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Objective Based on our previous established cohort of myelodysplastic syndrome (MDS), we investigated the potential effect of beta-tubulin (TUBB) gene in the transformation of MDS into acute leukemia Methods From our nested case-control study cohort of MDS patients, we chose 11 paired transformed and nontransformed MDS patients.TUBB gene expression was tested by quantitative real-time PCR.TUBB-siRNA transfection was used to down-regulate TUBB gene expression in SKM-1 cell line.The function of TUBB gene in SKM-1 cell line was evaluated by cell proliferation, soft agar clone formation and electron microscope.Results TUBB gene expression in MDS patients in transformed group were significantly higher than that in control group (2.91 ± 0.41 vs 0.90 ± 0.23, P ＜0.01).After TUBB-siRNA transfection, A450/630nm of SKM-1 cells at 24 h, 48 h and 72 h were 0.299 ± 0.045, 0.526 ± 0.034 and 0.652 ± 0.035, respectively, which were significantly decreased than those in negative-siRNA group (0.438 ±0.074, 0.858 ±0.064 and 0.974 ±0.044) (P ＜0.05).Soft agar clone formation in TUBB-siRNA group was (7.0 ±0.2)％, which was significantly reduced than that of negative-siRNA group (25.0 ± 0.2)％ (P ＜ 0.01).Electron microscope showed significant apoptotic signs in TUBB-siRNA group, including vacuoles in cytoplasm and karyorrhexis.Conclusion Our results indicate that TUBB gene may play a role in the transformation of MDS into acute leukemia by affecting the proliferation of malignant clones.
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<p><b>OBJECTIVE</b>To identify methylation profiles in myelodysplastic syndrome (MDS) and to provide the biomarkers for the early diagnosis and differential diagnosis of MDS.</p><p><b>METHODS</b>Genes were screened for hypermethylation by genome-wide DNA methylation profiles. Transcription down-regulation was determined with a gene expression microarray. Methylation-specific, real-time, and bisulfite-sequencing PCR cloning and sequencing were performed to validate selected genes in MDS cases and non-malignant hematologic diseases (controls). Diagnostic test, such as sensitivity and specificity, was used to evaluate the value of methylation patterns.</p><p><b>RESULTS</b>A draft of methylation patterns was established and refined to 6 genes after validation in 211 patients and 60 controls. The hypermethylated genes were ABAT (97%), DAPP1 (98%), FADD (89%), LRRFIP1 (96%), PLBD1 (89%), and SMPD3 (85%). A combination of 5 or more than 5 genes showed a specificity of 95% and sensitivity of 91.4% for the diagnosis of MDS. The accuracy of diagnosis was 92.3%.</p><p><b>CONCLUSION</b>We demonstrated here that the ABAT, DAPP1, FADD, LRRFIP1, PLBD1 and SMPD3 genes are hypermethylated and downregulated in MDS. The six genes could be the markers of the methylation patterns in MDS, as a noninvasive approach for the diagnosis of MDS.</p>
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Humans , DNA Methylation , Down-Regulation , Gene Expression , Genome, Human , Myelodysplastic Syndromes , Diagnosis , GeneticsABSTRACT
Objective To establish a novel glucocorticoid (GC)-resistant human diffuse large B lymphoma(DLBCL) cell line Toledo/dexamethasone (DEX) by the exposure to DEX,and observe the biological characteristics of resistant and parental cell line,investigate the mechanisms of glucocorticoid-resistance.Methods Toledo/DEX was established by the exposure to DEX,the dose of which was increased gradually and intermittently for long periods of time.Toledo,in the logarithmic growth phage,was incubated in the culture medium containing DEX at the concentration of 1×10-8 mol/L at first.The medium without DEX was replaced after for 96 hours until the cell line re-entered the logarithmic growth phase.Repeat the above steps to acquire the ultimate concentration of DEX in the medium as 1.024 ×10-5 mol/L.The biological characteristics of resistant and parental cell lines were evaluated.Results Toledo/DEX was more invasive in the aspects of ultrastructure,tumorigenicity and drug sensitivity.Meanwhile,Toledo/DEX achieved some stable biological characteristics such as morphology,karyotypes and immunophenotype.Furthermore,GC receptor (GR) α and GR β protein expression analysis showed that GR was involved in the mechanism of the GCresistance.Conclusions Toledo/DEX is a drug-resistant cell line with a stable biology backgroud.These results may help shed light on the knowledge of GC-resistance and lay the groundwork for searching new therapeutics to reverse drug-resistance.
