Preliminary exploration of the metabolic profile and metabolic pathways in newly diagnosed multiple myeloma / 中华肿瘤杂志

Objective: To explore the metabolite profile and metabolic pathways of newly diagnosed multiple myeloma (MM). Methods: Gas chromatography-mass spectrometry (GC-MS) was employed for the high-throughput detection and identification of serum samples from 55 patients with MM and 37 healthy controls matched for age and sex from 2016 to 2017 collected at the First Affiliated Hospital of Soochow University. The relative standard deviation (RSD) of quality control (QC) samples was employed to validate the reproducibility of GC-MS approach. The differential metabolites between patients with MM and healthy controls were detected by partial least squares discrimination analysis (PLS-DA), and t-test with false discovery rate (FDR) correction. Metabolomics pathway analysis (MetPA) was employed to construct metabolic pathways. Results: There were 55 MM patients, including 34 males and 21 females. The median age was 60 years old (42-73 years old). There were 30 cases of IgG type, 9 cases of IgA type, 1 case of IgM type, 2 cases of non-secreted type, 1 case of double clone type and 12 cases of light chain type, including 3 cases of kappa light chain type and 9 cases of lambda light chain type. The result of QC sample test showed that the proportion of compounds with the RSD of the relative content of metabolites < 15% was 70.21% obtained by the reproducibility of GC-MS experimental data, which implied that the experimental data were reliable. A total of 17 metabolites were screened differently with the healthy control group, including myristic acid, hydroxyproline, cysteine, palmitic acid, L-leucine, stearic acid, methionine, phenylalanine, glycerin, serine, isoleucine, tyrosine, valine, citric acid, inositol, threonine, and oxalic acid (VIP>1, P<0.05). Metabolic pathway analysis suggested that metabolic disorders in MM patients comprised mainly phenylalanine metabolism, glyoxylic acid and dicarboxylic acid metabolism, phosphoinositide metabolism, cysteine and methionine metabolism, glycerolipid metabolism, glycine, serine, and threonine metabolism. Conclusion: Compared with normal people, patients with newly diagnosed MM have obvious differences in metabolic profiles and metabolic pathways.

The Expression of WTAP Gene in Acute Myeloid Leukemia and Its Clinical Significance / 中国实验血液学杂志

OBJECTIVE@#To investigate the expression of WTAP gene in acute myeloid leukemia (AML) and its clinical significance.@*METHODS@#74 acute myeloid leukemia patients with non-M3 type and 19 normal donors were selected, and real-time quantitative polymerase chain reaction was used to detect the mRNA expression level of WTAP gene in their bone marrow cells. The relationship between the mRNA expression level of WTAP gene and the clinical characteristics was analyzed.@*RESULTS@#The relative mRNA expression of WTAP gene in the non-M3 AML group was significantly higher than that in the healthy control group, and the difference showed statistically significant (P0.05) according to the classification of FAB. The mRNA expression level of WTAP gene in FLT3-ITD mutated AML patients was higher than that in FLT3-ITD unmutated group (P=0.016), and the mRNA expression level of WTAP gene in AML patients with CEBPα mutation was lower than that in CEBPα unmutated group (P=0.016). The expression level of WTAP mRNA was positively correlated with WT1 expression (r=0.6866, P0.05). The expression level of WTAP mRNA showed no obvious effect on the complete remission of patients after first treatment. The different expression level of WTAP gene at initial diagnosis showed also no effect on the overall survival time of patients.@*CONCLUSION@#The expression level of WTAP gene is increasing in new diagnosed non-M3 acute myeloid leukemia. There is a positive correlation between the expression level of WTAP gene and the expression level of WT1 fusion gene. WTAP mRNA always shows higher expression in patients with FLT3-ITD mutation than that in patients without FLT3-ITD mutation, and shows lower expression in patients with CEBPα mutation than that in unmutated group.

Application of multiplex PCR assay to study early multiple herpesviruses infection during HSCT / 中华血液学杂志

