Research On Therapeutics And Mechanisms Of Takayasu Arteriti

Part Ⅰ Comparison of the efficacy of 3 immunosuppressants monotherapy for maintenance treatment in Takayasu arteritisBackground and Objective Takayasu arteritis(TAK)presents high rates of relapse.However,the optimal therapeutic agents for the maintenance of disease remission remain unclear.This study aimed to investigate the efficacy and safety of 3 different conventional immunosuppressants,as maintenance therapy,in preventing relapse during the remission period under real-world conditions.Material and Methods The present study was a single-center,retrospective cohort study.TAK patients who were in remission and were given glucocorticoids(≤10mg/day prednisone or equivalent)and immunosuppressant monotherapy were included.The beginning of taking immunosuppressant monotherapy during the remission phase was taken as the baseline time point.Demographic data,clinical characteristics,treatment regimen,and follow-up data were collected.Study outcomes were defined as relapse or no relapse.The efficacy of maintenance treatment was compared among mycophenolate mofetil(MMF),azathioprine(AZA)and methotrexate(MTX).Univariate survival curves were drawn by the Kaplan-Meier methods,and the statistical difference between survival curves was assessed by the Log-rank test.Univariate Cox regression analysis was used to screen the risk factors of TAK recurrence.Multivariate Cox proportional hazards regression analysis was used to adjust for confounders and calculate the hazard ratio(HR)of each factor with 95%confidence intervals(CI).Results A total of 232 cases who met the inclusion criteria were enrolled in the study,207 were females(89.2%)and the median(IQR)onset age was 26.0(20.9-31.6)years.Sixty cases used MMF(25.9%),75 used AZA(32.3%),and 97 used MTX(41.8%).The median(IQR)follow-up period from baseline to the study’s endpoint was 26.8(16.7-44.5)months.A total of 69 cases experienced recurrence during the follow-up period and their median(IQR)time to recurrence was 19.1(11.1-26.4)months.The cumulative recurrence rate at 26.8-month follow-up was 27.2%.No significant difference was observed in the major relapse rate among the 3 groups(P=0.962).Using MMF as the reference group,Kaplan-Meier survival curves revealed a significant difference among the 3 groups(Log-rank P=0.007).The estimated cumulative recurrence rate for MMF,AZA and MTX at 24-month follow-up was 35.9%,16.1%and 17.8%,respectively,and 57.3%,35.6%and 34.7%,respectively,at 48-month follow-up.After adjusting for confounders,the severity of vascular involvement was an independent risk factor for TAK relapse(HR=1.048,95%CI 1.001-1.096,P=0.044).After adjusting for the severity of vascular involvement,AZA showed a 54.9%decreased risk of relapse(HR=0.451,95%CI 0.249-0.818,P=0.009),and MTX showed a 51.7%decreased risk of relapse(HR=0.483,95%CI 0.277-0.844,P=0.011),compared with MMF.During the follow-up period,72 cases attempted to taper off glucocorticoids.Of them,9 cases relapsed(12.5%),and 55 cases achieved glucocorticoid-free remission(23.7%of all cases),Twelve cases withdrew non-glucocorticoid immunosuppressants and retain low-dose glucocorticoid monotherapy,and 4 cases of them relapsed(33.3%).During follow-up,adverse reactions were mainly recorded as an abnormal hepatic function for MTX(7.2%)and MMF(1.7%),and myelosuppression for AZA(5.3%).Conclusions Compared with MMF,AZA and MTX showed significantly better relapse prevention as maintenance therapies while in the remission periods of TAK.The severity of vascular involvement was an independent risk factor for TAK relapse.Nearly a quarter of cases could achieve glucocorticoid-free remission during remission maintenance periods.No severe adverse drug reaction occurred with MMF,AZA and MTX during treatment.Part Ⅱ Altered glycosylation profiles of serum IgG in Takayasu arteritisBackground and Objective Takayasu arteritis(TAK)is an autoimmune inflammatory disorder with an undefined etiology.The lectin microarray is a novel tool for glycan analysis that enables obtaining global glycosylation patterns in a rapid and highly sensitive way.This study aimed to characterize the glycosylation profiles