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Objective To survey the incidence of acquired adult aplastic anemia (AA) in Shanghai, China. Meanwhile, we compared it with the previous data from China in 1986 and other countries in order to explore the trends. Methods Newly diagnosed AA patients were registered in 6 districts (Jingan, Xuhui, Huangpu, Changning, Putuo, Yangpu ) in Shanghai from 2004 to 2006. Then we calculated the crude and age-adjusted incidence of AA according to the population data from Shanghai Statistic Yearbook. Results There were 38 adult patients with acquired AA. The average crude incidence of AA was 0. 33/100 000 from 2004 to 2006. The incidences per 100 000 persons per year were 0. 40, 0. 14and 0. 64 in 18-34, 35-59 and ≥ 60 years, respectively. The rate of severe AA was 0. 17/100 000.Conclusion The incidence of severe AA has no marked change, but the total rate is a little decreased compared with the data from China in 1986.
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Objective To determine risk factors of environment and lifestyles involved in acute hematology departments of 29 secondary-care and tertiary-care hospitals in Shanghai. There were 206 hospitalized patients with de novo AML during June 2003 to April 2007 meeting the diagnostic criteria for AML by the World Health Organization, excluding those with definite history of radiotherapy or chemotherapy, and 412 age- and gender-matched controls with non-hematological other diseases from the same hospitals. All the participants were interviewed face-to-face using a standard structured questionnaire,including personal medical history, family medical history, history of medications, traditional Chinese herb medicines, exposure to occupational hazards, lifestyle factors, etc. in the past five years prior to onset of the disease. Conditional univariate and multivariate logistic regression analyses were performed to identify risk factors for AML. Results Logistic regression analysis showed that taking traditional Chinese herb medicine might be a protective factor for AML, with an odds ratio (OR) of 0. 39 and its 95% confidence interval (CI) of 0. 16-0. 98. Potential risk factors for AML included history of diabetes and taking hypoglycemic agents ( OR: 1.39, 95% CI: 0.67 -2. 87), occupational exposure to metal ( OR: 1.73, 95% CI: 0. 77 -3. 91 ), glue, paint and other coating materials, etc. ( OR: 1.72, 95% CI: 0. 93 - 3.2), raising animals (OR: 1.33, 95% CI: 0. 79 -2. 24). No association of living within a distance of 100 meters from high voltage power transmission lines, home renovation and decoration in the past year, hair dyeing, smoking,alcohol drinking with AML in adults was found. Conclusions Factors mentioned above such as occupational exposure to metal, glue, paint and other coating materials potentially increase risk for AML in adults,however taking traditional Chinese herb medicine is possibly a protective factor for it.
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Objective To investigate the characteristics of dysplasia in myeledysplastic syndrome (MDS). Methods 716 samples of adult patients with abnormal blood routine and unelear cause were collected between July 04, 2003 and March 14, 2007. Based on the gold diagnostic standard of WHO MDS classification, all eases were detected on cytomorphology, cytochemical stain, bone marrow pathological assay,cytogenetics, flow eytometry et al. The cytological study of bone marrow on some abnormal hematopaietie cells has a diagnostic value to determine clonal or non-clonal diseases and assess sensitivity and specificity. Results in the complicated various dysplasia of hematopeiefic ceils, the following characteristics can be the main basis of cytomorphological diagnosis: one of granular Auer bodies, micronuclens (MN), or nuclear budding, erythroid nuclear budding, megakaryocytes presented in peripheral blood, myeloblast or prorubricyte exhibited in peripheral blood, ringed sideroblasts>1%. The subordinate basis of cytomorphological diagnosis development of nuclei, ring-shaped nuclei, and aggregation of nuclear chromafin, erythroid multi-nuclei, odd nucleus, mother-daughter nucleus, nuclear fragmentation, vacuole, anisoeytosis and mieromegakaryocytes.Conclusion Cytomorphologic assay is the base for the diagnosis of MDS, however, it presents certain limit,especially when eytomnrphoiogical change does not possess specificity for early MDS. Hereby, it requires to combine other deteetion methods.
