| Background and Objective: Myelodysplastic syndromes (MDS) are a group of clonal stem/progenitor cells diseases.Currently, a combination of morphology, to detect multilineage dysplasia in the bone marrow and peripheral blood, and bone marrow biopsy, to detect the proliferation of bone marrow and cytogenetics, to detect characteristic clonal abnormalities are the main methods in establishing a diagnosis of MDS. However, the clinical manifestations are various and the various of morphology depend upon the experiences of haematologists, and only approximately 40%-50% patients were detected with abnormal cytogenetics. So, to make the diagnosis of MDS is difficulty sometimes. Recently, flow cytometry (FCM) is widely used to detect the immunophenotype of malignant hematological disease (such as leukemia and lymphoma), which can provide objective adjuvant evidence for diagnosis, classification, therapy and prognosis. MDS is a heterogeneous group of myeloid neoplasms. The aim of this study is to use multiparametric flow cytometry to analysis the immunophenotypic characteristic of MDS patients and establish the flow cytometry scoring system, explore the clinical significance of this scoring system for diagnosis, classification and prognosis.Methods: 41 bone marrow samples were collected from MDS patients and 30 bone marrow samples were obtained from non-MDS patients, applying multiparametric flow cytometry and adopting an extensive panel of monoclonal antibodies and CD45/SSC gating strategy, to detect the expressions of antigens of bone marrow cells and the quantity of blast. Comparing the quantity of blast, the expressions of antigens and the FCM scores between MDS and non-MDS with Independent-Samples T Test and among MDS subtypes with One-Way ANOVA, analyzing the feature of immunophenotyping and establishing FCM scoring system as well, combining the results of hematology, morphology, cytogenetics and other methods to evaluate this FCM scoring system with ROC Curve and Spearman rank order correlation analysis.Results: Comparing with the non-MDS group, the proportion of blast, monocyte, lymphocyte and nucleated erythrocyte were significantly increased in the MDS group (P=0.000, P=0.000, P=0.001, P=0.029, respectively), but percentage of granulocyte was significantly decreased (P=0.021); The proportion of blast was increased grandually in RCMD, RAEB-I and RAEB-II and there were significant differences in RCMD vs RAEB–I , RCMD vs RAEB–II and RAEB–I vs RAEB-II(P=0.020, P=0.000, P=0.010, respectively). Other cell populations were no significant differences(P>0.05).The expressions of CD34, CD123 and CD133 on blast cells were much higher than non-MDS group (P=0.000, P=0.003, P=0.000, respectively), the expressions of CD38, HLA–DR, CD13, CD33, CD7 and CD117 were no significant differences (P=0.223, P=0.306, P=0.536, P=0.273, P=0.577 and P=0.368, respectively). Between RCMD vs RAEB-I, RCMD vs RAEB-II and RAEB-I vs RAEB-II, the expressions of CD34 were significantly increased (P=0.022, P=0.000, and P=0.000, respectively), Meanwhile the expressions of CD133, CD117, CD13 and HLA-DR on blast of RAEB–II were significantly increased than RCMD(P=0.001, P=0.000, P=0.001, P=0.047, respectively). Other expressions of antigens were no significantly differences among the subtypes of MDS(P>0.05).The expressions of CD10, CD11b, CD15, CD16 and CD66d on granulocytes were much higher than non-MDS group(P=0.012, P=0.027, P=0.016, P=0.005, P=0.024, respectively); there were no significantly differences of the expressions of CD33, CD13, HLA-DR, CD56 and CD117(P=0.058, P=0.337, P=0.649, P=0.096, P=0.081, respectively); the expression of CD10 in RAEB-II was decreased significantly than RAEB-I(P=0.001); the expressions of CD13 and HLA-DR in RCMD were significantly decreased than RAEB-I and RAEB-II(P=0.013, P=0.005 and P=0.018, P=0.003, respectively). Other expressions of antigens were no significantly differences among the subtypes of MDS(P>0.05).The expressions of CD15, CD33 and CD64 on monocytes were significantly higher than non-MDS group(P=0.001, P=0.007, P=0.000, respectively), however, there were no significantly differences of the expressions of CD7, HLA-DR, CD14, CD13, CD61 and CD56 on monocytes(P=0.075, P=0.066, P=0.408, P=0.153, P=0.137, P=0.322, respectively). Expressions of all antigens were no significantly differences among the subtypes of MDS(P>0.05).The expressions of CD25+CD8+, CD7 and CD8 on lymphocytes were significantly increased than non-MDS group(P=0.000, P=0.002, P=0.004, respectively), while the expressions of CD5 and CD19 were significantly decreased(P=0.011, P=0.000, respectively); the expressions of CD25+CD4+, CD4, CD25 and CD56 were no significantly differences(P=0.524, P=0.508, P=0.942, P=0.978, respectively); the expressions of CD5 in RCMD and RAEB-II was significantly increased than RAEB-I(P=0.006, P=0.037, respectively). Other expressions of antigens were no significantly differences among the subtypes of MDS(P>0.05).The expressions of GPA+CD71+, CD71, CD105 and GPA on nucleated erythrocytes were significantly increased than non-MDS group(P=0.026, P=0.020, P=0.033, and P=0.024, respectively), while the expressions of GPA+CD105+ and CD71+CD105+ were no significantly(P=0.050, P=0.938, respectively). Expressions of all antigens were no significantly differences among the subtypes of MDS(P>0.05).7 out of 41 MDS patients had complicate chromosome karyotype and all these 7 patients were RAEB-II, account for 17% of MDS patients and 47% of RAEB-II patients. There was good positive correlation between abnormal chromosome karyotype and IPSS scores(r=0.703, P=0.000).Base on the proportion of blast and the expressions of antigens on cell populations, we establish this BM FCM scoring system preliminarily. Using this scoring system for MDS diagnosis had a good coincidence rate(P=0.000), 95% confidence interval was 0.921～0.997,sensitivity and specificity were 87.8% and 86.7%. The BM FCM scores of MDS group was significantly higher than non-MDS group(P=0.000),as disease progression, the score was increased(P=0.000). Meanwhile, there were good positive correlations between FCM scores and MDS classification, karyotype, IPSS scores and WPSS scores respectively(r=0.656, r=0.410, r=0.791 and 0.425, P=0.000, P=0.008, P=0.000 and 0.006 respectively). However, there were no significantly differences between FCM scores of BM cell populations and absolute counts of PB(P>0.05).Conclusions: Using multiparametric flow cytometry to analysis the expressions of antigens of bone marrow cells from MDS patients could help us know much more about the immunophenotypic characteristics of MDS. Through synthetical evaluation the immunophenotypic abnormalities of MDS base on our preliminary study on establishing this BM FCM score system, we may could provide a practical, objective, reliable and adjuvant approach. |
