Influence of E-cadherin methylation on prognosis in children with acute lymphoblastic leukemia / 中国当代儿科杂志

OBJECTIVES@#To study the significance of E-cadherin and the association between E-cadherin methylation status and prognosis in children with acute lymphoblastic leukemia (ALL) by examining the mRNA and protein expression of E-cadherin and its gene methylation status in bone marrow mononuclear cells of children with ALL.@*METHODS@#The samples of 5 mL bone marrow blood were collected from 42 children with ALL who were diagnosed for the first time at diagnosis (pre-treatment group) and on day 33 of induction chemotherapy (post-treatment group). RT-qPCR, Western blot, and methylation-specific PCR were used to measure the mRNA and protein expression of E-cadherin and the methylation level of the E-cadherin gene. The changes in each index after induction chemotherapy were compared.@*RESULTS@#The mRNA and protein expression levels of E-cadherin in the post-treatment group were significantly higher than those in the pre-treatment group (P<0.05), while the positive rate of E-cadherin gene methylation in the post-treatment group was significantly lower than that in the pre-treatment group (P<0.05). At the end of the test, the children with negative methylation had significantly higher overall survival rate and event-free survival rate than those with positive methylation (P<0.05).@*CONCLUSIONS@#E-cadherin expression is associated with the development of ALL in children, and its decreased expression and increased methylation level may indicate a poor prognosis.

Clinical Significance of SFRP1 Gene Methylation in Patients with Childhood Acute Lymphoblastic Leukemia / 中国实验血液学杂志

OBJECTIVE@#To investigate the clinical significance of SFRP1 gene and its methylation in childhood acute lymphoblastic leukemia (ALL) .@*METHODS@#Methylation-specific PCR (MSP) was used to detect the methylation status of SFRP1 gene in bone marrow mononuclear cells of 43 children with newly diagnosed ALL before chemotherapy (primary group) and when the bone marrow reached complete remission d 46 after induction of remission chemotherapy (remission group), the expression of SFRP1 mRNA was detected by quantitative real-time polymerase chain reaction (qRT-PCR), the expression of SFRP1 protein was detected by Western blot, and clinical data of children were collected, the clinical significance of SFRP1 gene methylation in children with ALL was analyze.@*RESULTS@#The positive rate of SFRP1 gene promoter methylation in the primary group (44.19%) was significantly higher than that in the remission group (11.63%) (χ2=11.328, P<0.05). The relative expression levels of SFRP1 mRNA and protein in bone marrow mononuclear cells of children in the primary group were significantly lower than those in the remission group (P<0.05). Promoter methylation of SFRP1 gene was associated with risk level (χ2=15.613, P=0.000) and survival of children (χ2=6.561, P=0.010) in the primary group, children with SFRP1 hypermethylation had significantly increased risk and shortened event-free survival time, but no significant difference in other clinical data.@*CONCLUSION@#Hypermethylation of SFRP1 gene promoter may be involved in the development of childhood ALL, and its hypermethylation may be associated with poor prognosis.

Expression and Significance of Low-Density Lipoprotein-Related Receptors 5 and 6 in the Wnt/β-Catenin Signaling Pathway in Childhood Acute Lymphoblastic Leukemia / 中国实验血液学杂志

OBJECTIVE@#To investigate the significance of low-density lipoprotein receptor-related protein 5 and 6 (LRP5/6) in the Wnt/β-catenin signaling pathway in the pathogenesis and prognosis of childhood acute lymphoblastic leukemia (ALL).@*METHODS@#A total of 43 children who were newly diagnosed and achieved complete remission after remission induction therapy were enrolled. The children before treatment were included in incipient group, and after treatment when achieved complete remission included in remission group. A total of 39 children with immune thrombocytopenia were enrolled in control group. Three milliliter bone marrow samples were collected from above-mentioned each group. QRT-PCR was used to determine the mRNA expression of LRP5 and LRP6 in blood mononuclear cells of bone marrow. Western blot was used to detect the protein expression of LRP5 and LRP6. According to the protein expression levels of LRP5 and LRP6, the children were divided into low-expression group and high-expression group, and the clinical biological characteristics were compared between these two groups. Survival analysis was performed by Kaplan-Meier method.@*RESULTS@#Both mRNA and protein expression levels of LRP5 and 6 were upregulated in the incipient group compared with the control and remission group (P<0.05). The mRNA and protein expressions of LRP5 and LRP6 in the high-risk group were higher than those in the medium-risk group (P<0.05), it is the same as in the medium-risk group than the low-risk group (P<0.05). The mRNA and protein expressions of LRP5 and 6 positively correlated with risk degree in the incipient group (r@*CONCLUSION@#The high expression of LRP5/6 may be one of the pathogenesis of childhood ALL, and the degree of LRP5/6 increase may be related to the risk level.

