Expression And Function Of IL-3Rβ In Apl Cells

The complex network of haematopoietic growth factors (HGFs) regulate a precise and important balance of normal hematopoiesis between proliferation, differentiation,and self-renewal.The human IL-3 receptor is composed of bothαsubunits and a commonβsubunit,which shares with IL-5 and GM-CSF receptors.To date, rare experiential data exist about the role of IL-3Rβgene expression in the differentiation of AML cells.The subjects of the thesis were focused on the following aspects:1.Study the expression of IL-3R in APL patients using real-time quantitative RT-PCR, comparing with healthy donors.2.Study any potential effect on proliferation, differentiation on IL-3Rβknockdown NB4 cells induced with ATRA .The main results are as following:1.IL-3RβmRNA expression levels in APL patients were significantly lower than in normal donors and complete remission patients(P<0.05).And the IL-3RβmRNA expression was increased significantly after ATRA treatment compared to the untreated NB4 cells(P<0.05).2.The shRNA expression plasmids targeting on IL-3RβmRNA were successfully constructed.Stable clones were obtained after gene transfection and G418 screening,and IL-3RβshRNA plasmid showed obviously efficacy in decreasing IL-3Rβexpression with the detection of Real-time RT-PCR and Western Blot.3.Knockdown of IL-3Rβexpression in NB4 cells resulted in significant inhibition of cell proliferation as compared to the control cells.Cell cycle analysis demonstrated knockdown of IL-3Rβexpression in NB4 cells lead to a reduced proportion of S-phase cells, and the increased G1,G2-phase cells.For NB4-IL-3RβshRNA cells induced with ATRA,an accumulation of cells in the G2-phase was also detected,as compared to the control group cells.But no significant difference on cell differentiation was observed in NBT assays and cells surface marker CD11b for both cell lines induced with or without ATRA.In addition,with the down-regulation of IL-3Rβexpression,the level of IL-3Rαand PML-RARαwere decreased,compared to the control group cells.In conclusion,abnormalities of IL-3Rβexpression were detected in APL patients. Knockdown of IL-3Rβrendered NB4 cells treated with ATRA proliferation inhibited rather than differentiation.Moreover,down-regulation of IL-3Rβcaused the decrease of IL-3Rαand PML/RARαexpression.The up-regulation of IL-3Rβexpression may be the self-adjustment of NB4 cells in the stage of cells differentiation for survival.