Tetrahydroxystilbene glucoside protects against sodium azide-induced mitochondrial dysfunction in human neuroblastoma cells / 中草药·英文版

Objective: Mitochondrial dysfunction is evident in the early stage of Alzheimer's disease (AD). Therefore development of drugs that protect mitochondrial function is a promising strategy for AD. The present work was to investigate the effects of 2, 3, 5, 4′-Tetrahydroxystilbene-2-O-β-d-glucosides (TSG) on a mitochondrial dysfunction cell model induced by sodium azide and elucidate the underlying mechanisms. Methods: Mitochondrial membrane potential (MMP) was detected by a fluorescence method. Cellular adenosine triphosphate (ATP) level was measured using a firefly luciferase-based kit. Reactive oxygen species (ROS) was detected using dichlorofluorescin diacetate (DCFH-DA). The expression levels of Bcl-2 and Bax were measured by Western blotting assay. Flow cytometry was utilized to measure apoptosis. Results: Pretreatment of TSG (25–200 μmol/L) for 24 h significantly elevated MMP and ATP content, reduced ROS level and Bax/Bcl-2 ratio, and inhibited apoptosis in SH-SY5Y cells exposed to sodium azide. Conclusion: These results suggest that TSG protects SH-SY5Y cells against sodium azide-induced mitochondrial dysfunction and apoptosis. These findings are helpful to understand the protective effect of TSG on mitochondria, which are involved in the early stage of AD.

Genetic characteristics of human acute lymphoblastic leukemia cell line Molt-4 / 中国实验血液学杂志

This study was aimed to investigate the genetic characteristics of human acute lymphoblastic leukemia cell line Molt-4, and evaluate its application in measuring telomere length by Flow-FISH. Molt-4 cell line was cultured in suspension and subcultured regularly. Eight different passages of Molt-4 cells in exponential stage were selected.The growth curves were drawn by cell counting method, meanwhile calculating the population doubling times of cells,DNA ploidies were determined by flow cytometry,karyotypes were analyzed by G-banding and telomere lengths were measured by Southern blot. The results showed that the population doubling time of Molt-4 cell line was (1.315 ± 0.062) d, DNA ploidy index was (2.085 ± 0.0093) , and the telomere length was (32.05 ± 5.27) kb. There were no significant difference among different passages (P = 0.931,0.888 and 0.935 separately). The karyotypes showed that the chromosome numbers of Molt-4 cell line were from 91 to 99 in different metaphases, and the majority of them were hypertetraploid, and stable and recurrent structural abnormalities of chromosomes could be kept. It is concluded that the stable genetic characteristics and the longer telomere length of Molt-4 cell line makes it be a feasible control cells in measurement of telomere length by Flow-FISH.

Evaluation of neutrophilic CD64 index as a diagnostic marker of bacterial infection in blood diseases / 中国实验血液学杂志

This study was aimed to investigate the value of neutrophilic CD64 index (nCD64 index) as a diagnostic marker of bacterial infection in hematologic diseases. Experimental data of 232 patients with hematologic diseases were analyzed retrospectively. The nCD64 index was detected by flow cytometry and was compared with the levels of erythrocyte sedimentation rate (ESR), C reaction protein (CRP) and fibrinogen respectively. The results showed that the nCD64 index in clinical infection group were significantly higher than that in non-infection group and autoimmune disease group (P < 0.0001 respectively). The nCD64 index in blood culture positive group was also significantly higher than that in blood culture-negative group (P < 0.01). The result of ROC curve analysis showed that the optimal critical values of nCD64 index, ESR, CRP and Fib were 4.96, 21.5 mm/h, 8.56 mg/dl and 4.42 mg/dl, respectively. The sensitivity and specificity of nCD64 index were 0.928 and 0.933, while the sensitivities of ESR, CRP and Fib were 0.725, 0.754 and 0.594, and the specificities of CRP, ESR and Fib were 0.625,0.837 and 0.77, respectively. It is concluded that nCD64 index is possessed of much higher in sensitivity and specificity, compared with ESR, CRP and Fib in diagnosis of bacterial infection of hematologic diseases. nCD64 index can be used as an effective diagnostic marker for bacterial infection of hematologic diseases.