Expression and Significance of Leucine-rich Repeat-containing G-protein Coupled Receptor 5/6 in Wnt Pathway in Children with Acute Lymphoblastic Leukemia / 中国医学科学院学报

Objective To study the expression and significance of leucine-rich repeat-containing G-protein coupled receptor(LGR)5/6 in childhood acute lymphoblastic leukemia(ALL). Methods A total of 39 children who had ALL and achieved complete remission on day 33 after induction therapy were enrolled.The children before induction therapy were considered as the incipient group,and those who achieved complete remission on day 33 by induction therapy were considered as the remission group.According to the degree of risk,they were assigned into 3 groups:low-risk(

Expression and significance of dishevelled proteins in the Wnt pathway in childhood acute lymphoblastic leukemia / 中国当代儿科杂志

OBJECTIVE@#To study the significance of dishevelled (DVL) proteins in the Wnt signaling pathway in the pathogenesis and prognosis of childhood acute lymphoblastic leukemia (ALL).@*METHODS@#A total of 33 children with new-onset ALL were enrolled as the case group. According to the degree of risk, they were divided into 3 groups: low-risk (n=14), intermediate-risk (n=5) and high-risk (n=14). A total of 29 children with immune thrombocytopenia were enrolled as the control group. At diagnosis and on day 33 of induction therapy, 2 mL bone marrow samples were collected from the case and control groups, and qRT-PCR was used to measure the mRNA expression of DVL1, DVL2 and DVL3 in blood cells of bone marrow.@*RESULTS@#The mRNA expression of DVL1, DVL2 and DVL3 in the case group in the incipient stage was significantly higher than that in the remission stage and the control group (P<0.05). Compared with the control group, the case group had a significant increase in the mRNA expression of DVL2 in the remission stage (P<0.05). The mRNA expression of DVL2 was significantly higher than that of DVL1 and DVL3 in both remission and incipient stages (P<0.05). The high- and intermediate-risk groups had significantly higher mRNA expression of DVL1 and DVL2 than the low-risk group (P<0.05). The mRNA expression of DVL2 was significantly higher than that of DVL1 and DVL3 in the low-, intermediate- and high-risk groups (P<0.05).@*CONCLUSIONS@#The change in the expression of DVL, especially DVL2, in the Wnt signal pathway has certain significance in the pathogenesis and prognosis of childhood ALL.

Expression of Wif-1 and β-catenin in the Wnt pathway in childhood acute lympho-blastic leukemia / 中国当代儿科杂志

<p><b>OBJECTIVE</b>To investigate the expression and possible roles of Wnt inhibitory factor-1 (Wif-1) and β-catenin in the Wnt pathway in childhood acute lymphoblastic leukemia (ALL).</p><p><b>METHODS</b>The clinical data of 35 children who had newly-diagnosed ALL and achieved complete remission on day 33 of remission induction therapy were retrospectively reviewed. The children before treatment were considered as the incipient group, and those who achieved complete remission on day 33 were considered as the remission group. Fifteen children with non-malignant hematologic diseases were enrolled as the control group. RT-PCR was used to measure the mRNA expression of Wif-1 and β-catenin. ELISA was used to measure the protein expression of Wif-1.</p><p><b>RESULTS</b>Compared with the control and remission groups, the incipient group had significantly lower mRNA and protein expression of Wif-1 and significantly higher mRNA expression of β-catenin (P<0.05). In the incipient and remission groups, high-risk children showed significantly higher mRNA expression of β-catenin and significantly lower mRNA and protein expression of Wif-1 than the medium- and low-risk children (P<0.05). In the incipient and remission group, the children with T-cell acute lymphoblastic leukemia showed significantly higher mRNA expression of β-catenin and significantly lower mRNA and protein expression of Wif-1 compared with those with B-lineage acute lymphoblastic leukemia (P<0.05). In each group, there was a negative correlation between the mRNA expression of Wif-1 and β-catenin (P<0.05).</p><p><b>CONCLUSIONS</b>Reduced expression of Wif-1 and increased expression of β-catenin may be involved in the pathogenesis of childhood ALL, and the degree of reduction in Wif-1 and/or increase in β-catenin may be related to prognosis.</p>

Effects of matrine on the proliferation and apoptosis of human rhabdomyosarcoma RD cells / 中国当代儿科杂志

