Function Of Delayed Neutrophil Apoptosis By Fc?R In RA

Objective: To identify the apoptotic status and Fc?R expression of neutrophil in synovial fluid and peripheral blood from RA patients,and investigate the role of Fc?R expression of neutrophil in its apoptosis.Methods: 22 RA patients and 11 healthy control were enrolled from March 2016 to March 2017 in the Department of Rheumatology and Immunology,West China Hospital,Sichuan university.The peripheral blood(PB)and synovial fluid(SF)were collected from healthy control or RA patients.In order to isolate and purify neutrophil(PMN),density gradient centrifugation was adopted by ficoll-Hypaque Solution and ice pure water.BD FITC AnnexinV-Apoptosis Detection Kit I was used to detect apoptotic rate of SF PMN,PB PMN from RA patients,and PB PMN from HC at different time points(4 h,8 h,12 h and 24 h).HC neutrophil was co-cultured with 50% RA SF and apoptotic rate was measured afterwards.APC-H7 anti-human CD16(Fc?RIII),PE anti-human CD16b(Fc?RIIIb),FITC Anti-human CD32(Fc?RII)and BV510 Anti-human CD64(Fc?RI)were applied to detect Fc?R expression in neutrophil from RA patients' PB,SF or HC PB.Fc?R expression was also detected after HC and RA neutrophil was co-cultured with 50% RA SF.The demographic data(gender and age)was analyzed by Fisher definite probability method and Mann-Whitney U Nonparametric rank sum test.The AnnexinV positive rate was analyzed by Two-way ANOVA,and the Post-test was performed by Bonferroni.The Fc?R expression was analyzed by One-wayANOVA,and the Post-test was performed by Bonferroni.Results:(1)There was no difference on gender proportion and age proportion among RA SF group,RA PB group and HC PB group.And there was no difference on course of disease and disease activity between RA SF group and RA PB group.(2)Compared with HC PB PMN,RA patients' PB PMN displayed delayed apoptosis.There was no difference in neutrophil apoptosis between RA patients' SF and PB.(3)The apoptotic rate of PMN from HC or RA peripheral blood and synovial fluid at different time points(4 h,8 h,12 h and 24 h)was showed as following: HC PB PMN 49.2±8.5%,59.3±9.1%,66.5±8.3% and 79.0±6.3%;RA PB PMN 38.3±12.3%,44.0±4.5%,38.4±8.0% and 51.4±8.7%;RA SF PMN 36.9±7.7%,41.9±8.5%,42.9±8.7%and 44.7±9.1%.(4)RA SF can significantly inhibit the apoptosis of HC PB PMN(P<0.01).Synovial fluid incubation could reduce the apoptotic rate of HC PB PMN to 51.7±7.1%(4 h),53.1±6.8%(8 h),58.7±8.9%(12 h)and65.9±6.7%(24 h).(5)Fc?RIIIb was highly expressed in PMN from HC or RA peripheral blood,and no difference of Fc?R expression was found between these two groups(P>0.05).Compared with RA PB PMN,the expression of Fc?RI on RA SF PMN was increased(P<0.05).RA SF PMN showed Fc?RIIIbhigh and Fc?RIIIblow populations,and the proportion of Fc?RIIIbhigh was lower than Fc?RIIIblow(P<0.05).(6)RA SF can influence the sub-population expression of Fc?RIIIb on HC and RA PB PMN: the proportion of Fc?RIIIblow was promoted and the formation of Fc?RIIIbhigh was inhibited(P<0.05).(7)RA SF can inhibit Fc?RII expression on HC and RA PB PMN(P<0.05).Conclusions:(1)Neutrophil apoptosis in synovial fluid and peripheral blood of RA patient was delayed compared to HC control.(2)The synovial fluid of RA patient can inhibit the apoptosis of neutrophil.(3)There was difference in the Fc?R expression of PMN from RA PB and SF,suggesting that Fc?R maybe involved into the apoptosis.