Objective: To investigate herpesvirus infection in early stage of hematopoietic stem cell transplantation (HSCT) by multiplex polymerase chain reaction (PCR), and to explore the association between multiple herpesviruses infection and clinical characteristics in HSCT patients and its impact on post-transplant complications and prognosis. Methods: A total of 734 peripheral blood samples were collected from 90 patients undergoing HSCT in the Department of Hematology, the First Affiliated Hospital of Soochow University between February 2017 and August 2017. The peripheral blood specimens were obtained before and within 90 days after transplantation at different time points. Lab-Aid824 Nucleic Acid Extraction Mini Reagent was used to extract DNA and multiplex PCR assay was used to simultaneously detect 8 kinds of human herpesviruses from genomic DNA. The incidence of various herpesvirus infections, its correlation with clinical features and effects on post-transplant complications and prognosis were analyzed. Results: The median follow-up time was 192 (range: 35-308) days. Among the 90 patients before transplantation, the incidence of herpes virus infection was 35.6% (32/90), including 12.2% (11/90) with one herpes virus infection and 23.3% (21/90) with multiple viruses infection. The incidence of herpes virus infection after transplantation was 77.8% (70/90), including 20.0% (18/90) with one herpes virus infection and 57.8% (52/90) with multiple herpes virus infection. Among the 52 patients with multiple herpes viruses infection, 30 (57.7%) patients were infected by 2 kinds of viruses, 18 (34.6%) patients by 3 kinds of viruses and 4 (7.7%) patients by 4 kinds of viruses. There was a correlation between HHV-6 and HHV-7 herpesvirus infection (OR=13.880, Q=0.026). EBV infection was related to HHV-7 infection (OR=0.093, Q=0.044). The age of patients was correlated with the incidence of HHV-1 infection before transplantation. There were 24 patients in our study experienced clinical symptoms associated with viral infection. The main manifestations were hemorrhagic cystitis (HC), interstitial pneumonia, enteritis, viral encephalitis and fever of unknown origin. EBV infection was related to HLA incompatibility and the inconsistent of the ABO blood group and grade Ⅱ-Ⅳ aGVHD after transplantation. HLA incompatibility and the unrelated donor and grade Ⅱ-Ⅳ aGVHD were related to multiple viruses infection. Conclusion: Multiple herpesviruses were common in patients undergoing HSCT, which were closely related to HLA mismatch, unrelated donor and grade Ⅱ-Ⅳ aGVHD.

Current situation and prospect of resource evaluation and sustainable utilization on Bletilla / 中国中药杂志

The plants of Bletilla are one of the groups in Orchidaceae with the highest economic value. As the traditional Chinese medicinal material, Bletillae Rhizoma exhibits excellent efficacy in hemostatic, antibiosis, detumescent, anticancer activities and regenerating tissue to heal wound, which has great development potential. However, Bletillae Rhizoma is mainly collected from wild resources. At present, the quantity of wild resources of Bletilla plants has sharply decreased and is far from meeting the needs. Resource appraisal and breeding and cultivation of excellent germplasms of Bletilla plants are important for scientific utilization of the resources of the genus. This paper reviewed the following researches of Chinese Bletilla resources: species and distribution, genetic diversity, active ingredient evaluation, breeding, as well as seeding production and cultivation techniques. Suggestions were also provided in further researches on the resources evaluation, sustainable development and efficient utilization of Chinese Bletilla plants.

Amplification Ex Vivo and Cytocidal Activity of Leukemia Tumor-Associated Antigen-Specific Cytotoxic T Lymphocytes / 中国实验血液学杂志

<p><b>OBJECTIVE</b>To explore the feasibility of amplifying the leukemia tumor associated antigens-specific cytotoxic T lymphocytes (TAA-CTL) ex vivo and to evaluate the cytotoxicity of TAA-CTL.</p><p><b>METHODS</b>The peripheral blood mononuclear cells were enriched by density gradient centrifugtion; TAA-CTL were generated by stimulation of PBMNC with peptide-pulsed DC at an effector-to-target ratio of 10:1; immunophenotype of TAA-CTL was analyzed by flow cytometry; cytotoxicity assay was used to evaluate the cytotoxic activity of TAA-CTL against peptide-pulsed autologous target cells (PHA-Blasts).</p><p><b>RESULTS</b>TAA-CTL expanded from volunteer showed a mean expansion of 3.81±1.61, the phenotyping of the TAA-CTL was predominantly CD3+ (97.22±0.71)% with varying content of CD4+ (41.47±27.08)% and CD8+ (56.40±11.15)% T cells, it also contained few nature killer cells (0.50±0.31)% and rare residual B cells (0.14±0.20)%; the subpopulations of TAA-CTL and CTL were not statisticaly significantly different in the proportion (P>0.05); the detection of intracellular cytokines after stimulation with peptide showed that the secretion rates of IFN-γ and TNF-α in CD8+ TAA-CTL were (27.67±2.21)% and (34.2±0.71)%, while the secretion rates were (21.6±2.55)% and (9.97±3.44)% in CD4+ TAA-CTL. Compared with the CD8+ TAA-CTL group, the secretion rates of IFN-γ and TNF-α were (1.36±0.04)% and (5.58±0.03)% in CD8+ CTL, the rates of IFN-γ and TNF-α were (0.91±0.06)% and (1.60±0.07)% in CD4+ CTL. The secretions of IFN-γ and TNF-α in CTL were both significantly lower than those in TAA-CTL (P<0.01); the specific killing efficiency of the TAA-CTL against TAA-pulesd target cells were (77.00±1.00)%, (67.40±3.60)%, (60.55±2.45)% and (26.85±5.25)%, when the effecto-target ratios were 40:1, 20:1, 10:1 and 5:1, and there was negligible lysis of TAA-CTL for PHA-blast (P<0.01).</p><p><b>CONCLUSION</b>TAA-CTL can be successfully induced and generated ex vivo from the healthy volunteer peripheral blood, and the TAA-CTL possess a specific killing activity.</p>

