OBJECTIVETo investigate the methylation rate of cyclin-dependent kinase inhibitor 2A (CDKN2A) and cyclin-dependent kinase inhibitor 2B (CDKN2B) in the 9P21 region in children with acute myeloid leukemia (AML) and the association of gene methylation with clinical features and outcomes.

METHODSThe clinical data of 58 children who were newly diagnosed with AML between January 2010 and December 2012 were retrospectively analyzed. Thirty-eight healthy children were recruited as the control group. Genomic DNA was extracted from bone marrow or peripheral blood of the 58 patients and 38 healthy children. The methylation status of CDKN2A and CDKN2B was analyzed by methylation-specific multiplex ligation-dependent probe amplification.

RESULTSGene methylation was not found in healthy children. Methylation probes of 44 patients were detected in 58 patients. The methylation of CDKN2A was detected with 136 bp and 237 bp methylation probes. The methylation of CDKN2B was detected with 130 bp, 210 bp, 220 bp, and 417 bp methylation probes. The methylation rate of CDKN2A was 5%, while the methylation rate of CDKN2B was 76%. The methylation detected by some probes was associated with sex, hemoglobin, and platelet count at the first visit.

CONCLUSIONSThe methylation of CDKN2B is a common event in children with AML, while the methylation of CDKN2A is relatively rare.

Adolescent ; Child ; Child, Preschool ; Cyclin-Dependent Kinase Inhibitor p15 ; genetics ; Cyclin-Dependent Kinase Inhibitor p16 ; genetics ; DNA Methylation ; Female ; Humans ; Infant ; Leukemia, Myeloid, Acute ; genetics ; Male

OBJECTIVE: To analyze the relationship between and gene methylation with aging in the general population. METHODS: We collected peripheral blood samples from 284 male and 246 female healthy subjects for detection of methylation levels of and genes using quantitative methylation-specific PCR (qMSP). The relationship between the methylation levels of and genes and aging was analyzed using Spearman or Pearson correlation test. RESULTS: We found a significant positive correlation between the methylation levels of the two genes in these subjects ( < 0.05). In the overall population as well in the female subjects, methylation was found to be inversely correlated with age ( < 0.05). The methylation levels of and genes were inversely correlated with TG, ApoE, Lp(a) and AST in the overall population ( < 0.05). In both the female and male subjects, the methylation levels of the two genes were inversely correlated with Lp(a) ( < 0.05). In the male subjects, methylation was inversely correlated with AST ( < 0.05), while methylation was inversely correlated with HDL and ApoE ( < 0.05). In the female subjects, methylation was positively correlated with LDL and inversely correlated with ApoE and AST ( < 0.05). CONCLUSIONS The methylation levels of and are closely related to age and the levels of multiple proteins in healthy subjects.
Aging ; Cyclin-Dependent Kinase Inhibitor p15 ; metabolism ; Cyclin-Dependent Kinase Inhibitor p16 ; metabolism ; DNA Methylation ; Female ; Humans ; Male ; Real-Time Polymerase Chain Reaction

OBJECTIVETo investigate the effect of Oviductus Ranae (OR) on the expressions of CyclinD1, CDK6 and P15 in the liver of aged male rats.

METHODSEighteen male SD rats were randomly divided into 3 equal groups, namely the OR group, VE group and ageing model group. The rats received subcutaneous injection of D-galactose for 6 weeks to establish the aging models, and another 6 rats were injected daily with normal saline (NS) to serve as the normal control group. From the third week of the experiment, the rats were given oral OR or Vitamin E (VE) accordingly till the sixth week. After completion of the drug administration, all the rats were sacrificed for detecting the expressions of CyclinD1, CDK6 and P15 in the liver tissue by Western blotting.

RESULTSThe relative expression levels of CyclinD1, CDK6 and P15 in the liver of the rats in the OR group were 41.73-/+0.54, 23.29-/+0.30 and 1.49-/+0.30, respectively, significantly up-regulated as compared with those in the ageing model group (P<0.01). The expressions of the proteins were obviously down-regulated in the model group in comparison with those in the normal control group.

CONCLUSIONSOR treatment can lower the expressions of Cyclin D1 and CDK6 in the liver to enhance the liver cell proliferation in aged male rats. OR also promotes the expression of P15 through a feedback mechanism to prevent excessive proliferation of the cells. The effect of OR against ageing is mediated possibly by up-regulation of the proteins associated with the cell proliferation in the liver, a mechanism different from that of VE.