Application of interphase FISH on cell smears in detection of hematological diseases / 中国实验血液学杂志

The study was aimed to investigate the application value of interphase fluorescence in situ hybridization (FISH) on cell smears in hematological diseases. Both interphase FISH on peripheral blood smears and bone marrow smears treated by methanol/acetic acid, and routine interphase FISH of bone marrow cells dropped on slides were done at the same time, in order to detect Ph chromosome by BCR/ABL dual color, dual fusion probe in 20 patients with chronic myelogenous leukemia or acute lymphoblastic leukemia which had been proven to display Ph chromosome positive. The results indicated that as compared with routine interphase FISH, the interphase FISH on cell smears could also offer reliable result. It is concluded that interphase FISH on cell smears is a kind of reliable and time-saving technique, which is also suitable for retrospective research and worthy to further apply in clinic.

Multiparameter flow cytometric evaluation of bone marrow involvement in B cell non-Hodgkin's lymphoma / 中国实验血液学杂志

The objective of study was to evaluate the clinical values of multiparameter flow cytometry (MPFC) and cytomorphology of bone marrow aspiration(BMA) in detecting bone marrow involvement in patients with B cell Non-Hodgkin's lymphoma (B-NHL). 96 bone marrow samples from the patients with B-NHL were measured by MPFC using CD45/SSC and CD20/SSC gating strategy combined with anti-kappa and anti-lamda monoclonal antibodies, and then compared with results acquired by cytomorphologic analysis of BMA. The results showed that the bone marrow involvement was confirmed by MPFC in 38 cases (39.6%), while it was detected by cytomorphologic analysis of BMA only in 12 cases (12.5%). There was a significant difference between the two methods (p<0.05). 12 positive cases detected by cytomorphologic analysis of BMA were also positive by MPFC. There was no difference of 3-year overall survival rate between negative and positive cases detected by MPFC, but their 4-year overall survival rate was 73.18+/-6.65% and 44.13%+/-19.55% respectively (p<0.05). It is concluded that the MPFC is a more sensitive method for detecting bone marrow involvement in patients with B-NHL than cytomorphologic analysis of BMA. The 4-year overall survival rate of the patients without bone marrow involvement was significant higher than those of patients with bone marrow involvement. Bone marrow involvement in B-NHL detected by MPFC can be useful for clinical evaluation and prognosis prediction.

Abnormality of immunophenotyping in patients with myelodysplastic syndrome / 中国实验血液学杂志

The study was aimed to investigate the abnormality of immunophenotypes in patients with myelodysplastic syndrome (MDS) and its role in the identification of MDS. The cell immunophenotypes of 136 patients with hypocytosis accompanied by abnormal hematopoiesis of bone marrow were detected by flow cytometry, the detected results were evaluated by flow cytometric scoring system (FCSS), and the sensitivity and specificity of positive results were determined by FCSS also. The correlation of results detected by FCSS to traditional diagnosis method was analysed. The results indicated that 111 out of 136 cases were diagnosed as MDS, and 25 were diagnosed as non-MDS. Among 111 MDS cases, 85 cases were FCSS positive, 18 cases were FCSS intermediate and 8 cases were FCSS negative, whereas in 25 non-MDS cases 24 cases were FCSS negative, 1 case was FCSS intermediate and no case was FCSS positive. The sensitivity of FCSS in identification of MDS was 76.6%, and the specificity of FCSS was 100%. There was a good correlation of FCSS to traditional method (R = 0.613, p = 0.000). It is concluded that the various abnormalities of immunophenotyping are found in patients with MDS, in which the main immunophenotype abnormality and the abnormality involving two cell lineages are key points to distinguish MDS from non-MDS.

