Quantitative Analysis on Immunophenotype of CD34 / 中国实验血液学杂志

OBJECTIVE@#To investigate the quantitative expression of immunophenotype of CD34@*METHODS@#Multi-parameter flow cytometry (FCM) was used to detect the proportion and mean fluorescence intensity (MFI) of each antigen of bone marrow CD34@*RESULTS@#Bone marrow blast cell proportion (P<0.01), RBC level (P<0.01), and Hb level (P<0.05) of high-risk MDS patients were higher, while EPO level (P<0.05) was lower than those of low-risk patients. The proportion of CD34@*CONCLUSION@#The immunophenotype of CD34

Effect of Signal Transduction Pathway Gene Mutations on the One- course Induced Remission Rate and Analysis of Clinical Characteristics in Patients with CBF-AML / 中国实验血液学杂志

OBJECTIVE@#To investigate the effect of other gene mutations outside the fusion gene on the first complete remission (CR) induced by one course of induction chemotherapy in patients with core binding factor-associated acute myeloid leukemia (CBF-AML).@*METHODS@#DNA was extracted from bone marrow or peripheral blood samples of newly diagnosed CBF-AML patients admitted to the Hematology Department of the Second Hospital of Shanxi Medical University from January 2015 to January 2019. Next-generation sequencing was used for detection of 34 kinds of hematologic malignancy-related gene mutations in patients with CBF-AML, the effect of related gene mutations on the first complete remission (CR) rate in one course of induction chemotherapy was analyzed by combineation with clinical characteristics.@*RESULTS@#34 kinds of genes in bone marrow or peripheral blood of 43 patients were detected by high throughput sequencing and the gene mutations were detected in 16 out of 34 genes. The mutation rate of KIT gene was the highest (48.8%), followed by NRAS (16.3%), ASXL1 (16.3%), TET2 (11.6%), CSF3R (9.3%), FLT3 (9.3%), KRAS (7.0%). The detection rates of mutations in different functional genes were as follows: genes related with signal transduction pathway (KIT, FLT3, CSF3R, KRAS, NRAS, JAK2, CALR, SH2B3, CBL) had the highest mutation frequency (72.1% (31/43); epigenetic modification gene mutation frequency was 30.2% (13/43), including ASXL1, TET2, BCOR); transcriptional regulation gene mutation frequency was 7.0% (3/43), including ETV6, RUNX1, GATA2). Splicing factor related gene mutation frequency was 2.3% (1/43), including ZRSR2). The CR rate was 74.4% after one course of induction chemotherapy. At first diagnosis, patients with low expression of WT1 (the median value of WT1 was 788.9) were more likely to get CR (P=0.032) and the RFS of patients who got CR after one course of induction chemotherapy was significantly longer than that of patients without CR [7.6 (2.2-44.1) versus 5.8 (1-19.4), (P=0.048)]. The rate of CR in the signal transduction pathway gene mutation group was significantly lower than that in non-mutation group (64.5% vs 100%) (P=0.045), while the level of serum hydroxybutyrate dehydrogenase (HBDH) was significantly higher than that in non-mutation group [(418 (154-2702) vs 246 (110-1068)] (P=0.032). There was no difference in CD56 expression between the two groups (P=0.053), which was limited to the difference between (≥20%) expression and non-expression. (P=0.048).@*CONCLUSION@#CBF-AML patients with signal transduction pathway gene mutation are often accompanied by high HBDH level and CD56 expression, moreover, the remission rate induced by one course of treatment is low.

Expression of CD25 in acute B cell lymphoblastic leukemia and its clinical significance / 中国实验血液学杂志

This study was purposed to investigate the relation of CD25 with the acute B cell lymphoblastic leukemia (B-ALL) and its clinical significance. A totol of 88 newly diagnosed B-ALL patients were enrolled in this study. The immunophenotype of leukemic myeloblasts were detected by flow cytometry, including interleukin 2 receptor α chain (CD25), β chain (CD122), γ chain (CD132), CD19, CD20, CD10, CD34, CDIgM, CD79a, CD22 and CDTDT. The expression of BCR/ABL fusion gene was detected by qualitative PCR. The expression of IL2RA (CD25 gene) was detected by real-time qualitative RT-PCR. The results showed that there was no significant statistical difference in WBC count, Hb level, PLT count, marrow blast rate, peripheral blast rate, hepato-lienal infiltration, lymph node infiltration, levels of CD10, CD20, CD22, CD34, CD79a, CDTDT, CDIgM expression between B-ALL patients with CD25(+) and B-ALL patients with CD25(-), while the CD19 expression level in B-ALL patients with CD25(+) was higher than that in B-ALL patients with CD25(-). Out of 88 B-ALL patients, 21 patients showed BCR/ABL(+)(21/88) and their CD25(+) expression level was 66.7% (14/21); 67 patients showed BCR/ABL(-) and their CD25(+) expression level was 4.5% (3/67), there was statistical difference between these two groups (P < 0.05), but the expression level of IL2RA mRNA was not statistical different between CD25(+) and CD25(-) groups (P > 0.05). Among 21 BCR/ABL(+) B-ALL patients the remission rate and relapsed rate were not statistical different between CD25(+) an CD25(-) groups.In BCR/ABL(+) B-ALL patients 8 patients relapsed, the relapsed rate was 38.1% (8/21). In BCR/ABL(-) B-ALL patients 9 patients relapsed, the relapse rate was 13.4% (9/67), there was statistical difference between BCR/ABL(+) and BCR/ABL(-) two groups (P < 0.05). In BCR/ABL(+) group the RFS (relapse free survival) was 21 months, in BCR/ABL(+) CD25(+) patients the RFS was 15 months, while in BCR/ABL(+) CD25(-) patients the RFS was 21 months, in BCR/ABL(-) CD25(-) patients, the RFS was 24 months. It is concluded that the CD25 expresses at high level in B-ALL patients with BCR/ABL(+), which may serve as a predictive marker for the presence of BCR/ABL fusion gene, and relates with relapse, CD25(+) may serve as a adjuvant indicator for poor prognosis.

