| Section1Objective This study was designed to investigate the pathogenesis of Autoimmune Hemolytic Anemia (AIHA).Methods:The following indexes were detected by flow cytometry using blood samples from54patients with AIHA. They were the amount of CD19+and CD5+CD19+B lymphocytic cells, the amount of CD4+and CD8+T lymphocytic cells,the amount of NK cell, the amount of Thl and Th2T helper cells(CD4+cells), the amount of T regular cells and T effective cells (CD25+and HLA-DR+T cells), the amount of mDC and pDC dendritic cells. Patients were divided into hemolytic attack group and remission group. The indexes from each group were statistically analyzed and compared. Measurement data were described as mean±standard deviation. Applying SPSS13.0statistics software, we compared two normal distribution data based on t-test, abnormal distribution data based on non-parametric test (Kruskal-Wallis test), we analyzed three groups by analysis of variance (one-way ANOVA). Statistical significance was defined as P<0.05.Results1. B cells:The amount of B lymphocytic cells (CD19+cells) from hemolytic attack group was significantly higher than that from normal control (P=0.000,<0.01). CD5+CD19+B cells from hemolytic attack group were in excess of those from normal control (P=0.033, P<0.05). CD19+and CD5+CD19+B cells from remission group were significantly lower than from hemolytic attack group (P=0.000, P=0.000, P<0.01). The ratio of CD5+CD19+/CD19+from remission group was lower than from hemolytic attack group(P=0.004, P<0.01). The amount of CD19+cells,CD5+CD19+cells and the ratio of CD5+CD19+/CD19+were lower than from health control group (P=0.008, P=0.000, P=0.028; P<0.01, P<0.01, P<0.05).We also divided the patients based on treatment whether using rituximab. The amount of CD19+cells from rituximab treatment group was lower than the counterpart group (P=0.017, P<0.05). There was no statistically difference of CD5+CD19+cell quantity between rituximab group and the counterpart group. The ratio of CD5+CD19+/CD19+from rituximab group was higher than the counterpart group (P=0.032, P<0.05)2. T cells:There were no significant differences of the amount of CD4+cells, CD8+cells,NK cells, CD25+cells and HLA-DR+T cells between patients and healthy controls. The ratio of CD4/CD8was not significantly different between the two groups either. The amount of Thl cells from hemolytic attack group (P=0.002, P<0.05)and remission group(P=0.021, P<0.05)was both higher than normal control. There was no significantly differences of the amount of Thl cells between hemolytic attack group and remission group (P>0.05). The percentage of Th2cells from hemolytic attack group was higher than control (P=0.000, P<0.01). Meanwhile, the percentage of Th2cells from remission group was lower than hemolytic attack group (P=0.002, P<0.01). This indicated that after immunosuppressive therapy, Th2cells tended to become normal while the disease tended into remission.3. Dendritric cells:The amount of CD123+cells from patients was lower than control (P=0.000, P<0.01), the percentage of CD11c+cells from hemolytic attack group was lower than remission group (P=0.011, P<0.05). There was no statistically difference of CDllc+cells between remission group and control group. The percentages of CD123+cells from both hemolytic attack group and remission group were lower than control (P=0.000, P=0.001). There was no statistically significance of the amount of CD123+cells between hemolytic attack group and remission group. The ratio of CD11c+/CD123+from both hemolytic attack group (P=0.022, P<0.05) and remission group (P=0.002, P<0.01) were higher than control. There was no significantly differences of the ratio between hemolytic attack group and remission group.Conclusions The pathogenesis of AIHA is associated with activation of B lymphocytic cells(CD5+CD19+). During the hemolytic attack, the percentages of CD19+B cells and CD5+CD19+B cells substantially elevate, indicating the proliferation of B lymphocytic cells. The amount of CD19+lymphocytic cells from rituximab group was lower than the counterpart group, while there was no statistically significance of CD5+CD19+cells between rituximab group and the counterpart group. this suggests that rituximab exhibits a strong inhibition effect towards immune system. The regulation of T lymphocytic cells (especially Th2proliferation) is also related to the mechanism of AIHA. The percentages of Thl and Th2cells both increased during hemolytic attack, and Th2decreased when remission was achieved while Thl showed no alteration. The pathogenesis of AIHA is related to DC cells. The percentages of DC1and DC2cells both decreased during the attack, yet DC1tended to become normal