A Study On The Changes And Significances Of Neutrophil Extracellular Traps In The Peripheral Blood From Patients With MPO-ANCA Associated Vasculitis

Objective To investigate the level of neutrophil extracellular traps(NETs)in patients with anti-neutrophil cytoplasmic myeloperoxidase antibody-related vasculitis(MPO-AAV)and its relationship with innate immunity and adaptive immunity.To explore the relationship between the changes of NETs level and the damage of MPO-AAV organ function and disease activity.Methods 79 newly diagnosed patients with MPO-AAV were selected,including 72 MPA patients(91.14%),7 GPA patients(8.75%),male: female = 37:42,and age65.09±14.30(51-79)years old.Among the major organ damage,64(81.01%)were kidney damage,63(79.74%)were lung damage,and the course of disease was 0.5-180([11,16])months.The BVAS score was 19.00±7.52(11-27)points.A healthy control group of 40 patients was selected,male: female = 17:22,age 63.05±11.81(52-75 years old).Statistical test results showed that there was no significant difference in the distribution of gender and age between the two groups(P>0.05),indicating that the two groups of patients were comparable.Simultaneous detection of NETs(MPO-DNA levels)(Opticaldensity,OD values)in peripheral blood of case and healthy controls using enzyme-linked immunosorbent assay(ELISA).Flow cytometry(FCM)was used to detect the expression of Tfh in peripheral blood of patients and controls(percentage of CXCR5+CD4+T cells).The level of Ba and the level of MPO-ANCA were measured by ELISA.Clinical data including Birmingham Vasculitis Activity Score(BVAS)were collected.At last,we compared the expression differences of the various indicators between the two groups,and analyzed the correlation between these indexes in MPO-AAV patients,and detected the relationships between NETs,Tfh,Ba and the clinical damages.Results1.The levels of NETs in peripheral blood of patients in the case group were significantly higher than those in healthy controls(0.545(0.383,0.681)vs 0.216(0.171,0.237),t =-6.706,P<0.001);and NETs levels and complement The degree of activation of the complement pathway(Ba level),expression of follicular helper T cells(percentage of c Tf H cells),MPO-ANCA and percentage of neutrophils in peripheral blood,BVAS score,lung BVAS score and kidney BVAS score were positively correlated(r=0.342,P<0.001;r=0.387,P<0.001;r=0.272,P=0.016;r=0.314,P=0.005;r=0.361,P<0.001;r=0.304,P=0.007;r=0.233,P=0.030).2.The expression levels of c Tfh cells,ICOS+c Tfh cells,and PD-1+c Tfh cells in the case group were higher than those in the control group(respectively 25.79±3.72 vs19.98±4.72,t=7.66,P<0.001,1.83±1.06 vs 0.82±0.60,t= 4.55,P<0.001,10.10±2.85 vs 8.17±2.66,t= 2.45,P=0.018);The expression level of c Tfh cells was positively correlated with MPO-ANCA levels(r=0.761,P<0.001).and the expression of c Tfh cells and ICOS+c Tfh cells were positively correlated with the BVAS score,(respectively r = 0.384,P = 0.037;r = 0.455,P = 0.015).3.Compared with the healthy control group,the level of Ba in the patient group was significantly higher(122.05(89.41,204.26)vs 35.12(14.69,65.02),t=-6.756,P<0.001),and was positively correlated with the BVAS score.(r = 0.383,P <0.001).4.There was a positive correlation between MPO-ANCA level and BVAS score(r=0.359,P=0.027).Conclusion1.The neutrophil extracellular traps(NETs)in patients with MPO-AAV was significantly increased.2.Increased NETs may promote the proliferation and differentiation of follicular helper T cells,and the increase number and activity of follicular helper T cells will assist B cells to differentiate into plasma cells to produce specific antibodies MPO-ANCA participate in MPO-AAV patient-specific immunity.3.Increased NETs may be involved in activation of the complement pathways system,attracting,pre-activation of more neutrophils,and degranulation occurs with the complement of the neutrophil membrane complement C5 a receptor involved in innate immunity of MPO-AAV.4.This study supports the activation of complement pathways involved in the pathogenesis of MPO-AAV.MPO-ANCA may act as pathogenic antibodies and participate in the pathogenesis and clinical damage of MPO-AAV.5.The level of NETs in blood around MPO-AAV patients is significantly increased,which may promote the pathogenesis of MPO-AAV by enhancing adaptive immunity and innate immunity,respectively.Therefore,inhibition of NETs levels may be one of the new potential therapeutic approaches for MPO-AAV.