Aging ; metabolism ; Animals ; Cyclin D1 ; metabolism ; Cyclin-Dependent Kinase 6 ; metabolism ; Cyclin-Dependent Kinase Inhibitor p15 ; metabolism ; Liver ; metabolism ; Male ; Materia Medica ; pharmacology ; Rats

Gastric cancer is the result of multiple risk factors, including environmental factors, genetic factors and the interaction between them. The environmental factors mainly include dietary, Helicobacter pylori infection and family history of gastric cancer. Genetic factors mainly refer to the susceptible genes that cause epigenetic alterations in oncogenes, tumor suppress genes, cell cycle regulators, DNA repair genes and signaling molecules. This paper summarizes the susceptible genes of gastric cancer and explores the genetic basis of it.
Cyclin-Dependent Kinase Inhibitor p15 ; genetics ; Genes, Tumor Suppressor ; Genes, p16 ; Humans ; Oncogenes ; Stomach Neoplasms ; etiology ; genetics

OBJECTIVETo investigate the association of p15 and pl6 genes deletion and STKI5 gene overexpression in primary hepatocellular carcinoma (PHC).

METHODSThe carcinoma tissue and the adjacent normal tissue were taken from 30 PHC patients during operations who had had neither chemotherapy nor radiotherapy preoperatively. DNA was extracted from the tissues and PCR was used to determine the homozygous deletion of p15 exon2 (pl5E2) and pl6 exon 2 (pl6E2). RNA was extracted, cDNA was synthesized by RT-PCR, and the expression of STKI5 gene was tested by PCR. Beta-actin was used as an internal control. Average density value (ADV) of STK15 gene and that of beta-actin gene were determined in both carcinoma tissue and the adjacent normal tissue.

RESULTSThe rate of p15E2 deletion was 13.3% (4/30) and the rate of p16E2 deletion was 16.7% (5/30) in the carcinoma tissue. The p15E2 and pl6E2 co-deletion rate was 6.7% (2/30). In 19 of the 30 cases (63.3%) the expression of STK15 gene in carcinoma tissue was higher than that in the adjacent normal tissue. The ratio of ADV of STK15 gene to ADV of beta-actin gene (1.53+/-0.31) in the carcinoma tissue was significantly higher than that (0.91+/-0.25) in the paired adjacent normal tissue (t = 2.86).

CONCLUSIONThe homozygous deletion of p15E2 and p16E2 and overexpression of STKI5 gene may play a role in the oncogenesis and malignant progression of PHC.

Aurora Kinase A ; Aurora Kinases ; Carcinoma, Hepatocellular ; genetics ; Cyclin-Dependent Kinase Inhibitor p15 ; genetics ; Cyclin-Dependent Kinase Inhibitor p16 ; genetics ; Female ; Gene Deletion ; Gene Expression Regulation, Neoplastic ; Humans ; Liver Neoplasms ; genetics ; Male ; Protein-Serine-Threonine Kinases ; biosynthesis ; genetics

OBJECTIVETo detect the methylation pattern of the p15(INK4B) gene and to explore its significance in the pathogenesis of acute leukemia (AL) and leukemic transformation of myelodysplastic syndromes (MDS).

METHODSA total of 49 AL cases and 22 MDS cases were analyzed by methylation specific polymerase chain reaction (MSP) for methylation patterns in CpG islands of the p15(INK4B) gene.

RESULTSHypermethylation of the p15(INK4B) gene was found in 90% (26/29) of newly diagnosed AL, including 46% with complete methylation and 54% with partial methylation. All 3 evolved AL from MDS and 9 relapsed AL showed a methylated p15(INK4B) gene and the proportion of complete methylation was 67% and 56% respectively. Only 5 of 11 (45%) AL in remission, including 2 in complete remission (CR) and 3 in partial remission (PR), were partially methylated. The frequency of p15(INK4B) gene methylation in newly diagnosed or relapsed AL was significantly higher than that in AL in the remission stage (P = 0.002) p15(INK4B) gene methylation was found in 5 of 13 (38%) low-risk MDS (RA/RAS) patients and 80% of them showed only partial methylation. However, p15(INK4B) gene methylation was found in all 9 cases in the high-risk group (RAEB/RAEB-T), including complete methylation in 56%, significantly different from the low-risk MDS group (P = 0.002).

