Impaired Immunosuppressive Effect Of Bone Marrow Mesenchymal Stem Cell-derived Exosomes On T Cells In Aplastic Anemia

Background:Previous studies have verified the dysfunction of mesenchymal stem cells(MSCs)for immunoregulation in acquired aplastic anemia(AA)patients.Exosomes derived from MSCs can partially substitute MSC acting as immune regulator.Dysfunction of exosomes(Exos)derived from AA-MSC(AA-Exos)may play a key role in immunologic dissonance.Objectives:This study aims to investigate the interaction between exosomes derived from mesenchymal stem cells and T cells of patients with aplastic anemia by examining the immunosuppressive effects of exosomes on T cells both in vitro and in vivo.Besides,this study intends to explore the differently expressed miRNA of exosomes and mRNAs of T cells between patients with AA and healthy donors(HD)to better understand the mechanisms of AA on molecular lever.Method:In this study,CD3+T cells were collected and cocultured with AA-Exos and exosomes derived from HD-MSC(HD-Exos).The proliferation,differentiation and activation of CD3+T cells were detected to compare the immunosuppressive effects between AA-Exos and HD-Exos.An immune-mediated murine model of AA was structured to compare the therapeutic effect of AA-Exos and HD-Exos.Furthermore,total RNA including miRNA from exosomes we purified and total RNA of CD3+T cells were extracted for RNA-seq in order to construct the miRNA-mRNA network for interactions and functional analysis.Results:AA-Exos had impaired inhibition effects on CD3+T cells in terms of cell proliferation,activation and differentiation compared with HD-Exos.HD-Exos other than AA-Exos can rescued the AA mice.Importantly,we identified some differentially expressed miRNA involved in immune response,such as miR-199,miR-128,miR-486 and miR-375.The Gene Ontology analysis of differentially expressed genes(DEGs)revealed involvement of various cellular processes,such as lymphocyte chemotaxis,lymphocyte migration and response to interferon-gamma.The Kyoto Encyclopedia of Genes and Genomes analysis illustrated upregulation of critical pathways associated with T cell function after co-culturing with AA-Exos compared with HD-Exos,such as graft-versus-host disease,Th 17 cell differentiation,and JAK-STAT signaling pathway.A miRNA-mRNA network was established to visualize the interaction between them.Conclusion:In summary,AA-Exos had impaired immunosuppressive effect on T cells,less ability to rescue AA mice and differently expressed miRNA profile,which might partly account for the pathogenesis of AA as well as provide a new target of AA treatment.Background:Platelet transfusion is important for preventing and treating bleeding in patients with aplastic anemia(AA),but repeated platelet transfusions may lead to platelet transfusion refractory in some patients.Objective:To analysis the clinical features of severe aplastic anemia(SAA)patients with platelet transfusion refractory(PTR-SAA)and evaluate the hematological response and survival prognosis of these patients following immunosuppressive therapy(IST).Methods:From January 2011 to December 2020,a cohort of 281 SAA patients in our center following antithymocyte/lymphocyte globulin(ATG)-based IST were enrolled in this study.We collected their clinical data and retrospectively analyzed their hematological response rates and long-term survival.Results:Twenty patients developed PTR.Very severe aplastic anemia accounted for 70%of PTR-SAA patients,which was significantly higher than that in patients without PTR(46.4%,p=0.04).Additionally,PNH clones presented in up to 40%PTR-SAA patients,which was remarkedly higher than that in patients without PTR.At the onset of disease,the platelet counts(PLT)and absolute neutrophil counts(ANC)were much lower in PTR-SAA than those in patients without PTR(p<0.001 and p=0.04,respectively).The median 24h corrected count increment(CCI)was 594×109/L.Eighteen patients safely accomplished the ATG course.What’s more,the status of PTR was corrected in all patients after ATG.The response rates at 3 months and 6 months after ATG were 27.8%and 44.4%in PTR-SAA patients,which were lower than those in patients without PTR(49.2%and 60.7%,respectively)but without significant difference.Older(>40 ages)and VSAA patients had inferior response rates at 3 and 6 months than those younger and SAA patients(p=0.02 and p<0.001).The 10-year overall survival(OS)was 60.9%which was significantly lower than that in patients without PTR(80.0%,p=0.03).Multivariate analysis revealed that SAA but not VSAA and short interval from diagnosis to IST(<50 days)were positive prognostic factor for OS(RR=2.7,95%CI 1.4-4.9,p=0.002.RR=1.9,95%CI 1.13.3,p=0.02).Conclusions:PTR-SAA was a special subtype of SAA with a relatively poor prognosis.Adequate therapy targeting PTR could bridge SAA patients to accomplish ATG safely.ATG could eliminate the PTR.What’s more,up to half of PTR-SAA patients may be cured with normal peripheral blood counts without relapse and clonal evolution.