Effect Of Decitabine On The Proliferation Of T Cell Lymphoma Through BIM Gene And Its Related Mechanism

Objective To explore the methylation pattern of BIM gene in T cell lymphoma,and the effect of Decitabine(DAC)on cell proliferation and BIM gene expression in PTCL cells.Methods1.Target genes and cell lines were screened by bioinformatics metho ds.Data used for bioinformatics analysis were obtained by the Gene Expr ession Omnibus(GEO)platform;cytohubba calculations are performed in Cytoscape software to screen for target genes.The Human Protein Atlas(HPA)database was used to screen cell lines according to mRNA levels.2.Bone marrow biopsy samples from clinical patients were selected,including 16 cases of peripheral T-cell lymphoma(PTCL)and 10 cases of normal control.Patients with malignant lymph node reactive hyperplasia were excluded from the normal group.To explore whether BIM gene methylation is related to certain clinical characteristics,including gender,age,stage,type,etc.3.Cell proliferation was detected by CCK-8 method.Changes in absorbance values of Hut-78 and Jurkat cells treated with(0,2.5,5,10,25,50,100,200)μM decitabine for 24,48,72 h were detected by CCK-8 method,so as to screen the optimal treatment concentration and time.4.Hut-78 and Jurkat cells were divided into control group and decitabine treatment group,and positive control and negative control were set.The effect of decitabine treatment on the methylation level of BIM gene in cells was detected by MSP method.5.Hut-78 and Jurkat cells were divided into control group and decitabine treatment group,respectively.Western Blot was used to detect the effect of decitabine treatment on BIM gene protein levels in cells.6.Hut-78 and Jurkat cells were divided into control group and decitabine treatment group,respectively.qRT-PCR was used to detect the effect of decitabine treatment on BIM mRNA level in cells.Results1.A potential new molecular marker BIM for peripheral T-cell lymphoma was identified by screening data from public data sets.Hut-78 and Jurkat cells were screened from HPA database according to BIM mRNA levels.2.BIM gene showed a higher methylation frequency in PTCL patients,which was significantly different from that in the normal control group,and the results were statistically significant.There was no significant correlation between BIM gene methylation pattern and clinical characteristics of PTCL patients collected,which may be related to the small number of samples.3.The results of cell proliferation activity measured by CCK-8 showed that decitabine inhibited the proliferation of Hut-78 and Jurkat cells in a concentration and time dependent manner.The above cells were treated with decitabine for 48 h and reached half inhibitory concentration at 200μmol/L.4.Methylation-specific PCR results showed that BIM showed hypermet hylation in Hut-78 and Jurkat cells,and decitabine treatment could signific antly reduce the methylation level of BIM gene promoter.5.Western Blot results showed that BIM protein expression was up-regulated after decitabine treatment compared with the control group,and the results were statistically significant.6.qRT-PCR results showed that BIM mRNA expression level increased after decitabine treatment compared with the control group,and the results were statistically significant.Conclusions1.BIM gene methylation level is higher in PTCL patients;2.Decitabine can induce the demethylation of BIM gene and up-regulate the expression levels of BIM protein and mRNA.