CONCLUSIONSHypermethylation of the p15(INK4B) gene occurs frequently in leukemia and high-risk MDS. It is possible that hypermethylation of this gene is related to the pathogenesis and development of AL and MDS. It may be used as a gene marker to detect minimal residual disease, relapse of AL and leukemic transformation in MDS.

Cell Cycle Proteins ; genetics ; Cyclin-Dependent Kinase Inhibitor p15 ; Cyclin-Dependent Kinase Inhibitor p16 ; genetics ; DNA Methylation ; Humans ; Leukemia, Myeloid, Acute ; genetics ; Myelodysplastic Syndromes ; genetics ; Precursor Cell Lymphoblastic Leukemia-Lymphoma ; genetics ; Tumor Suppressor Proteins

Among tumor suppressor genes, p15(INK4b) gene is gaining more attention for its important role in the progression of myelodyplastic syndrome (MDS). Serial studies demonstrated that highly frequent hypermethylation of p15(INK4b) gene, which is located at the 5'CpG island in the promoter region of exon 1 and is the main reason of inactivation of p15(INK4b) gene, occurs during the development of MDS towards AML. The assay of methylation-specific PCR (MSP) is sensitive to this pattern of methylation which is restricted to the MDS clone. Apoptosis mediated by cytokines such as Fas antigen and TGF-beta, and bHLH proteins is inhibited by the inactivation of p15(INK4b) gene. This may result in the evolution of MDS clone to AML. In as much as the close relationship between p15(INK4b) gene methylation and MDS, modulation of the methylation status of p15(INK4b) gene may be considered as a noval treatment modality for MDS.
Cell Cycle Proteins ; genetics ; Cyclin-Dependent Kinase Inhibitor p15 ; Cyclin-Dependent Kinase Inhibitor p16 ; genetics ; DNA Methylation ; Gene Deletion ; Genes, Tumor Suppressor ; Humans ; Mutation ; Myelodysplastic Syndromes ; etiology ; genetics ; therapy ; Tumor Suppressor Proteins

OBJECTIVETo investigate the frequency of p16 and p15 gene methylation in multiple myeloma (MM), and its relationship with bone marrow cell apoptosis and clinical outcome.

METHODSTwenty-two patients with MM were studied to detect p16 and p15 gene methylation. Methylation-specific polymerase chain reaction (MSP) was used to detect gene methylation, and terminal transferase-mediated dUTP nick end-labeling (TUNEL) was used to detect cell apoptosis.

RESULTSp16 and/or p15 gene methylatoin was detected in 10 of 22 patients (45.4%). There were 3 patients with p16 gene methylation, 9 patients with p15 gene methylation, and 2 patients with both genes methylation. The incidence of methylation of p15 gene was higher than that of p16 gene (P < 0.05). The patients with p16 and/or p15 gene methylation had a delayed cell apoptosis, poor response to chemotherapy, and a short over-all survival (OS).

CONCLUSIONThe methylation of p16 and/or p15 gene plays a key role in MM apoptosis pathogenesis. The patients with both p16 and p15 gene methylation had a poor prognosis.

Apoptosis ; Cell Cycle Proteins ; Cyclin-Dependent Kinase Inhibitor p15 ; Cyclin-Dependent Kinase Inhibitor p16 ; genetics ; DNA Methylation ; DNA, Neoplasm ; genetics ; Gene Silencing ; Genes, p16 ; Humans ; Multiple Myeloma ; genetics ; pathology ; Prognosis ; Transcription Factors ; genetics ; Tumor Suppressor Proteins

OBJECTIVETo explore the proliferation and invasive effects of inhibitors of kinase 4(INK4)(P15(ink4b) and P16(ink4a)/CDKN2) gene protein activation on RKO human colorectal cell in vivo and in vitro.

METHODSRKO human colorectal cell line was exposed to the specific DNA methyltransferase inhibitor 5-Aza-CdR and INK4(P15(ink4b) and P16(ink4a)/CDKN2) protein expression was detected by Western blotting. Soft agar cloning experiment and Transwell chamber assay were used to detect the proliferative and invasive ability in vitro. Tumorigenicity in nude mice was analyzed in vivo.