<p><b>OBJECTIVE</b>To investigate the effects of matrine on the proliferation and apoptosis of human rhabdomyosarcoma RD cells in vitro, and to explore the mechanism of matrine inducing apoptosis of RD cells.</p><p><b>METHODS</b>MTT assay was used to measure the proliferation inhibition rates of RD cells that were treated with matrine (final concentrations= 0.5, 1.0, 1.5, and 2.0 mg/mL). Flow cytometry was used to evaluate the apoptosis of RD cells treated with the four concentrations of matrine. RT-PCR was used to measure the mRNA expression of cyclin D1 and survivin in RD cells treated with 0.5, 1.0, and 1.5 mg/mL of matrine.</p><p><b>RESULTS</b>The RD cells treated with various concentrations of matrine showed significantly higher proliferation inhibition rates and apoptotic rates than those that were not treated with matrine (P<0.01), and with increased matrine concentration, the proliferation inhibition rate of RD cells increased gradually, thus exhibiting a dose dependence. The mRNA expression of cyclin D1 and survivin was seen in all RD cells, but was significantly lower in RD cells treated with matrine than in those that were not treated with matrine (P<0.01). There were significant differences in cyclin D1 mRNA level among the RD cells treated with 0.5, 1.0, and 1.5 mg/mL of matrine (P<0.05), while there was significant difference in survivin mRNA level between the RD cells treated with 0.5 and 1.5 mg/mL of matrine (P<0.05).</p><p><b>CONCLUSIONS</b>Matrine can significantly inhibit proliferation and induce apoptosis of RD cells, which may be related to downregulating the mRNA expression of cyclin D1 and survivin.</p>

Effects of vector-mediated targeting human telomerase reverse transcriptase gene RNA interference on proliferation and apoptosis of leukemia HL-60 cells / 肿瘤

Objective: To investigate the effects of vector-mediated targeting human telomerase reverse transcriptase ( hTERT) gene RNA interference (RNAi) on the proliferation and apoptosis of leukemia HL-60 cells. Methods: The targeting hTERT gene RNAi recombination vector (pSilencer1.0-U6/hTERT) was transfected into HL-60 cells. The HL-60 cells transfected with empty vector (pSilencer1.0-U6), RNAi-mate or HL-60 cells were used as the controls. The expression of hTERT protein was detected by Western blotting. The proliferation and apoptosis of HL-60 cells were measured by MTT assay and flow cytometry, respectively. Results: The expression levels of hTERT protein in HL-60 cells were lower and the proliferation inhibitory rate and apoptosis rate of HL-60 cells were higher than those of each control group after transfection with pSilencer1.0-U6/hTERT for 24, 72, and 120 h ( P0.05). Conclusion: Vector-mediated targeting hTERT gene RNAi can down-regulate the expression of hTERT protein and inhibit the proliferation and induce apoptosis of HL-60 cells in vitro.

Effect of proteasome inhibitor MG-132 on L1210 cell apoptosis and its mechanism / 中国实验血液学杂志

This study was aimed to investigate the effect of proteasome inhibitor MG-132 on apoptosis of L1210 cells and its mechanism. L1210 cells were treated with MG-132 of different concentrations (0, 2.5, 5, 10, 10 micromol/L). Cell viability was tested by MTT assay, apoptosis rate was detected by using flow cytometry, activity of caspase 3 was detected by colorimetry, the expression of NF-kappaB nuclear protein was detected by Western blot. The results showed that the growth inhibition of L1210 cells treated for a same time (24 hours) was enhanced along with increasing of MG-132 concentrations (0, 2.5, 5, 10, 20 micromol/L); the inhibitory rate, apoptosis rate and activity of caspase 3 increased also along with raising of MG-132 concentrations; while the expression of NF-kappaB nuclear protein decreased along with raising of GM-132 concentrations. It is concluded that MG-132 can induce the apoptosis of L1210. The mechanism of apoptosis may be related to the down-regulation of the expression of NF-kappaB and the activation of caspase 3.

Effect of proteasome inhibitors MG-132 at different doses on cultured K562 cell apoptosis / 中国实验血液学杂志

This study was aimed to investigate the effect of proteasome inhibitor MG-132 at different doses on cultured K562 cell apoptosis. MTT assay was used to observe the activity of K562 cell proliferation inhibition rate after treating for 48 hours at different doses (0, 2, 4, 8, 16, 32 micromol/L). Immunocytochemistry was used to detect the NF-kappaB activity and glucocorticoid receptor (GR) expression. Flow cytometry was used to determine the K562 cell apoptosis. The results indicated that proliferation inhibition rate of K562 cells after treated for 48 hours showed dose-dependent, the inhibitory rates of cell proliferation in test groups were significant higher than that in control group, and the effect in 32 micromol/L test group was the most obvious (45.24 +/- 4.12)% (p < 0.05). The NF-kappaB activity and GR expression after treating for 48 hours showed dose-dependent. Compared with control group, the NF-kappaB activities in test groups were lower (p < 0.05), and the NF-kappaB activity in 32 micromol/L test group was the lowest (63.60 +/- 2.95); the GR expression in test groups was higher (p < 0.05), and the GR expression in 16 micromol/L test group was the highest (75.62 +/- 2.70). The K562 cell apoptosis rate after treating for 48 hours also showed dose-dependent. Compared with control group, the K562 cell apoptosis rates in test groups were higher (p < 0.05), the K562 cell apoptosis rate in 32 micromol/L test group was the highest (21.37 +/- 2.02)%. It is concluded that the MG-132 may induce K562 cell apoptosis and proliferation inhibition through up-regulation of NF-kappaB activity and down-regulation of GR expression both in dose-dependent manner.