Prognosticating relapse risk based on multiparameter flow cytometric assessment of minimal residual disease in patients with acute myeloid leukemia / 中国实验血液学杂志

The objective of this study was to investigate the prognosticating value of multiparameter flow cytometry in detection of minimal residual disease (MRD) and relapse risk of patients with acute myeloid leukemia (AML). Multiparameter flow cytometry (MPFC) analysis was used to detect the leukemia-associated aberrant immunophenotype (LAIP) of the pretreated patients with AML and to assess the levels of MRD after remission induction (Post-Ind MRD) and consolidation therapy (Post-Cons MRD). The results showed that the definite LAIP could be detected in 94.3% of the patients (115/122) with AML (except APL). Among 115 cases only one LAIP was identified in 15 cases (13.0%), but two or more LAIP were identified in other 100 cases (87.0%). The most frequent LAIP identified was cross-lineage antigen expression (40.9%). The percentages of asynchronous antigen expression, antigen over-expression and antigen lack expression were 20.9%, 27.0%and 34.8% respectively. MRD frequency was monitored in 41 AML patients with CR after remission induction chemotherapy and 2 or more cycles of consolidation chemotherapy. 24 patients were Post-Ind MRD(+) and 17 patients were Post-Ind MRD(-). The percentages of relapse in cases of Post-Ind MRD(+) and Post-Ind MRD(-) were 75.0% (18/24) and 29.4% (5/17) respectively after consolidation chemotherapy. The relapse free survival (RFS) times of the patients with Post-Ind MRD(+) and Post-Ind MRD(-) were 49.06 +/- 6.53 months and 11.92 +/- 1.64 months (p < 0.0001) respectively. 18 patients were Post-Cons MRD(+) and 23 patients were Post-Cons MRD(-). The percentages of relapse in cases of Post-Cons MRD(+) and Post-Cons MRD(-) patients were 100% (18/18) and 21.7% (5/23) respectively after consolidation chemotherapy. The RFS times of the patients with Post-Cons MRD(+) and Post-Cons MRD(-) were 41.74 +/- 5.52 months and 10.06 +/- 1.72 months (p < 0.0001) respectively. It is concluded that the levels of post-Ind MRD and post-Cons MRD identified in the patients with AML was highly associated with their RFS. The detection of MRD by MPFC provides prognostic information in AML patients.

Diagnostic significance of immunophenotyping of bone marrow cells in myelodysplastic syndrome without an increase of marrow blasts / 中国实验血液学杂志

This study was aimed to investigate the characteristics of immunophenotypes in the patients with myelodysplastic syndrome (MDS) without an increase of marrow blasts, and to confirm their diagnostic significance. Marrow cells from 222 patients with pancytopenia, dysplastic changes in one or more hematopoietic lineages and blast cells less than 5% were analyzed by multiparametric flow cytometry(FCM). The abnormal immunophenotypes were evaluated in asynchronous antigen expression (CD34 or CD117 in mature granulocytes or mature monocytes, HLA-DR in mature granulocytes), in cross-lineage antigen expression (CD7 or CD56 in granulocytes or monocytes), in aberrant light-scatter (CD45/SSC in mature granulocyte or monocyte) and in abnormal expression of differentiation antigen (CD13/CD16 pattern in granulocytes and HLA-DR under-expression in monocytes). The sensitivity and specificity of abnormal immunophenotypes were determined on diagnosis. Among 222 cases, 127 cases were diagnosed as MDS by traditional diagnostic method and 95 cases were non-MDS (drug-related neutropenia, autoimmune cytopenia and idiopathic thrombocytopenia). In mature granulocyte gate, the sensitivity of asynchronous, cross-lineage antigen expression, aberrant light-scatter of CD45/SSC and abnormal expression of differentiation antigen were 31.5%, 30.7%, 49.6% and 60.6% respectively, and the specificity were 100%, 100%, 88.4% and 52.6% respectively. In monocyte gate, the sensitivity of asynchronous, cross-lineage antigen expression, aberrant light-scatter of CD45/SSC and abnormal expression of differentiation antigen were 2.3%, 11%, 37% and 12.6% respectively. The specificity was 100% in all of them. Among 8 above mentioned items, sensitivity of more than 2 abnormalities was 77.9%, and specificity was 95.8%. The positive predictive value was 96.1%. It is concluded that the abnormal expression of asynchronous, cross-lineage antigen expression, aberrant light-scatter of CD45/SSC have a high specificity and a low sensitivity for diagnosis of MDS. The abnormal expressions of differentiation antigens have a high sensitivity and a low specificity; however, the detection of multiple expression abnormalities possesses the high sensitivity and specificity for diagnosis of MDS.