RESULTSINK4(P15(ink4b) and P16(ink4a)/CDKN2) protein expression of RKO human colorectal cells after exposure to 1×10(-7), 5×10(-7) and 1×10(-6) mol/L 5-Aza-CdR concentrations(A, B, C groups) were 1.13, 1.38, 1.92 folds and 1.11, 1.45, 2.14 folds compared to positive control group respectively. Soft agar cloning experiment showed the number of cell colony significantly decreased from 36.8±5.1(positive control group) to 32.4±7.2, 21.3±5.4 and 19.5±6.4 (3 experiment groups, all P<0.05) respectively. Transwell chamber assay showed that migrated cell number in positive control group(67.4±7.2) was significantly higher than those of 3 experimental groups(35.3±4.6, 29.5±7.3 and 25.3±6.2, respectively). The tumor volume of metastasis model in nude mice was inhibited in experimental groups, but not significantly lower compared to control group (P>0.05). There were significant differences of tumor weight and inhibition rate between control group and 3 experimental groups in nude mice respectively(all P<0.01).

CONCLUSIONINK4(P15(ink4b) and P16(ink4a)/CDKN2) protein activation can inhibit tumor proliferation, migration and suppress the tumor formation ability.

Animals ; Cell Line, Tumor ; Cell Proliferation ; Colorectal Neoplasms ; metabolism ; pathology ; Cyclin-Dependent Kinase Inhibitor p15 ; genetics ; metabolism ; Cyclin-Dependent Kinase Inhibitor p16 ; genetics ; metabolism ; Humans ; Mice ; Mice, Inbred BALB C ; Neoplasm Metastasis ; Neoplasm Transplantation ; Transcriptional Activation

OBJECTIVETo detect p15(INK4B) methylation levels and the kinetics of the methylation status before and after decitabine to explore its influences on prognosis and response to decitabine in myelodysplastic syndromes (MDS) patients.

METHODSWe examined 261 MDS patients (143 male and 118 female) with the median age of 52 years (32-78). Of them, 172 cases were low-risk group (low-risk 104 cases, intermediate-1 68 cases), 89 cases high-risk group (intermediate-2 52 cases, high risk 37 cases). Collections of bone marrow mononuclear cells of MDS patients and extracted the genomic DNA, the methylation status of p15(INK4B) was detected by methylation-specific PCR (MSP) method. Survival analysis was conducted according to the level of p15(INK4B) methylation in the cohort of patients. The kinetics of the methylation levels of p15(INK4B) in 58 MDS patients before and after 2 courses of decitabine have been assessed with the method of MSP.

RESULTSThe methylation level of p15(INK4B) in low-risk group patients was significantly lower than that in high-risk group (117.22 vs 157.63, P<0.05 ). The expected 2-year survival rate of p15(INK4B) methylation positive patients was lower than that of negative ones (91.8% vs 69.8%, P<0.05); the expected 2-year survival rate of p15(INK4B) methylation positive patients was shorter than that of negative ones in low-risk group（78.2% vs 92.0%, P<0.05）, meanwhile there was no significant difference in terms of expected 2-year survival rate and median expected survival between p15(INK4B) methylation positive and negative patients in high-risk group [35.6% vs 38.5%, (17.0±9.3) month vs (18.0±5.7) month, P>0.05]. Multivariate analysis showed p15(INK4B) methylation degree was an independent prognostic factor for overall survival. No statistical difference of overall response (OR) rates were found between p15(INK4B) methylation positive patients and negative patients before decitabine（65.9% vs 76.5%, P>0.05）, and complete remission (CR) rates between these two groups also showed no statistical difference（22.0% vs 29.4%, P>0.05）. p15(INK4B) methylation levels had no obvious change before and after treatment in decitabine responders（P>0.05）.

CONCLUSIONThe survival of newly diagnosed MDS patients with positive p15(INK4B) methylation was comparatively shorter, but p15(INK4B) methylation had no influence on response to decitabine.

Adult ; Aged ; Azacitidine ; analogs & derivatives ; Cyclin-Dependent Kinase Inhibitor p15 ; genetics ; DNA Methylation ; Female ; Humans ; Male ; Middle Aged ; Myelodysplastic Syndromes ; drug therapy ; genetics ; Prognosis ; Survival Rate ; Treatment Outcome