Promoter Methylation Of EDNRB Gene In Acute Leukemia

BACKGROUND and PURPOSEThe only natural way of DNA modification of vertebrate is DNA methylation, DNA Methylation is an important part of the Epigenetics, and play an important role in the stability of genetic structure, gene expression and regμlation, etc,and are closely linked with tumor occurrence and development. Some genes found in Leukemia cells, mainly are tumor suppressor genes (such as ER, p15,p16, etc.),if the promoter region of CpG islands occurs abnormal methylation,will resμlt in gene expression reduced or closed down,affect the normal function of cells,and at last leading to the occurrence of the disease.The gene of Endouthelin Receptor B is located in 13q22, Approximately 24kb, it often has gene deletion of many cancer in this area, containing seven exons and six introns, Encoding a 442 amino acid protein, is a G protein-coupled transmembrane receptor protein.Endothelin is a vasoactive peptide, has three different forms of ET1, ET2,and ET3.In addition, EDNRB involved in the process of activating NO synthase. Studies have found that the genes associated with normal growth and development. EDNRB gene as a tumor suppressor gene, play an important role in a variety of tumor occurrence and development, the gene's expression reduced in many tumors. It has been confirmed that it has allelic loss of heterozygosity (LOH) and replication error (REP) in variety of tumors.The study found:The tumor tissue in hepatocellμlar carcinoma, bladder cancer, nasopharyngeal cancer, prostate cancer, breast cancer, etc, exists aberrant hypermethylation in the promoter CpG island of EDNRB gene, while the corresponding benign tissue and normal tissue had no hypermethylation of the gene, after demethylation of drug treatment can restore the expression of the gene.Taiwan's Hsiao has reported the hypermethylation status of the p16 and the EDNRB gene in the untreated leukemia and mμltiple myeloma cells. In this study, we will further tests the methylation status of the EDNRB gene in newly diagnosed acute leukemia, complete remission of acute leukemia and relapsed leukemia, And analysis the relationship between the gene and clinical features, and then further explore the relationship between the methylation pattern's change of the EDNRB gene promoter CPG islands and the occurrence and development of leukemia.MATERIALS AND METHODSObjectall the specimens of cases were 39, all derived from Hematology outpatient and inpatient who have diagonosed acute leukemia (AL) in the First Affiliated Hospital of Zhengzhou University on February to September 2009,all the confirmed cases were characterized by cell morphology, histochemistry, immunology, molecμlar biology and (or) cell genetics.21 males and 18 females aged 16-70 years old.Experimental group1.The untreated group of acute leukemia have 17 patients (AML 10 cases, ALL 7 cases);2.Complete remission group have 13 cases; 3.the relapsed or refractory group of acute leukemia have 9 cases; 4. The normal control group has 8 cases.2ml bone marrow blood specimen frome each patient of the untreatment group, complete remission group and relapsed or refractory group,In the normal control group,2ml bone marrow specimens from the healthy donors who present hematopoietic stem cell transplantation for patients, EDTA tubes were used to save and backup in -20℃.MethodsUsing methylation-specific polymerase chain reaction (MS-PCR) to detected methylation status of the promoter CpG islands of the EDNRB gene in mononuclear cells of the normal bone marrow blood, As well as the blood of patients with acute leukemia bone marrow.Data processingUsing SPSS17.0 software to analysis the experimental data statistically, Using x2 test for the statistics, P=0.05 for the test. RESMLTS1.There is no methylation of EDNRB gene in any cases of the normal control group.2.The methylation rate of EDNRB was 82%(14/17),31%(4/13),89%(8/ 9).respectively in untreated AL group, complete remission AL group and in relapsed AL group.There was significant difference amoun the three groups, p=0.003.Further compared between the two, we found that the untreated group compared with the complete remission group, the relapsed or refractory group compared with the complete remission group, there were significant differences, p<0.05.There was no significant difference between the untreated group and the relapsed or refractory group, p>0.05.3.The methylation of EDNRB gene was not related to gender, age, and the type of leukemia, There was no significant difference, p>0.05.CONCLUSIONS1.The aberrant hypermethylation pattern of EDNRB gene involved in the pathogenesis of AL.2.The aberrant methylation of EDNRB gene may be related to the states of untreated AL, complete remission AL and relapsed AL, well there was not correlated with gender, age, and the type of leukemia.3.Methylation-specific polymerase chain reaction (MS-P) is a sensitive experimental method to detecte the methylation status of EDNRB gene.