Megakaryocytic dysplasia and leukemia associated phenotype in elderly patients with acute myeloid leukemia / 中国实验血液学杂志

This study was purposed to investigate the megakaryocytic dysplasia and leukemia-associated phenotypes (LAP) of acute myeloid leukemia (AML) in the elderly. The megakaryocytic dysplasia, lineage infidelity, asynchronous antigen expression, total WBC count, and karyotypes were observed in the 147 none M(3)-AML patients. Logistic regression were used to analyzed the difference between the elderly (age > or = 60) and the control. The results showed that out of the total 147 patients (66 elderly patients, and 81 younger patients) 124 patients accepted induction chemotherapy, in which 70 cases achieved complete remission (elderly 18, younger 52, p = 0.008); megakaryocytic dysplasia was found in 32 patients (21.8%); CD33 and CD19/CD7 (lineage infidelity) was co-expressed in 55 patients (37.4%), CD34 and CD11b (asynchronous antigen expression) was co-expressed in 65 patients (44.2%); white blood cell count > 25 x 10(9)/L was found in 52 patients (35.4%). By the Logistic regression, compared with the control, in the elderly patients there was difference in the megakaryocytic dysplasia, and the co-expression of CD33/CD19/CD7 and CD34/CD11b (OR = 4.315, 2.761, 0.397; p = 0.001, 0.006, 0.020), but there was no difference in the total WBC count and karyotypes (OR = 0.802, 1.096; p = 0.646, 0.813). It is concluded that the incidence of megakaryocytic dysplasia, such as lineage infidelity, and asynchronous antigen expression, in elderly patients is higher than that in younger patients.

Aberrant immunophenotypes in 126 patients with acute leukemia / 中国实验血液学杂志

The existence of leukemia aberrant immunophenotypes (LAIP) has been suggested to be a valuable tool for the detection of minimal residual disease (MRD), as they could distinguish leukemic cells from normal hematopoietic progenitors. This study was purposed to analyze the characteristics of LAIP in acute leukemia and further explore the proportion of different types of LAIP in acute leukemia patients. Flow cytometry (FCM) with four color and CD45/SSC gating were used to detect the antigen expression in samples of bone marrow from 126 patients with acute leukemia. The results showed that definite LAIP could be detected in about 76% patients. The LAIP could be divided into four groups as cross-lineage antigen expression, asynchronous antigen expression, antigen overexpression and antigen lack expression. The percentages of these LAIPs were 39%, 46%, 21% and 29% respectively. About 11% out of analyzed cases showed the existence of only one aberrant phenotype while two or more of aberrant phenotypes could be detected in majority cases. It is concluded that the LAIP with four subgroups can be detected in the majority of patients with acute leukemia and immunophenotyping based on LAIP is applicable for the detection of MRD.

