The Effect Of Methylation Inhibitor Exert On SHP-1, JAK3, TYK2 Genes In Leucocythemia Cells

Objectives: In this essay, we investigated the influence of the expression about methylation inhibitor to JAK3 and TYK2 which are the members of Janus kinase(JAK) family and hematopoietic cell phosphatase (SHP-1) in leukemia cell line, and explored their role in the pathogenesis of leukemia.Methods: In this study, the investigation object were the chronic myelocytic leukemia cell line K562 and leukemia cell line U937.These cells were suspension cultured in RPMI1640 with 10% heat inactivation (56℃,30min) calf serum. These cells in exponential phase of growth were transferred to the cell cultivante board. There were 1×10⁶/ml cells in each hole. The concentration of 5-aza-CdR were 0.5, 1, 2μmol/l (all 3 holes); As₂O₃ were 1, 2.5, 5μmol/l (all 3 holes); As₂O₃ 1μmol/l and 5-aza-CdR 0.5μmol/l,As₂O₃ 2.5μmol/l and 5-aza-CdR 1μmol/l , As₂O₃ 5μmol/l and 5-aza-CdR 2μmol/l (every 1 hole); no drug in 1 hole. There were two holes cells with 10 different treated conditions. These cells with drugs were incubated under 37℃,5%CO2 in 24h,48h,72h.After that, cells were harvested, washed, extracted total RNA, detected the expression of JAK3, TYK2, SHP-1 with the method of Real-time Quantitative Polymerase Chain Reaction(RQ-PCR). Results:1 Expression of SHP-1 mRNA in K562 cell line: When As₂O₃ and 5-aza-CdR single or combine effected, the expression of SHP-1 mRNA was dose and time dependence. The gene expression became higher. It had obviously difference and statistical significance (p<0.05). 2 Expression of JAK3 mRNA in K562 cell line: When As₂O₃ and 5-aza-CdR single or combine effected, the expression of JAK3 mRNA was dose and time dependence. The gene expression was downtrend. It had obviously difference and statistical significance(p<0.05). 3 Expression of TYK2 mRNA in K562 cell line : When As₂O₃ and 5-aza-CdR single or combine effected ,the expression of TYK2 mRNA was dose and time dependence. The gene expression was downtrend. It had obviously difference and statistical significance(p<0.05).4 Expression of SHP-1 mRNA in U937 cell line :When As₂O₃ and 5-aza-CdR single or combine effected , the expression of SHP-1mRNA was dose and time dependence. The gene expression became higher. It had obviously difference and statistical significance (p<0.05).5 Expression of JAK3 mRNA in U937 cell line :When As₂O₃ and 5-aza-CdR single or combine effected , the expression of JAK3 gene was dose and time dependence. The gene expression was downtrend. It had obviously difference and statistical significance (p<0.05).6 Expression of TYK2 mRNA in U937 cell line :When As₂O₃ and 5-aza-CdR single or combine effected , the expression of TYK2 mRNA was dose and time dependence. The gene expression was downtrend. It had obviously difference and statistical significance (p<0.05).7 Dependablities of SHP-1 and JAK3,TYK2: SHP-1 gene educed down-regulation to JAK3 gene , TYK2 gene. The contribution JAK3 gene was suffered was more significanct than TYK2 gene.Conclusions: 1 Both As₂O₃ and 5-aza-CdR can raise up the expressions of SHP-1 mRNA in the cell lines of K562 and U937, and degrade the expressions of JAK3 mRNA and TYK2 mRNA. 2 SHP-1 gene maybe inhibit Janus kinase-signal transducer and activator of transcription signaling pathway. JAK3 gene was suffered was more significanct than TYK2 gene . In Janus kinase-signal transducer and activator of transcription signaling pathway, JAK3 gene maybe educe prior contribution.