The Expression Of PD-land Its Ligands In Immune Thrombocytopenia

[Background and Objective]Immune thrombocytopenia (ITP) is a common autoimmune disease in which autoreactive T and B cells are activated by platelet autoantigens resulting in immune-mediated platelets destruction and/or suppression of platelets production. In recent years, more and more evidences indicate that, ITP is a heterogeneous disease. The pathogenesis of ITP include both humoral immune disorders and cellular immunity disorders, wherein T lymphocytes play an important role. T cell activation requires histocompatibility antigens (MHC)-peptide complex stimuli (the first signal), but also need the co-stimulatory signal (known as a second signal). Lacking of the latter can lead to T cell responses incompetent, or even programmed cell death. PD-1 (programmed death-1) and its ligand PD-L1 (programmed death ligand-1) is an important negative costimulatory molecules found in recent years, and can mediate negative costimulatory signal. Which is thought to be closely related with transplantation rejection, autoimmune diseases, lentivirus infection, tumor immune escape and other diseases.PD-1 which providing an inhibitory signal to T cell activationis is an immune inhibitory receptors and their structure and function is similar to CILA-4 (cytotoxic T lymphocyte-associated antigen 4). PD-1 plays a critical role in establish and/or maintain peripheral tolerance through restraining T cell immunization:1?Inhibit transformation from autoreactive naive T cells to effector T cells(Teff).2?Induce conversion from Teff to Treg.3 ?Activate Treg/DCs to inhibit Teff.PD-1 has two ligands, PD-L1 and PD-L2. PD-1 combination with PD-L1/PD-L2 can obvious block TCR mediated proliferation, cytokine secretion and stay cells in G0/G1 phase. PD-1/PD-L is another negative costimulation after CTLA-4. Whose functions are impair, limit and/or stop the activation of T cell, B cell and marrow cell. Recent clinical trials have shown that aberrant expression of PD-1 signal pathway can result in breakout of the peripheral tolerance and autoimmune diseases. As a tissue-specific autoimmune diseases, if ITP relate to this signal pathway, there have currently no reports at home and abroad. In our research, we detected the expressions of PD-1, PD-L1 and PDL-2 on T cell and Dendritic cells(DCs), we also measured the concentrations of sPD-1 in plasma, the expressions of sPD-1 mRNA. Ultimately we analyze the different forms of PD-1 in the pathogenesis of ITP.[Methods]Thirty patients with newly diagnosed and recurrent ITP patients (twenty two females and eight males, age range 14-76 years, median 39.1 years, platelet counts that range from 0 to 27x109/l, with median counts of 8.7×109/l) were enrolled for flow cytometry and enzyme-linked immunosorbent (ELISA) assay. Peripheral blood sample of another 16 newly diagnosed patients with ITP were collected (ten females and eight males, age range 16-72 years, median 39.5 years years, platelet counts that range from 2 to 19×109/l, with median counts of 10×109/l) for the real-time RT-PCR assay. They had not been treated with any immunosuppressive agents for at least one week before sampling. The healthy controls consist of 36 adult healthy volunteers (tween females and sixteen males, age range 19-72 years, median 38 years; platelet counts range from 129 to 241× 109/l, with a median count of 184×109/l). Enrollment took place between November 2014 and April 2015 at the Department of Hematology, Qilu Hospital, Shandong University. All of the cases were selected and diagnosed by our study group, and all met the diagnosis criteria of primary ITP as previously described. Informed consent was obtained from each patient and healthy control in accordance with the Declaration of Helsinki. Ethical approval for the study was obtained from the Medical Ethical Committee of Qilu Hospital, Shandong University. The expressions of PD-1, PD-Lland PDL-2 on T cell and Dendritic cells(DCs)were detected by flow cytometry. The concentrations of sPD-1 in plasma were measured by enzyme-linked immunosorbent assay ELISA. The expressions of sPD-1 mRNA were measured by real-time RT-PCR assay.[Results]1. Expressions of PD-1 on T cellsThe percentage of PD-1 positive cells was significantly increased in CD4+cells (P<0.01) and CD8+cells (P<0.01) in patients with ITP when compared with healthy controls.2. Expressions of PD-L1, PD-L2 on DCsCompared with those in healthy controls, the MFI of PD-L1 on DCs was significantly reduced (p<0.05). Similarly, the MFI of PD-L2 on DCs of the ITP patients was significantly lower than that of healthy controls (p<0.01).3. Concentration of plasma sPD-1The level of plasma sPD-1 in ITP patients was significantly higher than that in healthy controls (P<0.01).4.The level of sPD-1 mRNACompared with healthy control, ITP patients show significantly increased in the level of sPD-1 mRNA(P<0.01).[Conclusions]These results suggest that the aberrant expressions of PD-1, PD-L1 and PD-L2 may play a role in ITP. The decreased expressions of PD-L1 and PD-L2 on DCs, the increased level of sPD-1 mRNA, increased plasma concentration of sPD-1 might link to a pathogenic mechanism involved in reduced ligation of PD-1 and PD-LI, PD-L2 between T cells and antigen presenting cells thus leading to activation of autoreactive T cells in ITP.