Clinical outcomes after autologous haematopoietic stem cell transplantation in patients with progressive multiple sclerosis / 中华医学杂志(英文版)

<p><b>BACKGROUND</b>Multiple sclerosis (MS) is a continuously disabling disease and it is unresponsive to high dose steroid and immunomodulation with disease progression. The autologous haematopoietic stem cell transplantation (ASCT) has been introduced in the treatment of refractory forms of multiple sclerosis. In this study, the clinical outcomes followed by ASCT were evaluated for patients with progressive MS.</p><p><b>METHODS</b>Twenty-two patients with secondary progressive MS were treated with ASCT. Peripheral blood stem cells were obtained by leukapheresis after mobilization with granulocyte colony stimulating factor. Etoposide, melphalan, carmustin and cytosine arabinoside were administered as conditioning regimen. Outcomes were evaluated by the expanded disability status scale and progression free survival. No maintenance treatment was administered during a median follow-up of 39 months (range, 6 to 59 months).</p><p><b>RESULTS</b>No death occurred following the treatment. The overall confirmed progression free survival rate was 77% up to 59 months after transplantation which was significantly higher compared with pre-transplantation (P = 0.000). Thirteen patients (59%) had remarkable improvement in neurological manifestations, four (18%) stabilized their disability status and five (23%) showed clinical recurrence of active symptoms.</p><p><b>CONCLUSIONS</b>ASCT as a therapy is safe and available. It can improve or stabilize neurological manifestations in most patients with progressive MS following failure of conventional therapy.</p>

Expansion ex vivo of human bone marrow mesenchymal stem cells and cord blood CD34+ cells / 中国实验血液学杂志

The aim was to study the roles that the bone marrow mesenchymal stem cells (MSC) and cytokines play in cord blood CD34(+) cell expansion ex vivo and the influence of culture ex vivo on expression of the adhesive molecule of CD44. CD34(+) cells sorted from cord blood cells had been cultured in each well of 24 well culture plates containing culture medium supplemented with mesenchymal stem cells layer or/and cytokines for a week, and then all kinds of indexes of different groups were compared. The results showed that as for cord blood cell expansion, there was no significant difference between the groups with cytokines SDF-1alpha + SCF + TPO + FL and SCF + TPO + FL no matter if MSC layer existed or not. The groups with MSC layer and cytokines were superior to the corresponding groups without MSC layer. In addition, the expression of the adhesion molecule CD44 had no distinct change after culture. It is concluded that SDF-1alpha has no distinct influence on the effect of cytokines SCF + TPO + FL on cord blood cell expansion ex vivo. MSC enhance the effect of cytokines on cord blood cell expansion ex vivo. Such expansion ex vivo may not influence the expression of the adhesive molecule CD44 on cord blood cells.

The effect of granulocyte colony stimulating factor on T-cell subsets in peripheral blood and its correlation with the mobilization effect of CD34+ cells / 中国实验血液学杂志

The objective was to observe the effect of G-CSF as a mobilizer of hematopoitic stem cells on the absolute counts of T-cell subsets in peripheral blood and their relevance with the mobilized CD34(+) cells. The examples of peripheral blood from 26 patients performed of autologous stem cell transplantation were taken before and after mobilization by G-CSF. Flow cytometry was used for detecting CD3(+), CD3(+)CD4(+), CD3(+)CD8(+), CD3(+)CD4(+)CD8(+) and CD3(+)CD4(-)CD8(-) cells. Concurrently, their correlations with mobilized CD34(+) cells in peripheral blood were compared. The results showed that after the mobilization by G-CSF, the amounts of CD3(+), CD3(+)CD4(+), CD3(+)CD4(+)CD8(+) and CD3(+)CD4(-)CD8(-) cells in peripheral blood increased by 2.23, 2.62, 2.99 and 10.96 fold respectively, but that of CD3(+)CD4(-)CD8(+) cells was nearly no changed (P = 0.243). The correlation coefficient of CD3(+)CD4(-)CD8(-) cells and mobilized CD34(+) cells was 0.796, (P = 0.000) and no correlation with other T-cell subsets. It was concluded that when CD34(+) cells were mobilized by G-CSF from bone marrow to peripheral blood, the absolute counts of the peripheral T-cell subsets got changed. The increase of CD3(+)CD4(-)CD8(-) cells had correlated with mobilization effect of CD34(+) cells into peripheral blood.