while DC2decreased after immunosuppressive therapy. This indicates that DCs maybe migrate into peripheral lymphoid tissue when hemolysis attacks.Section2. Objective To better assess the efficacy and safety of monoclonal anti-CD20antibody rituximab combined cyclophosphamide with in treatment of refractory and recurrent autoimmune hemolytic anemia. Cases The study included7cases of autoimmune hemolytic anemia (including1case of Evans syndrome), which all of them were refractory or recurrent. Methods rituximab:375mg/m2, once per week,2-6times; Cyclophosphamide:1g/10days,2-7times; combined with intravenous immunoglobulin (IVIG)5g/week, given1day after rituximab treatment. Results All7patients showed good responses.6patients achieved complete remission (CR) and1patient achieved partial remission (PR). Responses occurred1to10months after the first dose of rituximab and the mean effective time is approximately2.5months. Average follow-up for the patients is around27months. All patients remained in remission at the12-month follow-up visits. At the time of24-month visits,2patients showed elevated indirect bilirubin and increased reticulocyte counts. One patient achieved CR after additional rituximab therapy, and the other patient just wait and watch without additional therapy. At the time of36-month visits,1patient relapsed and was retreated with3cycles of rituximab and eventually reached PR. All patients tolerated the treatment well with only mild side effects. Conclusions rituximab combined with and recurrent autoim cyclophosphamide is highly effective and relatively safe in patients with refractory mune hemolytic anemia. Additional treatment can be given in patients with relapse after1-2years.Section3. Objective To study the clinical and laboratory features of the patients with immuno-related pancytopenia(IRP). Methods The risk factors,manifestations, blood cell counts,bone marrow phenotypes,autoantibodies and immunosuppressive therapy response of157patients with IRP were analyzed.Then they were followed up for (3～48) monthS,to see their long-term outcome and the prognostic factors. Results The infection,anaphylaxis and pregnancy were highly suspected to be the risk factors of IRP.Most of these patients were with pancytopenia73.2%(115/157), some patients were with anemia and thrombocytopenial8.5%(29/157)];61.3%(95/155) of them were anemic with large MCV,32.9%(51/155) of them were anemic with normal MCV;78.9%(124/157) of them were with leukopenia,23.6%(37/157) of them had fever. Thrombocytopenia was common[91.7%(144/157)],but serious bleeding was rare.68.8%(108/157)of these cases were with nomal or increased bone marrow cellularities and increased normoblasts.They were all found to have positive results of bone marrow mononuclear cell antibody Facs test,negative results of rutine hemolysis tests and no evidence of malignant clonal hematopoiesis.42.6%(67/157) of these patients had C3decreased, and18.5%(29/157) C4decreased. Immunosuppressive therapy was administered to157IRP patients,The response rate at36months was87.5%(28/32). Conclusions Immuno-related pancytopenia(IRP) is an acquired multiclonal autoimmunoantibody conducted bone marrow failure syndrome, featuring pancytopenia without reducing of reticulocytes and neutrophilic granulocytes. More than half of the patients used to have infections or allergies. A few patients’bone marrow have higher proliferation at least one site, with mild hemolysis yet negative hemolytic test. The following abnormalities often complicates with IRP, such as low amount of complements, positive ANA. Those patients may receive good responses to immunosuppressants and hemopoiesis improving treatment.Conclusions The pathogenesis of AIHA is closely associated with B lymphocytic cells and CD5+B lymphocytic cells. Th2cells, the regulating cells of B lymphocytic cells, may play an important part in hemolysis. The amount of mDC and pDC decrease during hemolytic attack, which caused by DC peripheral migration. Refractory AIHA has good response to Rituximab,but sometimes relapse1year later. Immuno-related pancytopenia is an acquired multiclonal autoimmunoantibody conducted bone marrow failure syndrome, featuring pancytopenia without reducing of reticulocytes and neutrophilic granulocytes. More than half of the patients used to have infections or allergies. A few patients’bone marrow have higher proliferation at least one site, with mild hemolysis yet negative hemolytic test. The following abnormalities often complicates with IRP, such as low amount of complements, positive ANA. Those patients may receive good responses to immunosuppressants and hemopoiesis improving treatment. |
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