Adolescent ; Child ; Child, Preschool ; Core Binding Factor Alpha 3 Subunit ; genetics ; metabolism ; DNA Methylation ; Female ; Humans ; Infant ; Lymphoma ; genetics ; metabolism ; Male ; Promoter Regions, Genetic

OBJECTIVETo investigate the methylation status of zonula occludens-1 (ZO-1) gene promoter and its clinical significance in children with stage IV non-Hodgkin lymphoma (NHL) and to provide a basis for further etiological study and early diagnosis of this disease.

METHODSFifty-five children with a confirmed diagnosis of stage IV NHL (40 cases of T-NHL and 15 cases of B-NHL) were selected as the case group, and 20 children with diseases other than hematologic malignancies were selected as the control group. Bone marrow samples were collected from these subjects. Methylation-specific PCR (MS-PCR) was applied to evaluate the methylation status of ZO-1 gene promoter, and the integrated optical density (IOD) was determined. RT-PCR was used to measure the mRNA expression of ZO-1.

RESULTSMS-PCR showed that the methylated bands of ZO-1 gene promoter were found in 39 (70.9%) of 55 patients in the case group before treatment, while no ZO-1 gene promoter methylation was detected in the control group. With close tracking of 47 cases in the study group, consisting of 32 cases of T-NHL and 15 cases of B-NHL, the rates of ZO-1 gene promoter methylation prior to treatment were 72% and 67%, respectively, (P>0.572). The cases of T-NHL and B-NHL showed no significant changes in methylation rate in the early and middle phases of chemotherapy (P>0.05), but they showed significant changes in methylation rate in the late phase of chemotherapy (P<0.05). RT-PCR showed that the NHL cases carrying methylated ZO-1 gene had no mRNA expression of ZO-1, while all children in the control group had mRNA expression of ZO-1. There was no linear relationship between the total number of peripheral blood leukocytes and ZO-1 gene IOD (r=0.093, P=0.575); a positive correlation was found between the number of malignant cells in bone marrow and ZO-1 gene IOD (r=0.669, P<0.001).

CONCLUSIONSZO-1 gene shows a hypermethylation status in children with NHL, and the methylation level is positively correlated with the number of malignant cells in bone marrow. ZO-1 may be used as a novel molecular marker in early diagnosis, outcome assessment, prognostic evaluation, and detection of minimal residual disease.

Adolescent ; Child ; Child, Preschool ; DNA Methylation ; Female ; Humans ; Infant ; Lymphoma, Non-Hodgkin ; genetics ; Male ; Promoter Regions, Genetic ; Zonula Occludens-1 Protein ; genetics

OBJECTIVETo study the changes and significance of plasma cardiotrophin-1 (CT-1) in neonates with hypoxic-ischemic encephalopathy (HIE) complicated by myocardial ischemic injury.

METHODSForty-five neonates with HIE (15 mild cases, 24 moderate cases and 6 severe cases) were enrolled and divided into two subgroups based on the presence of myocardial injury (n=19) and not (n=26). Twenty healthy neonates were used as the control group. Plasma CT-1 levels were measured using double-antibody sandwich enzyme immunoassay method. Serum creatinine kinase MB (CK-MB) and cardiac troponin I (CTnI ) levels were also measured.

RESULTSPlasma CT-1 levels in the mild HIE (169±20 pg/mL) and moderate/severe HIE subgroups (287±44 pg/mL) were significantly higher than those in the control group (30±8 pg/mL), and plasma CT-1 levels were associated with the severity of HIE (P<0.01). Plasma CT-1 levels were positively correlated with serum CK-MB and CTnI levels in neonates with HIE in the acute phase (r=0.565 and 0.621 respectively; P<0.01). Plasma CT-1 levels in neonates with myocardial injury were significantly higher than those without myocardial injury (249 ±35 pg/mL vs 177±26 pg/mL; P<0.01). Plasma CT-1 levels were significantly reduced in neonates with myocardial injury in the convalescent phase (157±19 pg/mL) compared with those in the acute phase (249±35 pg/mL; P<0.01).

CONCLUSIONSDetection of plasma CT-1 levels may be useful in the diagnosis of myocardial ischemic injury and the severity evaluation of HIE.

Creatine Kinase, MB Form ; blood ; Cytokines ; blood ; Female ; Humans ; Infant, Newborn ; Male ; Myocardial Ischemia ; blood ; Troponin I ; blood

