Study Of Complement And Regulatory T Cells Of The Patients With Immuno-related Pancytopenia

Objective:To investigate the bone marrow complement levels,the quantity and function of regulatory T cells in the IRP,then expore the mechanism of hematopoietic cells destruction and significance of regulatory T cells in the pathogenesis of IRP.Methods:Fifty-nine untreated IRP patients,65 recovered IRP patients and 24 healthy donors as controls were enrolled in this study.The bone marrow levels of CH50,C3, C4,C5b-9,IL-2,TGF-βwere tested with ELISA.The auto-antibodies on bone marrow hematopoietic cells(BMHC)and the quantities of regulatory T cells(CD4~+CD25~+/CD4~+ cells,CD4~+CD25~+CD127~-/CD4~+ cells)were examined with flow cytometric assay.The expressions of FoxP3 and Galectin-10 mRNA in BMMNC were measured with semiquantitive RT-PCR.Results:1.The level of C5b-9 in bone marrow of untreated IRP patients(119.82±53.95)was significiantly higher than that of recovered IRP patients(100.74±33.42)or normal controls(93.86±28.81)(p＜0.05),and there was no significant difference between recovered IRP patients and normal controls(p＞0.05);The levels of CH50 in bone marrow of untreated or recovered IRP patients(33.29±11.51,30.77±10.34)was significiantly higher than that of normal controls(24.09±6.37)(p＜0.05),and there was no statistic significance between untreated IRP patients and recovered IRP patients (p＞0.05);The level of C3 in bone marrow of untreated or recovered IRP patients (4.90±2.19,4.95±3.47)was significiantly lower than that of normal controls (6.98±5.56)(p＜0.05),and there was no statistic significance between untreated IRP patients and recovered IRP patients(p＞0.05);The bone marrow level of C4 among three groups was no statistic significance(p＞0.05);The level of complement in peripheral blood was consistent with in bone marrow.Bone marrow CH50 level of IRP patients was negatively correlated with C3 level(r=-0.303,p=0.007)and positively correlated with C5b-9 level(r=0.241,p=0.003).IgG autoantibody accounted for 55.17%,IgM 54.31%,IgG+IgM 31.90%in all patients,BMHC-IgM positive group accounted for 86.21%in all patients.The bone marrow level of C5b-9 in IRP patients with BMHC-IgM(117.59±55.69)was higher than that in IRP patients without BMHC-IgM(99.15±26.15)(p＜0.05).The positive rate of CD34~+-IgG or CD34~+-IgM of IRP patients was positively correlated with their C5b-9 levels(r=0.593 p=0.000,r=0.326 p=0.049).The percentage of reticulocyte(r=0.421,p=0.000)and the serum level of indirect bilirubin of IRP patients were positively correlated with their CH50 levels(r=0.230,p=0.032).After treatment,the percentage of reticulocyte reduced to normal(p=0.038)and the quantity of leukocyte increased to normal(p=0.002).The bone marrow level of C3 was higher than that of before treatment(p=0.003);and the positive rate of CD34~+-IgG or CD34~+-IgM was lower than that of before treatment(p=0.016,p=0.022).2.The bone marrow levels of IL-2 in untreated or recovered IRP patients(5.64±1.70, 6.19±2.53)was significiantly lower than that in normal controls(7.91±3.71)(p＜0.05); The bone marrow levels of TGF-βin untreated or recovered IRP patients(1.79±0.67, 1.86±0.77)was significiantly lower than that in normal controls(2.48±0.94)(p＜0.05), and there was no statistic significance between untreated IRP patients and recovered IRP patients(p＞0.05);The quantity of CD4~+CD25~+/CD4~+ cells in untreated IRP patients(22.46±9.47)was significiantly lower than that in normal controls (30.59±8.58)(p＜0.05);The quantity of CD4~+CD25~+CD127~-/CD4~+ cells in untreated IRP patients(7.18±2.72)was significiantly lower than that in normal controls (10.44±3.24)(p＜0.05),and there was no statistic significance between recovered IRP patients and normal controls(p＞0.05).The relative mRNA expressions of FoxP3 and Galectin-10 were 0.34±0.25,0.69±0.51,0.82±0.65 and 0.66±0.11,0.74±0.11, 0.76±0.09 in three groups.The expression of untreated IRP patients was significiantly lower than that of recovered IRP patients or normal controls(p＜0.05).Conclusions:The destruction mechanism of hematopoietic cells of IRP patients especially with BMHC-IgM have some relationship with auto-antibody activating complement.The regulatory T cells play important role in the pathogenesis of IRP.