| Background and objectiveIdiopathic thrombocytopenic purpura(ITP) is the most common hemorrhagic auto-immune disease in children due to excessive platelet destroyed. The pathogenesis of ITP is not completely clarified, we divide it into AITP and CITP clinically by its course more or less than 6 months. More and more research confirmed that part of immune cells and cytokine is related to the process of ITP. There is a exact, strict regulatory system in the normal organism regulatory immune reaction; if the regulatory system is disordered, it will make the occurance, persistence, strength of the normal immune out of control, and cause the autoimmune disease(AID) finally. Recently, in the model auto-antibody produced induced, we found that pessively importance of CD4~+CD25~+Treg can reduce or restrains the auto-antibody production. And later research indicated that CD4~+CD25~+Treg can play an important role by producing cytokine as IL-10, TGF-β; and by the restraining model cell-cell dependent, through CTLA-4. Finally, it can prevents the AID production by two ways: restraining the Th cell activation, differentiation; and restraining B cell directly producing antibody. NK cell is a kind of cell without T or B cell characteristic surface symbol, and it is a kind of important immune regulatory cell, its surface symbol is CD16, CD56 and so on. It play an immune regulatory role by secreting cytokine like IFN-γ, TNF-β, and GM-CSF and so on. Researches indicated that the NK cell activation reduced in peripheral blood of children with ITP, and it is minus related to CD4~+/CD8~+; and is closely related to the serious extent of the condition. The objective is to explore the pathogenesis further, by researching the roles of CD4~+CD25~+Treg cell and NK (CD16~+,CD56~+,CD16~+CD56~+) cell in children with idiopathic thrombocytopenic purpura, and to approach pathogenesy of ITP, to provide theory base to treat ITP.Materials and methods1. experimental object: 24 children cases with AITP and 21 children cases with CITP, each one was drawn 2 ml peripheral blood before treatment, anticagulated. The controls were 15 children cases eliminated hemotopathy diseases. 2. Index detection:①Staining: 2 ml fresh peripheral blood anticagulated by EDTA of children with ITP and controls, each test tube is appended with 10μl antibody, then appended with 50 Ml anicagulated complete blood, mixed, away from light 20～30 minutes.②Hemolysis: Each test tube is appended with 1μl hemolysin, mixed, ecaded from light 15 minutes, then centrifuged for 5 minutes, by 1000～1500 r/min, deserting the upper liquid.③Ablution: Putting PBS liquid 1 ml in above-mentioned residue liquid, mixed, centrifuged(ditto), deserting the upper liquid.④Awaiting to be detected: Putting PBS liquid 0.5 ml in above-mentioned residue liquid, awaiting to be detected. We choose 6000 cells from each semple, and detecting the expression of CD4~+,CD25~+,CD16~+,CD56~+ by flow cytometryCELLQUGST software. 3. Statistical assay, the data was dealt by the SPSS 11.0 statistical software, experimental data was indicated by mean±standard deviation( x±s, %),α=0.05 is the size of test. And the analysis of variance and linear correlation was used.Result1. CD4~+CD25~+Treg cells(%) in peripheral blood: The CD4~+CD25~+Treg cells(%) in children with AITP was 2.11±0.53, in CITP was 1.18±0.47, and in controls was 7.40+0.26. The level of CD4~+CD25~+Treg cells(%) in AITP and in CITP was lower than in controls. There was significance between the ITP and controls(p=0.000), and there was significance between the AITP and CITP(p=0.008), the level in AITP was higher than in CITP. 2. CD4~+T cells(%) in peripheral blood: The CD4~+T cells(%) in children with AITP was 22.95±7.34, in CITP is 21.53±5.59, and in controls was 36.84±6.49. The level of CD4~+T cells(%) in AITP was lower than in controls(p=0.004), the level of CD4~+T cells(%) in CITP was lower than in controls(p=0.003), but there was no significance between the AITP and CITP (p=0.832). 3. NK (CD16~+,CD56~+, CD16~+CD56~+) cells(%) in peripheral blood: The NK (CD16~+,CD56~+,CD16~+CD56~+) cells(%) in children with AITP was 8.55±1.68, in CITP is 7.30±1.47, and in controls was 14.22±2.41. The level of NK (CD16~+,CD56~+,CD16~+CD56~+) cells(%) in AITP was lower than in controls(p=0.004), the level of NK (CD16~+,CD56~+,CD16~+CD56~+) cells(%) in CITP was lower than in controls(p=0.003), but there was no significance between the AITP and CITP(p=0.803). 4. Correlation analysis between CD4~+CD25~+Treg cells(%) and NK (CD16~+,CD56~+,CD16~+CD56~+) cells(%): There was positive correlation in AITP group between CD4~+CD25~+Treg cells(%) and NK (CD16~+,CD56~+,CD16~+CD56~+) cells(%) (r=0.43); and there was no correlation in CITP group; and there was positive correlation in AITP group between CD4~+CD25~+Treg cells(%) and NK (CD16~+,CD56~+,CD16~+CD56~+) cells(%)(r=0.26), the correlation was little. 5. Correlation analysis between CD4~+CD25~+Treg cells(%), NK (CD16~+,CD56~+,CD16~+CD56~+) cells(%) and platelets: The correlation in AITP group between CD4~+CD25~+Treg cells(%) and platelets was the biggest(r=0.37).The correlation in AITP group between NK (CD16~+,CD56~+,CD16~+CD56~+) cells(%) and platelets, and above-mentioned two index in controls and platelets was less. There was no correlation in CITP group.Conclusion1. CD4~+CD25~+Treg and NK degraded, the function to inhibit Th and B cells decreased, which induced the Th and B cells ana-activated, cell-mediated immunity and humoral-mediated immunity immunologic derangement, ultimumly induced occurrence of ITP. 2. Compared to AITP, CD4~+CD25~+Treg degraded manifestly in CITP, it cued that the role of cell-mediated immunity predominant.
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