Complete mitochondrial genome of the longicorn Anoplophora horsfieldi Hope (Coleoptera: Cerambycidae).

The chinaberry yellow-banded longhorn beetle, Anoplophora horsfieldi Hope 1842 (Coleoptera: Cerambycidae) is an important pest on many economic tree species. In this study, the complete mitochondrial genome of A. horsfieldi was determined, which was 15,837 bp in length and contained 37 genes, including 13 protein-coding genes (PCGs), two rRNA, 22 tRNA genes, and a non-coding A + T-rich region. The phylogenetic analysis based on mitochondrial genomes showed that A. horsfieldi is sister to a clade formed by A. chinensis and A. glabripennis.

Complete mitochondrial genome of the olive weevil, Dyscerus cribripennis (Coleoptera: Curculionidae).

The olive weevil Dyscerus cribripennis (Coleoptera: Curculionidae) is an uncontrollable noxious insect to Olea europaea. The 15,977 bp complete mitochondrial genome of D. cribripennis contained 13 protein-coding genes (PCGs), 2 ribosomal RNA genes (rRNAs), 21 transfer RNA genes (tRNAs), and a control region (GenBank accession number MW023069). The trnI was not found in the D. cribripennis mitogenome. The phylogenetic analysis based on mitogenomes showed that D. cribripennis is closed related with Hylobitelus xiaoi.

[Impact of hepatitis B virus infection on liver function after hematopoietic stem cell transplantation].

To analyze the impact of hepatitis B virus (HBV) infection on liver function of patients after hematopoietic stem cell transplantation (HSCT), the transplantation outcome of 48 patients infected with HBV prior to transplantation among 185 patients received HSCT was investigated retrospectively. The results showed that during a follow-up for 6 months after HSCT, the alanine aminotransferase (ALT) peak average values of the patients with HBsAg(+), HBsAb(+) and control groups were (281.6 ± 414.6), (95.4 ± 79.9) and (65.1 ± 44.2) U/L, respectively. The incidences of abnormal liver function of the patients with HBsAg(+), HBsAb(+) and control groups were 61.54%, 40.00% and 30.23% respectively. There were no significant differences between any two groups (P > 0.05). The lethality of those patients at late period after transplantation was not related to HBV infection. The hepatocirrhosis and hepatocarcinoma caused by HBV infection have not become major problems in long-term survivors. It is concluded that in HBsAg(+) patients received HSCT, the damage of liver function is more severe than control group, possibly increasing the development of abnormal liver function. The measures against the liver function damage should be taken. The prophylactic administration of ganciclovir for virus may be effective to prevent the activation of HBV.

[Arsenic trioxide inhibits cell growth in imatinib-resistant bcr-abl mutant cell lines in vitro].

OBJECTIVE: To explore the effect of arsenic trioxide (As2O3) on the growth inhibition of imatinib (IM)-resistant bcr-abl mutant cell lines in vitro. METHODS: Cell growth of one IM-sensitive cell line, 32Dp210 and 15 IM-resistant cell lines including T315I and other 14 bcr-abl mutants were detected by MTT assay after treatment with IM and As2O3. The cell lines with 5 frequently observed mutants in CML patients were analyzed for apoptosis by flow cytometry with Annexin V and PI staining as well as the expression of bcr-abl fusion protein, phosphorylated CRKL protein and apoptosis-related proteins by Western blot. RESULTS: The fifty percent inhibition concentration (IC50) values of As2O3 for 15 IM-resistant cell lines were 2.6-5.3 fold lower than that for IM-sensitive cell line. For the 5 bcr-abl mutants frequently happened in CML patients, As2O3 significantly inhibited the expression of bcr-abl fusion protein and phosphorylated CRKL and induced apoptosis in a dose-dependent manner as compared with that for 32Dp210. Coincidently, the cell apoptosis was induced through caspase-3, 8 and 9 pathways. CONCLUSION: As2O3 remarkably inhibits cell growth and induces apoptosis of IM-resistant bcr-abl mutant cell lines in vitro, suggesting that it might be a potential therapeutic agent for IM-resistant CML patients.

[Effect of a novel tyrosine kinase inhibitor HHGV678 on growth inhibition of Bcr-Abl wild type and IM-resistant cell lines in vitro].

