Study On Clinical Significance Of Long Noncoding RNA In Ischemic Stroke And Its Molecular Mechanism In Stroke-Associated Immunodepression

Part One Expression profile and clinical significance of long noncoding RNAs in acute ischemic strokePurpose Long noncoding RNAs(lnc RNAs)have been highlighted to be involved in the pathological process of ischemic stroke(IS).The purpose of this study was to investigate the expression profile of lnc RNAs in peripheral blood mononuclear cells(PBMCs)of acute IS patients and to explore their utility as biomarkers of IS.Methods Distinctive expression patterns of PBMC lnc RNAs were identified by a lnc RNA microarray and individual quantitative real time PCR(q RT-PCR)in four independent cohorts for 206 IS,179 healthy controls(HCs),and 55 patients with transient ischemic attack(TIA).A biomarker panel(lnc RNA-based combination index)was established using logistic regression.Results1.Lnc RNA microarray analysis showed 70 up-regulated and 128 down-regulated lnc RNAs in IS patients(fold-chang>2,P<0.05).2.Individual q RT-PCR validation demonstrated that three lnc RNAs(linc-DHFRL1-4,SNHG15 and linc-FAM98A-3)were significantly up-regulated in IS patients compared to HCs and TIA patients(P<0.05).3.Longitudinal analysis of lnc RNA expression up to 90 days after IS showed that lincFAM98A-3 normalized to control levels by day 7,while SNHG15 remained increased(P<0.05).4.Receiver-operating characteristic(ROC)curve analysis showed that the lncRNA-based combination index outperformed serum brain-derived neurotrophic factor(BDNF)and neuron-specific enolase(NSE)in distinguishing IS patients from TIA patients and HCs with areas under ROC curve of more than 0.84(P<0.05).5.The combination index increased significantly after treatment and was correlated with neurological deficit severity of IS(r=0.427,P<0.001).Conclusions The panel of these altered lnc RNAs was associated with ischemic cerebrovascular diseases and could serve as a novel diagnostic method.Part Two The role and molecular mechanism of long noncoding RNA SNHG15 in stroke-associated immunodepressionPurposeThis study was to explore the role and molecular mechanism of long noncoding RNA SNHG15 in stroke-associated immunodepression.MethodsMagnetic beads separation,q RT-PCR and immunofluorescence were used to analyze SNHG15 expression in PBMCs of ischemic stroke.The binding capacity of SNHG15 and p-STAT6 was assessed by RNA pull-down and co-immunoprecipitation assays.Subsequently,the impact of SNHG15 on STAT6 phosphorylation was studied by western blot.The influence of SNHG15 on cytokines was estimated by ELISA and q RT-PCR.The binding capacity of p-STAT6 and SNHG15 promoter was explored by q RT-PCR,luciferase assay and CHIP.Results1.SNHG15 was highly expressed in monocyte-macrophages of ischemic stroke(P<0.001).2.SNHG15 was interacted with p-STAT6 and promoted STAT6 phosphorylation.3.SNHG15 overexpression partially inhibited M1 polarization of LPS-induced macrophage.SNHG15 deleption partially suppressed M2 polarization of IL-4-induced macrophage.4.The p-STAT6 was interacted with SNHG15 promoter region(~842-~856)after nuclear translocation of p-STAT6,which facilitated SNHG15 systhesis.Conclusions SNHG15 was involved in the macropahge polarization in monocyte-macrophages by promoting p-STAT6 synthesis.The p-STAT6 was interacted with SNHG15 promoter region after nuclear translocation of p-STAT6,which facilitated SNHG15 systhesis.It will further promote M2 polarization of macrophages.