Extracellular HMGB1 promotes the migration of cord Blood CD34⁺ cells via SDF-1/CXCR-4 axis / 中国实验血液学杂志

This study was aimed to investigate the effect of high mobility group box1(HMGB1) and/or stromal cell derived factor-1(SDF-1) on the migration of cord blood CD34⁺ cells, and to explore whether HMGB1 promotes cord blood CD34⁺ cell migration via SDF-1/CXCR4 axis. Cord blood mononuclear cells were isolated by Ficoll-Paque density centrifugation, CD34⁺ cells were collected by a positive immunoselection procedure (CD34 MicroBeads) according to the manufacturer's instructions, the purity of the isolated CD34⁺ cells was detected by flow cytometry. In vitro chemotaxis assays were performed using Transwell cell chambers to detect cells migration. 1 × 10⁵ cells/well cord blood CD34⁺ cells were added into the upper chambers. Different concentrations of HMGB1 and/or SDF-1 (0, 10, 25, 50, 100, 200 ng/ml) were used to detect the optimal concentrations of HMGB1 and/or SDF-1 for inducing migration of cord blood CD34⁺ cells. Freshly isolated cord blood CD34⁺ cells express CXCR4 (SDF-1 receptor), and HMGB1 receptor TLR-2,TLR-4 and RAGE. To explore which receptors were required for the synergy of HGMB1 and/or SDF-1 on cells migration, the anti-SDF-1, anti-CXCR4 and anti-RAGE antibodies were used to detect the effect of HGMB1 alone or with SDF-1 on cord blood CD34⁺ cells migration. The results showed that the purity of CD34⁺ cells isolated from cord blood mononuclear cells by magnetic cell sorting was 97.40 ± 1.26%, the 25 ng/ml SDF-1 did not induce migration of cord blood CD34⁺ cells, whereas the optimal migration was observed at 100 ng/ml. HMGB1 alone did not induce migration up to 100 ng/ml. The dose test found that the the best synergistic concentrations for cells migration were 100 ng/ml HMGB1 combined with 50 ng/ml SDF-1. The blocking test showed that both the anti-SDF-1 (4 µg/ml) and anti-CXCR4 (5 µg/ml) antibodies could block cell migration induced by HMGB1 or combined with SDF-1, but the cord blood CD34⁺ cells in the presence of anti-RAGE, anti-TLR-2 and anti-TLR-4 antibodies did not modify the response to SDF-1 in the presence of HMGB1. It is concluded that both HMGB1 and SDF-1 can induce cord blood CD34⁺ cells migration, HMGB1 enhances SDF-1-induced migration exclusively via CXCR4 and in a RAGE and TLR receptors-independent manner, the exact mechanism needs to be further explored.

Immunophenotyping characteristics of AML and their correlation with the curative effects / 中国实验血液学杂志

This study was aimed to explore the immunophenotyping characteristics of acute myeloid leukemia (AML) and their correlation with the curative efficacy. The bone marrow or blood samples were collected from 516 patients with newly diagnosed AML, and their immunophenotypes were analyzed by flow cytometry. The results showed that (1) In 516 cases, the ratios of myeloid antigen expression were higher, as follows: MPO 95.0%, CD33 93.0%, CD13 88.8%, CD117 69.4%; and the expressions of CD14, CD15, CD64 and CD71 were lower, meanwhile 145 cases were accompanied with lymphocyte antigen expression, the ratios were as follows: CD7 21.5%, CD19 6.0%, CD2 0.78%, CD10 0.58% and CD20 0.58%; the positive expression rate of CD71 in M6 was 100%, and that of CD64 in M5 was the highest (30.2%); the overall positive rate of CD34 was 57.8%. (2) After first chemotherapy, the complete remission (CR) rate was 64.7%, CR rate of CD34(+) patients was lower than that of CD34(-) in M3 group (P = 0.019). The CR rate of CD34(+) patients was significantly lower than that of CD34(-) in non-M3 group (P = 0.002). The CR rate of CD19(+) patients was higher than CD19(-) (P = 0.028); the CR rate of CD7(+) patients was significantly lower than that of CD7(-) (P = 0.002); the CR rate of CD71(+) patients was lower than that of CD71(-) (P = 0.013); the CR rate of MPO(+) patients was higher than that of MPO(-) (P = 0.015). Between the CR rate of CD11b, CD13, CD33-positive and-negative group, the difference was not statistically significant (P > 0.05). It is concluded that the phenotype is a prerequisite for the diagnosis of AML, and can help to guide the clinical typing, selection of treatment protocols and evaluation of prognosis.