Objective To study and discuss what part does methylated Id4 gene participate in malignant lymphoma stage Ⅳ by detecting the extent of how much Id4 gene has been methylated in afflicted children who suffer from malignant lymphoma.Methods Forty-two patients who had diagnosed with malignant lymphoma [Hodgkin's disease (HD),Non-Hodgkin's lymphoma(NHL)] were selected as study group.Their chemotherapy stages of pre-treatment,early-treatment,mid-treatment,post-treatment and clinical remissions or relapse throughout the entire treatment had been traced.At each stage the expression of methylated Id4 gene mRNA was detected by methylation-specific polymerase chain reaction (MS-PCR) and compared with the control group.The control group consisted of 20 non-neoplastic hematologic disorder affected children as sample donors.Results MS-PCR detection:in pre-treatment stage,there were 27 patients who were found methylated or partially methylated Id4 genes.Methylated ratio was thus at 64.3％ (27 patients out of a total of 42 patients).Those 27 patients were actively traced down along with different stages of treatment (21 NHL patients,6 HD patients).Before NHL there was 55.6％ methylated Id4 gene (15 cases out of 27 NHL patients).During chemotherapy treatment,there was 51.9％ of methylated Id4 gene positive (14 cases out of 27 patients).In post chemotherapy treatment,there was 48.1％ of methylated Id4 gene positive (13 cases out of 27 patients).Totally there were 9 patients showed clinical recovery after chemotherapy.There was 44.4％ of traceable methylated Id4 gene after recovered chemotherapy patients (4 recovered patients still carrying positive reading of methylated Id4 gene out of totally 9 recovered patients).There were 2 patients relapsed,with traceable methylated Id4 gene re-appeared in them afterwards.Throughout different treatment stages,there was no significant correlation in the treatment result and the appearance of methylated Id4 gene in early treatment stages (all P ＞ 0.05).But in latter treatment of chemotherapy,the correlation started to emerge (P ＜ 0.05) ; The overall statistics on NHL and HD share the same statistical pattern on different stages (all P ＞ 0.05).The control group had 20 patients,none of them had methylated Id4 gene.The study group showed noticeable difference in methylated Id4 gene before and after the experiment (P ＜ 0.001).RT-PCR result showed that before chemotherapy treatment,all of those who carried methylated Id4 gene had no expression of mRNA,by comparison to the control1 group which all had expression of mRNA.Conclusions Methylated Id4 gene is closely related in affected children who suffer from malignant lymphoma and its complications.The expression of Id4 gene is depressed when it has been methylated.The state of methylated Id4 gene is changed as patient's condition changed,so the methylated Id4 gene is thus a possible indicator of early diagnostic tool for children lymphoma.

OBJECTIVETo study methylation of Id4 gene and demethylation effect of arsenic trioxide (ATO) in Raji cells.

METHODSHuman Burkitt's Raji lymphoma cells were cultared and treated with ATO at different concentrations and different time points. Methylated degree of Id4 gene was detected by methylation specificity polymerase chain reaction (MS-PCR), Id4 mRNA expression in Raji cell by reverse transcription polymerase chain reaction (RT-PCR), the growth of cell by MTT assay, and cell apoptosis and cycle distribution by Flow Cytometry (FCM).

RESULTS(1) The Id4 gene exhaustive methylation in control group, and hypermethylation in experimental group were reversed by ATO in a dose-dependent manner. (2) Id4 mRNA expression in Raji cells treated with ATO for 48 h increased gradually with ATO concentration increasing in experimental group. (3) Raji cell growth inhibited rates after different concentrations of ATO treatment for 24, 48, 72 h were 12.15% ∼ 92.17% in the experimental group (P < 0.05). (4) Apoptosis peak emerged after ATO treatment for 48 hours in experimental group, while a much lower apoptosis in control group. (5) After ATO treatment for 48 h in experiment group, the cells were arrested at G(0)/G(1) phase in a dose-dependent manner.

CONCLUSIONId4 gene presents exhaustive methylation in Raji cells. ATO can reverse the hypermethylation of Id4 gene and recover the expression of Id4 mRNA. Hypermethylation of Id4 gene is one of the reasons of Raji cells malignant proliferations.

Apoptosis ; drug effects ; Burkitt Lymphoma ; genetics ; Cell Line, Tumor ; DNA Methylation ; Humans ; RNA, Messenger ; genetics

OBJECTIVES: To investigate the expression and significance of jumonji domain-containing protein 2B (JMJD2B) and hypoxia-inducible factor-1α (HIF-1α) in non-Hodgkin's lymphoma (NHL) tissues in children. METHODS: Immunohistochemistry was used to detect the expression of JMJD2B and HIF-1α in lymph node tissue specimens from 46 children with NHL (observation group) and 24 children with reactive hyperplasia (control group). The relationship between JMJD2B and HIF-1α expression with clinicopathological characteristics and prognosis in children with NHL, as well as the correlation between JMJD2B and HIF-1α expression in NHL tissues, were analyzed. RESULTS: The positive expression rates of JMJD2B (87% vs 21%) and HIF-1α (83% vs 42%) in the observation group were higher than those in the control group (P<0.05). The expression of JMJD2B and HIF-1α was correlated with serum lactate dehydrogenase levels and the risk of international prognostic index in children with NHL (P<0.05). The expression of JMJD2B was positively correlated with the HIF-1α expression in children with NHL (r_s=0.333, P=0.024). CONCLUSIONS JMJD2B and HIF-1α are upregulated in children with NHL, and they may play a synergistic role in the development of pediatric NHL. JMJD2B can serve as a novel indicator for auxiliary diagnosis, evaluation of the severity, treatment guidance, and prognosis assessment in pediatric NHL.
Humans ; Child ; Hypoxia-Inducible Factor 1, alpha Subunit ; Prognosis ; Hypoxia ; Lymphoma, Non-Hodgkin