（HDs）,then human PBMCs were isolated by lymphoprep separation liquid.The flow cytometry was used to measure the proportion of CD19⁺B cells and its subsets,as well as the expression of proBDNF and p75NTR.Meanwhile,we collected patients’relevant clinical data,and the correlations were analyzed between proBDNF MFI on ASCs and clinical data,including autoantibody titer,complement 3（C3）,erythrocyte sedimentation rate（ESR）,C-reactive protein（CRP）and SLE disease activity score（SLEDAI）.2)Six-week-old female C57BL/6 mice were divided into two groups:lupus group were immunized intraperitoneally with 0.5 m L pristane for induction of lupus in rodents;control group were intraperitoneally injected with 0.5 ml saline.After 8 weeks of immunization,spleens and lymph nodes were harvested from pristane immunized C57BL/6J mice and the controls.Flow cytometry were used to detect the expression of proBDNF and p75NTR in B cells and ASCs(CD44^hiCD138⁺)in spleen.Meanwhile,we evaluated the proBDNF level in spleen by Western Blot.Immunofluorescence were perfromed to measure the expression of proBDNF and p75NTR on B cells in lymph nodes.Results:1)Compared to HD,the percentage of CD19⁺B cells（P<0.01）and ASCs（P<0.01）in SLE patients was increased,and the expression of proBDNF（P<0.001）and p75NTR（P<0.01）in ASCs was increased.In SLE patients,the proBDNF（P<0.05）and p75NTR level（P<0.001）in ASCs was higher than that in CD19⁺B cells,memeroy B cells and naive B cells.The proBDNF expression on ASCs is related to whether the autoantibody is positive or not.And the proBDNF MFI on ASCs has a positive correlation with ESR（r=0.3821,P=0.0080）,CRP（r=0.5306,P=0.0004）and SLEDAI（r=0.481,P=0.0001）,and a negative correlation with C3（r=-0.3584,P=0.0106）.2)Compared to control mice,the proBDNF level on splenic CD19⁺B cells（P<0.05）and ASCs（P<0.05）of lupus mice were increased,and p75NTR level on splenic CD19⁺B cells（P<0.05）and ASCs（P<0.05）of lupus mice were increased;the results of Western Blot indicated the increased proBDNF expression in spleen of lupus mice compared to control mice.And immunofluorescence showed that the expression of proBDNF and p75NTR on B220⁺B cells were increased in lympho node of SLE mice compared to control mice.Conclusion:1)In the peripheral blood of patients with systemic lupus erythematosus,proBDNF and p75NTR in ASCs are up-regulated.The expression of proBDNF in ASC is positively correlated with disease activity,suggesting that it may become a clinical marker of disease activity.2)In the lupus model induced by pristane,the expression of proBDNF/p75NTR in B cells and ASCs was up-regulated,which provided a research basis for subsequent animal experiments.