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Objective To explore the polarization and proliferation of naive CD4+TT cells after preincubation with interleukin-2. Methods The expression of suppressor of cytokine signal (SOCS)-3 was detected by the real-time PCR in the naive T lymphocytes of DO11.10 TCR transgenic and C57BL/6N mice after they were preincubated by 50 U/ml of IL-2. The naive CD4+T T lymphocytes preincubated for 4 h by IL-2 were stimulated with ovalbumin (OVA) and inactivated BALB/c spleen cells, respectively. After 14 d, IL-12Rβ1, IL-12β2 and cytoplasm IL-4 (cyIL-4) were detected by flow cytometry. Results SOCS-3 in the naive CD4+T lymphocytes of DO11. 10 TCR transgenic and C57BL/6N mice reached the peak after preincu- bated by IL-2 for 6 h. The polarization to TH1 and the proliferation ability of naive CD4+ T lymphocytes stimulated with specific antigen and allogeneic antigen were inhibited conspicuously during the time of the peak of SOCS-3. Conclusion The expression of SOCS-3 in naive CD4+T lymphocytes can be upregulated by IL-2. The upregulation of SOCS-3 can suppress the polarization to TH1and the proliferation ability of naive CD4+T lymphocytes stimulated by specific antigen and allogeneic antigen.
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Hereditary hemolytic anemia(HHA)is a hereditary disorder with main clinical presentation of hemolysis and hemolytic anemia.It's the most common disease of hereditary disorders in the world.According to the pathogenic mechanism,the disorders can be divided into three groups:abnormalities of the red cell membrane,red cell enzyme disorders and hemoglobinopathies.The most common HHA are hereditary spherocytosis,G6PD deficiency and the thalassemias.The clinical presentation of HHA may be heterogeneity.No anemia may occur in the early stage or mild HHA.In order to detect HHA early,we should pay more attention to the morphology of red cell in the peripheral blood film.Recommendation of screening tests with high sensitivity and specificity will be improving the diagnosis of HHA.For correct diagnosis,the medical centres should establish red cell membrane protein electropheresis,direct quantitative assay of red cell enzyme,gene diagnosis and prenatal diagnosis of hemoglobinopathy.
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PurposeTo explore the relationship between expression of apoptosis-modulating proteins and chemotherapeutic efficacy in acute leukemia. MethodsImmunocytochemical method was used to detect the expression of Bcl-2、Bcl-XL、Bax and Bak in 36 cases of acute leukemia including previously untreated/ drug-sensitive group and refractory/relapse group. ResultsThe average positive cell rates of Bcl-2 and Bcl-XL in refractory/relapse group were (41.68 ± 14.39) % and (35.96 ± 9.95 ) %, while the rates in previously untreated/drug-sensitive group were (15.64 ± 8.51 )% and (12.91 ± 8.63 )%. Statistical analysis showed the average positive cell rates of Bcl-2 and Bcl-XL in refractory/relapse group were higher than those in previously untreated/drug-sensitive group (P ＜ 0.01 ). There was no significant difference in average positive cell rates of Bax and Bak between refractory/relapse group (25.28 ± 15.49) %, (15.53 ± 10.64) % and previously untreated/drug-sensitive group (21.55 ± 12.58)%, (13.23 ± 8.36)%. The Logistic regression of expression of Bd-2 、Bcl-XL、Bax and Bak to complete remission rate (CR) of 36 cases of acute leukemia showed that Bcl-XL was the most risk factor in reducing the CR.ConclusionsBcl-2 and Bcl-XL might play important roles in multi-drug resistance of acute leukemia and Bcl-XL was more important than Bcl-2.
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Purpose To explore the Relationship between expression of apoptosis-modulating proteins amdmultidrug resistance in K562/VCR cells. Methods Irnmunocytochemical methol and western blot wereused to analyze the expression of apoptosis-modulating proteins (Bcl - 2, Bcl-XL, Bax, Bak ) in multidrugresistant cell line K562/VCR and drugsensitive cell line K562. Results The positive cell rates ofapoptosis-suppressing protein Bcl-2 and Bcl-XL in K562/VCR were (40.0 ± 8.0) % and (60.0 ± 10.0) % .While the rates in K562 were (1.0 ± 0.3) % and (20.0 ± 4.0) %. There was significant difference in thepositive cell rates of Bcl - 2 and Bcl - XL between K562/VCR and K562 ( n = 3, P ＜ 0.05 ). It was alsofound there was no significant difference in expression of Bax between K562/VCR and K562. Furthemore,Bak was not expressed in both K562/VCR and K562 or the expression was very low. Conclusions Wesuggest that Bcl-2 and Bcl-XL play important roles in multidrug resistance in K562/VCR, while Bax and Bakmight not be important.