Cell Cycle Related Molecular Mds Patients

Objective The occurrence and development of cancer are extremely related to theaberrant cell cycle. We investigated the expression of Cyclin-dependent kinase inhibitors(p21cip1, p27kip1, p57kip2) and the changes of cell cycle in patients with MDS andexplored the possible mechanisms, to provide new ideas in the pathogenesis of MDS andtargeting therapy.Methods We performed flow cytometric analysis of cell apoptosis and cell cycle; Theexpression of p21cip1、p27kip1、p57kip2、p73and CXCR4mRAN were detected byreal-time quantitative PCR; Methylation of p73promoter was detected byMethylation-specific PCR (MSP); The results of MSP were confirmed by bisulfitesequencing; We chose samples from3low risk MDS,3high risk MDS and4healthyvolunteers and searched the role of SDF-1/CXCR4signal in gene expression (p21cip1、p27kip1、p57kip2) and changes of cell cycle in vitro. Primary cell from MDS patients weretreated with decitabine alone or together with cytarabine, then the changes of p73methylation status, the expression of p73and p21mRNA, cell apoptosis and cell cyclewere measured. We also explored the role of p73methylation in the prognosis of MDS bycorrelated it with clinical data. Moreover, the p73methylation status was analyzed both atthe time of diagnosis and after decitabine treatment in32patients with MDS.Results The expression of p21cip1was reduced in high risk MDS group and AMLand MDS-AML group compaired with healthy controls (P=0.008,0.029，respectively).However, there was no significant difference between low risk MDS groupand healthy control. The expression of p27kip1was no significant differences among thethree groups. The expression level of p57kip2were significantly reduced in low risk MDS,high risk MDS and AML and MDS-AML groupcompared with control group (P<0.001).Patients with abnormal karyotype showed lower expression of p57kip2gene, compared topatients with normal karyotype (P=0.045). Although the expression of CXCR4was no significant differences between MDS and healthy controls, we found a positive correlationbetween CXCR4and p57kip2（r=0.652, P<0.001, n=67）. There were more S phase cells inhigh risk MDS group and AML and MDS-AML group compaired with healthy controls（P<0.05）. p57expression in bone marrow mononuclear cells (BMMCs) from normalcontrols increased significantly when co-cultured with SDF-1in vitro, which could beblocked by AMD3100, whereas SDF-1induced a mild increase of p57expression in MDSwhen compared to normal controls. p73methylation was present in37.04%of these casesand patients with high risk MDS (RAEB-1and RAEB-2) exhibited a significantly higherfrequency of p73methylation than low risk MDS (29.7.8%vs.58.8%, P=0.002); Theexpression of p21cip1and p73mRNA in the methylated sample group was decreasedcompared to the unmethylated group (P=0.011,0.032, respectively). Additionally, apositive correlation between p21cip1and apoptosis was investigated in MDS patients(r=0.675, P<0.001). Decitabine treatment decreased the level of p73methylation, andincreased the level of p73transcripts and G2/M phase cell.When treated with decitabineand cytarabine together, decitabine treatment restored the p73expression and increased thesensitivity of MDS cells to cytarabine. Patients with p73gene methylation had asignificantly shorter survival than patients without methylation (P=0.002). In a Coxmultivariate analysis, p73gene methylation status and BM blast emerged as anindependent prognostic factor for OS. Although p73was demethylated after the decitabinetherapy, we did not find any significant association between p73methylation and theclinical responses.Conclusion The BMNC from patients with MDS exist abnormality in cell cycle andCyclin-dependent kinase inhibitors. SDF-1treatment significantly increased the p57kip2level dose dependently in BMMCs from healthy volunteers and Reduced responded toSDF-1contribute to the low expression of P57kip2in BMMCs from MDS patients. P73gene promoter hypermethylation, which was common in patients with MDS, may controlcell cycle and cell apoptosis by high in patients with MDS is relatively common, may beaffected by regulating the expression of downstream gene p21cip1.