| BackgroundPemphigus vulgaris is a chronic and potentially life-threatening autoimmune blistering disease,the most severe and common subtypes of pemphigus,characterized primarily by the presence of antibodies against desmosomal cadherins,desmoglein(Dsg)3,which lead to a loss of intercellular adhesion,resulting in intra-epidermal blister formation of skin and mucous membrane,its occurrence and development are closely related to the abnormal subsets of Th2 and Treg cells in peripheral blood.In pemphigus vulgaris,up-regulated autoreactive Th2 cells produce large amounts of interleukin(IL)-4,and then stimulate autoreactive B cells to produce excess IgG4,which in turn attacks the extracellular domain of Dsg3 and induces intercellular signals conduction,causing apoptosis and spinal cell lysis.T-regulatory(Treg)cells are a type of cell that play suppressive roles in autoimmune reactivity in vivo.They maintain selftolerance and immune homeostasis by inhibiting the activation,proliferation,and effector functions of various immune cells.Aberrant mTOR pathway activity is involved in a great many autoimmune diseases.Aberrant activation of mTOR pathway causes the activation of effector T and B cells,increased the production of IL-4,and depletion of CD4+CD25+FoxP3+ Treg cells in SLE.In patients with systemic sclerosis,mTOR activation contributes to type I collagen production by dermal fibroblasts.ObjectiveThe purpose of this study was to investigate the correlation of mTOR pathway activity with the loss of balance in Th2/Treg cells in the peripheral blood CD4+T cells of patients with pemphigus vulgaris,and to explore the role of mTOR in pemphigus vulgaris pathogenesis.MethodsPeripheral blood was collected from 15 patients with pemphigus vulgaris and 15 age-and gender-matched healthy controls,CD4+T cells were isolated,and the ratios of Th2/CD4+T cells and Treg/CD4+T cells were analysed by Flow cytometric,the activity of the mTOR pathway,and the expressions of Th2 and Treg cell transcription factors and cytokines were detected by RT-PCR,Western blot,Immunofluorescence staining and ELISA.Primary CD4+T cells from pemphigus vulgaris patients were cultured under Th2-or Treg-polarizing conditions with or without rapamycin in vitro.The Th2 and Treg cell percentages in CD4+T cells were measured by flow cytometry,Jecreted IL-4 and TGF-β levels in the culture medium supernatants were analyzed by ELISA.ResultsWe found that patients with pemphigus vulgaris when compared with healthy controls showed significantly elevated mRNA levels of PI3K,AKT,and mTOR;elevated protein levels of PI3K(P85),AKT,p-AKT(Ser473),mTOR,p-mTOR(Ser2448),p-p70S6K(Thr389),and GATA3;and reduced protein levels of FOXP3.Elevated serum IL-4 was positively correlated with anti-Dsg1/3 antibody titers and disease severity,and the serum TGF-β level was negatively correlated with anti-Dsg3 antibody titer and disease severity,increased Th2/CD4+T cell ratio,and decreased Treg/CD4+T cell ratio.After cultured CD4+T cells which selected from the peripheral blood of PV patients under Th2-or Treg-polarizing conditions in vitro in the presence of rapamycin or vehicle control DMSO for 4 days,we found that the percentage of Th2 cells differentiation and IL-4 secreted in the culture medium supernatants were significantly lower in RAPA group than the vehicle control DMSO group,whereas the percentage of Treg cells differentiation and TGF-β secreted in the culture medium supernatants were significantly higher in RAPA group than the vehicle control DMSO group.These results suggested that inhibiting mTOR pathway activation reversed the loss of Th2/Treg balance in PV patients.ConclusionWe provided evidence that activation of the mTOR pathway and the loss of balance in Th2/Treg cells(i.e.an increase in Th2 and decrease in Treg in peripheral blood CD4+T cells)occurs in PV patients.Inhibiting mTOR activation restored the Th2/Treg balance.This suggests a tight association between the mTOR pathway and Th2/Treg balance,which may present a new target for the treatment of PV. |
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