of serum immunoglobulin G(IgG)in patients with TAK.Material and Methods The discovery cohort included 164 patients with TAK(82 active and 82 inactive),128 patients with atherosclerosis used as disease controls(DCs),and 100 healthy controls(HCs).Lectin microarrays containing 56 types of lectins were used to detect the glycan levels of serum IgG.Differentially altered glycosylation patterns between TAK and control groups as well as between TAK subgroups were identified and further validated by lectin blot.The classification performance of the TAK-specific glycosylation change was measured by receiver-operating characteristic(ROC)curve analysis.Results Lectin microarray analysis revealed significantly increased N-Acetylgalactosamine(GalNAc)levels in the TAK group compared to the DC and HC groups(all P<0.01).No significant difference was found between the DC and HC groups(P>0.99).For TAK subgroups,significantly decreased mannosylation was observed in patients with active TAK compared to patients with inactive disease(P<0.01).These differences were validated by lectin blot.In addition,GalNAc levels exhibited a considerable potential for discriminating patients with TAK from patients with atherosclerosis,with an area under the curve of 0.749(P<0.001),a sensitivity of 71.7%,and a specificity of 73.8%.Conclusions Serum IgG in patients with TAK displayed disease-specific glycosylation alterations.Aberrant GalNAc glycosylation showed substantial value as a diagnostic biomarker.The potential pro inflammatory properties of the abnormal glycans may provide new insights into the role of humoral immunity in the pathogenesis of TAK.Part Ⅲ Single-cell RNA sequencing revealed increased CD160 expression on circulating natural killer cells in untreated Takayasu arteritisBackground and Objective Sustained activation of the innate and adaptive immune responses cooperatively lead to the pathogenesis of Takayasu arteritis(TAK).However,the research regarding the role of innate immune cells in TAK pathogenesis is currently limited.The present study performed single-cell RNA sequencing using peripheral blood mononuclear cells(PBMC)of untreated patients with TAK and aimed to primarily explore the alterations of immunological profiles and function of immune cells in the early stages of TAK.Material and Methods PBMC were collected from 5 untreated TAK patients and 2 gender-and age-matched healthy controls.10× Chromium single-cell platform(10× Genomics)was used to perform single-cell RNA sequencing.Bioinformatics tools were used to conduct reduction analysis and clustering analysis and visualize single-cell RNA-seq data.The immunological profiles of immune cells in PBMC were described.Differential gene expression analysis was performed between untreated TAK and healthy controls for each cell cluster.Flow cytometry was used to validate the selected candidate gene in untreated TAK,treated-active TAK,treated-inactive TAK patients and healthy controls.Results In total,80791 cells from 7 samples were retained for downstream analysis after quality control and filtration.Dimensionality reduction analysis identified 23 major clusters of 4 main categories.Data of T/Natural killer(T/NK)cells,B cells,and Monocytes/Dendritic cells were extracted separately and further analyzed.The fraction of regulatory T cells(Tregs),type 1 classical dendritic cells(cDC1)tended to decrease,and that of interferon(IFN)-activated B cells,plasma cells,CD56bright NK cells,CD1cCD141-dendritic cells,and plasmacytoid dendritic cells(pDC)tended to increase in untreated TAK.CD160 expressions on CD56dim NK cells were significantly higher in untreated TAK than that in healthy controls,and this result was verified using flow cytometry.Conclusions Compared with healthy controls,there are imbalances in the proportion of multiple peripheral immune cells in untreated TAK,which might be a possible reason for disturbed immune regulation.Although the fraction of cytotoxic CD56dim NK cells is decreased in PBMC of untreated TAK,the CD160 expression is remarkably increased.The overexpression of CD160 might lead to enhanced secretion of pro-inflammatory cytokines,such as IFN-γ,and thus participates in TAK pathogenesis.