Effect of N-tosyl-L-phenylalnylchloromethyl ketone and dexamethasone on expression of nuclear transcription factor-kappaB in childhood acute lymphoblastic leukemia and its significance / 中国实验血液学杂志

In order to investigate the effect of N-tosyl-L-phenylalnylchloromethyl ketone (TPCK) and dexamethasone (Dex) on expression of nuclear transcription factor-kappaB (NF-kappaB) in childhood acute lymphoblastic leukemia (ALL) and its significance, so as to provide the experimental basis for corresponding clinical treatment of ALL, in which NF-kappaB is taken as a target. The biotin-streptavidin method was used to detect the expression of NF-kappaB P65 protein and the effects of TPCK and Dex at clinically relevant dosage on activity of NF-kappaB P65 protein in 20 childhood ALL patients. The results indicated that the expression of NF-kappaB P65 protein was strongly diminished and reached to negative level at 2 hours by treatment with 40 micromol/L TPCK, the positive expression of NF-kappaB P65 protein was (2.5 +/- 1.6)%. TPCK had a time-dependent inhibitory effect on ALL cells cultured in vitro. The expression of NF-kappaB P65 protein in ALL cells was strongly inhibited by clinically relevant concentration of dexamethasone 5.0 microg/ml for 24 hours in vitro. The positive expression was (25.0 +/- 3.0)%, there was significant difference, as compared with untreated ALL cells (T=55, P<0.01). It is concluded that TPCK and Dex can inhibit NF-kappaB activity. Inhibition of NF-kappaB activity may be one of the effect mechanism of dexamethasone on ALL cells. Inhibition of NF-kappaB conduction pathway may have a significant value in childhood ALL treatment.

Different Doses of Calcium 5-Formyltetrahydrofolate for Protecting Enteral Mucosa after Chemotherapy of High-Dose Methotrexate in Rats / 实用儿科临床杂志

Objective To explore different doses of calcium 5-formyltetrahydrofolate(CF)for protecting enteral mucosa after chemotherapy of high-dose methotrexate(HD-MTX) in rats.Methods Sixty of 6 weeks old Wistar rats were divided into 5 groups in random,12 rats every group.Group A:control group,normal sodium(NS) intraperitoneal injection only;Group B to E:after HD-MTX intraperitoneal injection(120 mg/kg),1% CF(CF dose amounts to 1% of total MTX dose) for group B,2% CF for group C,8% CF for Group D and empty for group E.For group B、C and D,CF were intramuscular injected after 12 hours of MTX used,q6h?7 times.Rats were killed after 18 hours of the last time of CF.Morphous of jejunum dissection were observed and length of intestinal villus and depth of crypt were mea-sured.Results For group A,jejunum walls were thick and elastic and intestinal villus were close and orderly.Jejunum walls were congestive,swollen and thin,length of intestinal villus and depth of crypt reduced both in group B to E.These were most obvious in group E,and were secondary in group B.Statistical analysis showed that significant difference in effect existed between group B,C,D,E and group A(Pa0.05).Conclusion MTX can damage in intestinal mucosa of rats,CF can reduce this damage,excessive low doses of CF can't play this role.

Effect and Significance of Dexamethasone on Expression of Nuclear Transcription Factor-?B in Childhood Acute Lymphoblastic Leukemia / 实用儿科临床杂志

Objective To investigate the expression and significance of nuclear transcription factor-?B(NF-?B) in childhood acute lymphoblastic leukemia(ALL) and the effect of dexamethasone(DEX) on its expression,to provide the experimental base for corresponding clinical treatment of the ALL,in which NF-?B is taken as a target.Methods 1.The biotin-streptavidin method was used to detect NF-?B P65 protein on 20 childhood ALL patients and 20 healthy children.2.The effect of DEX at clinically relevant dosage on NF-?B P65 protein were also detected by the biotin-streptavidin method.Results 1.The positive expression rate of NF-?B P65 protein in childhood ALL patients was 85.50%,obviously higher than that in normal group(10.0%)(?~2=22.56 P