Establishment of diagnostic model to monitor minimal residual disease of acute promyelocytic leukemia by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry / 中华血液学杂志

<p><b>OBJECTIVE</b>To screen the potential protein biomarkers in minimal residual disease (MRD) of the acute promyelocytic leukemia (APL) by comparison of differentially expressed serum protein between APL patients at diagnosis and after complete remission (CR) and healthy controls, and to establish and verify a diagnostic model.</p><p><b>METHODS</b>Serum proteins from 36 cases of primary APL, 29 cases of APL during complete remission and 32 healthy controls were purified by magnetic beads and then analyzed by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS). The spectra were analyzed statistically using FlexAnalysis(TM) and ClinProt(TM) software.</p><p><b>RESULTS</b>Two prediction model of primary APL/healthy control, primary APL/APL CR were developed. Thirty four statistically significant peptide peaks were obtained with the m/z value ranging from 1000 to 10 000 (P < 0.001) in primary APL/healthy control model. Seven statistically significant peptide peaks were obtained in primary APL/APL CR model (P < 0.001). Comparison of the protein profiles between the two models, three peptides with m/z 4642, 7764 and 9289 were considered as the protein biomarker of APL MRD. A diagnostic pattern for APL CR using m/z 4642 and 9289 was established. Blind validation yielded correct classification of 6 out of 8 cases.</p><p><b>CONCLUSIONS</b>The MALDI-TOF MS analysis of APL patients serum protein can be used as a promising dynamic method for MRD detection and the two peptides with m/z 4642 and 9289 may be better biomarkers.</p>

Effect of Nrf2 and TrxR on proliferation of chronic myeloid leukemia cell and its mechanism / 中华血液学杂志

<p><b>OBJECTIVE</b>To explore the effect of nuclear factor erythroid-2 related factor 2 (Nrf2) and thioredoxin reductase (TrxR) gene on proliferation of chronic myeloid leukemia (CML) line cells and its mechanism.</p><p><b>METHODS</b>Four interfering sequences of Nrf2 and one negative control sequence were designed and synthesised based on the principle of target sequence of siRNA, then constructed lentivirus vectors, which were transfected into K562 cell lines. The transfection effect was observed by laser scanning confocal microscope (LSCM) and flow cytometer (FCM); The depressing effect of siRNA was analyzed by real-time PCR. The cell proliferation inhibiting rate was measured with CCK-8 assay, the apoptotic rate by Annexin V-PE/PI with FCM and the apoptotic morphology of cells by LSCM.</p><p><b>RESULTS</b>The transfection efficiency of lentivirus was 65%. One cell line K562-C3 which significantly inhibited Nrf2 mRNA was obtained by real-time PCR, Nrf2 relative quantitation (RQ) expressions were 1.003±0.093 and 0.344±0.032 in the control group and K562-C3 respectively; TrxR expression also decreased with RQ as 1.090±0.549 and 0.395±0.029 respectively. The cellular proliferation inhibition rates of K562-C3 were (4.74±0.39)%, (6.13±1.78)% and (25.36±3.77)%, respectively at 24, 48 and 72 h. The apoptotic rate induced by K562-C3 (29.9%) at 72 hours was obviously higher than in the control group (7.9%). The Annexin V-PE positive K562-C3 cells presented the following apoptotic characteristics, such as karyopyknosis, nuclear fragmentation and apoptotic bodies observed by LSCM.</p><p><b>CONCLUSION</b>Nrf2 specific siRNA could repress its expression at the cellular level and down-regulate the expression of its downstream antioxidant enzyme, such as TrxR, which lead to increased apoptotic rate and decreased cell proliferation.</p>

Expressions of Th17 cells and interleukin 17 in patients with primary immune thrombocytopenia and their clinical significance / 中国实验血液学杂志

The aim of this study was to detect the rate of T-helper (Th)17 cells and interleukin (IL)-17 level in peripheral blood of patients with primary immune thrombocytopenia (ITP) and to explore their clinical significance. The proportion of Th17 cells from 48 patients with ITP and 28 healthy controls was detected by flow cytometry, and the IL-17 level was evaluated by enzyme-linked immunosorbent assay (ELISA). The results showed that the percentage of Th17 cells in ITP group was (1.40 ± 1.35)%, which was significantly higher than that in healthy control group (P < 0.05), but in the glucocorticoid hormone-treated group it was significantly lower than that in treated group without glucocorticoid hormone(P < 0.05). The level of IL-17 expressed by Th17 cells in ITP patients was (19.624 ± 5.187) pg/ml, which was higher than that in the healthy control group (P < 0.05), it was lower in the glucocorticoid hormone treated group than that in treated group without glucocorticoid hormone, but there was no statistically significant difference between the glucocorticoid treated and treated group without glucocorticoid hormone (P > 0.05). It is concluded that the Th17 cells may involve in the pathogenesis of ITP, and the glucocorticoid hormone probably plays a therapeutic role through inhibiting Th17 cells.