Triptolide affect the methylation status of HL-60cells / 中华血液学杂志

ABSTRACT

<p><b>OBJECTIVE</b>To study the effect of triptolide (TP) on the methylation status of human promyelocytic leukemia cells (HL-60) and explore a preliminary demethylation mechanism.</p><p><b>METHODS</b>Normal HL-60 cells as control group, the cell proliferation level of HL-60 cells was detected by MTT assay, being treated by different concentration TP (3.125, 6.25, 12.5, 25 nmol/L) for 24 h or 48 h respectively; Choosing the 3.125 nmol/L and the 6.25 nmol/L TP affected HL-60 cells for 48 h, the cell apoptosis rate and cell cycle were determined by flow cytometry, the expressions of death-associated protein kinase 1 (DAPK-1) and methyltransferase DNMT1, DNMT3B mRNA were measured by real time-PCR (RT-PCR), LINE-1, DAPK-1 genes'methylation variations were analyzed by methylation specific PCR (MSP).</p><p><b>RESULTS</b>Compared with control group, the different concentration TP could significantly inhibit the proliferation of HL-60 in a time-dose dependent manner (P<0.05, P<0.01). After being treated by TP for 48 h, the cell early apoptosis rate of control group and 6.250 nmol/L TP group were (2.07 ± 1.91)%, (9.77 ± 3.52)%, respectively (P<0.05); When the TP concentration increased, DAPK-1mRNA expression increased (P<0.01), DNMT1, DNMT3B mRNA expression significantly dampened (P<0.01); the promoter of LINE-1, DAPK-1 genes were hypermethylation state in the control group, after being treated by TP for 48 h, the brightness of LINE-1, DAPK-1 genes'methylation strips weakened, and the non-methylation strips enhanced all in a dose-dependent manner.</p><p><b>CONCLUSION</b>TP could down-regulate the transcriptional expression of methyltransferase DNMT1/3B genes, indirect action to reduce the degree of DAPK-1, LINE-1 genes mathylation, thus promote DAPK-1 gene expression level and inhibit the HL 60 cell growth.</p>