The Study Of The Pathogenesis Of Abnormal Expression Of WT1Gene In The Paroxysmal Nocturnal Hemoglobinuria

Objective:To investigate the expression of WT1gene in the bone marrow cells of the patients with paroxysmal nocturnal hemoglobinuria (PNH). And to explore the role of abnormal expression of WT1gene in the pathogenesis of PNH.Methods:Eleven newly diagnosed PNH cases (including classic eight cases and three cases of AA-PNH patients) and eight normal controls were enrolled in this study. The expressions of WT1mRNA in CD59-and CD59+BMMNC were measured with semi-quantitive RT-PCR. WT1gene were knocked down with RNA interference (RNAi) technology and then evaluate the change of biological characteristics with Flow-cytometry before and after WTlgene knocked, and investigate its role in the pathogenesis of PNH.Results:1. The relative mRNA expression of WT1in59-BMMNC,59+BMMNC of PNH and BMMNC of normal controls were1.06±0.12,0.90±0.12and0.86±0.05, the expression of it in CD59-BMMNC was significantly higher than that in PNH CD59+BMMNC or normal controls BMMNC (P<0.05),and there was no significant difference between PNH CD59+BMMNC in PNH or normal control BMMNC (P>0.05). The relative mRNA expressions of WT1in BMMNC of PNH cases were positively correlated with the proportion of CD59cells number (coefficient correlation was0.490, P＜0.05), but had no relationship with the proportion of CD59+cell number.2. After the target gene WT1was transfected by siRNA on CD59BMMNC of the patients with PNH, WT1was low and its mRNA expression was decreased.The cells of G0/G1phase in PNH CD59-cell blank control group and siRNA-scr transfection group is (92.73±3.71)%and (93.06±4.14)%, the cells in S phase of them is (6.99±3.61)%and (6.73±4.08)%, respectively. The cells in G0/G1phase of SiRNA-WT1transfected group is (94.46±3.71)%, which in S phase decreased (5.40±3.55)%.There existed significant difference in the cell proportion of G0/G1phase and S phase among the control group, SiRNA-WTl transfected group and the siRNA-scr-transfected group (P<0.05). The cell apoptosis rate in SiRNA-WT1transfected group was significantly increased than that in control group and siRNA-scr transfected group [(35.91±22.36)%vs (26.12±17.10)%vs (27.39±18.99)%](P＜0.05).Conclusion:SiRNA-WT1can effectively suppress the expression of WT1gene of the CD59-clone in the patients with PNH and can also reduce its proliferative capacity,promote its apoptosis. WT1gene expression may participate in the process of PNH clone proliferation.