This study was aimed to compare HHGV678 with imatinib (IM) in growth inhibition of Bcr-Abl wild type and IM-resistant cell lines, investigate the possibility of replacing IM with HHGV678 in treatment of chronic myeloid leukemia (CML) and IM-resistant CML patients. Viability of two Bcr-Abl wild type cell lines (K562 and 32Dp210) and 16 IM-resistant cell lines (K562R and 15 Bcr-Abl point mutant cell lines) treated with HHGV678 and IM was analyzed by MTT. The apoptosis of those cells was identified by flow cytometry with Annexin V staining and DNA ladder analysis. Western blot was applied for detecting the expression of Bcr-Abl and phosphotyrosine protein levels. The results indicated that HHGV678 significantly inhibited the growth of two Bcr-Abl wild types and IM-resistant cell lines in dose-dependent manner except cell line of T315I point mutant. IC(50) results showed that the growth inhibition of HHGV678 was 15.5 and 28-fold higher than that of IM in K562, 32Dp210 and 1.4 to 124.3-fold higher than that of IM in 15 IM-resistant cell lines respectively. Compared with IM, HHGV678 more significantly inhibited phosphotyrosine kinase protein of the cells mentioned above at different concentrations. With most importance, HHGV678 of 10.0 micromol/L induced cell apoptosis of 40.06% and 33.32% in K562R and 32Dp210(T315I) cell lines, which were much higher than that of IM (19.77% and 10.68%). It is concluded that HHGV678 is more effective than IM in the growth inhibition of Bcr-Abl wild type cell lines and IM-resistant cell lines, especially in strongest IM-resistant cell lines. Further studies are needed to show whether HHGV678 may be a novel targeting drug in treatment of CML and IM-resistant CML patients.

[Late-onset noninfectious pulmonary complications after allogeneic peripheral blood stem cell transplantation].

The aim of study was to explore the incidence, risk factors, outcome and efficacious treatment of late-onset noninfectious pulmonary complications (LNIPC) after allogeneic peripheral blood stem cell transplantation (allo-PBSCT). Seventy patients received allo-PBSCT were analyzed retrospectively. The results showed that 9 out of 63 patients surviving more than 3 months occurred late-onset noninfectious pulmonary complications (14.3%). Five out of the 9 patients developed secondary pulmonary infections. In 4 patients, LNIPC caused death directly. Advanced stage of disease at transplantation and extensive chronic graft-versus-host disease (GVHD) happened in association with LNIPC. However, other transplantation-related factors including age at transplantation, gender of patient, conditioning regimen, HLA matching and GVHD prophylaxis were not significantly correlated with the incidence of LNIPC. It is concluded that performing pulmonary function test (PFT) and thoracic computer tomography should be taken routinely after transplantation. Most patients who get correct and early diagnosis for LNIPC will show a positive response to prednisone with or without CsA.

[Growth inhibition and differentiation of imatinib-resistant chronic myeloid leukemia cell induced by cell differentiation agent in vitro].

OBJECTIVE: To investigate the effects of uroacitide (CDA-2), a cell differentiation agent, on the growth inhibition and differentiation of imatinib-(IM) resistant chronic myeloid leukemia (CML) cells. METHODS: IM resistant CML cell line K562R was established from the line K562. K562 and K562R CML cells were cultured with CDA-2 of different concentrations. MTI method was used to detect the survival rates. Bone marrow cells of IM-resistant and non-IM-resistant CML patients were collected and co-incubated with K562 and K562R cells. MTT and colony-forming assays were used to evaluate the efficacy of CDA-2 treatment for cell growth in K562 and K562R cell lines, and IM-resistant or non-IM-resistant bone marrow cells of the CML patients; Annexin-V staining was employed to detect the apoptosis. Cell differentiation was assessed by flow cytometry analysis with CD11b/CD14 markers, reverse transcriptase PCR (RT-PCR) for mRNA levels of NCF-1 and ORM-1 genes and Giemsa staining for the observation in morphology. Cell cycle distribution was detected by stained with propidium iodide and then analyzed by flow cytometer. RT-PCR also was employed for the expression of DNA methyltransferase. RESULTS: Significant cell growth inhibition was found at a dose-dependent manner in the IM-resistant K562R cell line and IM-resistant bone marrow cells of the CML patients compared with the non-resistant K562 cell line and bone marrow cells of the CML patients following 7 days exposure to CDA-2. Although CDA-2 could significantly induce the apoptosis of K562R (15.38%) compared with K562 (5.28%) (P < 0.05), the major reason for the cell growth inhibition of K562R is CDA-2-induced cell differentiation, including the increase of expression of differentiation-related antigens CD11b/CD14, mRNA expression of NCF-1 and ORM-1, and cell cycle arrest in G1-phase at a dose-dependent manner. Because CDA-2 could significantly activate the p21 and p27 gene expression, downregulate the expression of cyclin D1, and down-regulate the expressions of DNMT1 and DNMT(3B) at mRNA level, CDA-2 might be a DNMT inhibitor for restoring some gene function that involved in cell cycle control by demethylation. CONCLUSION: Inhibiting the growth and inducing the differentiation of K562R cells, CDA-2 is very likely to be a potential agent for the treatment of IM resistance CML patients.

[Clinical study on pulmonary complications after allogeneic peripheral blood stem cell transplantation].

OBJECTIVE: To explore the incidence, pathogenesis, risk factors and effective treatment of pulmonary complications after allogeneic peripheral blood stem cell transplantation (allo-PBSCT). METHODS: Pulmonary complications in 70 patients received allo-PBSCT were analyzed. RESULTS: Thirty one episodes were observed in 26 patients. Among them episodes were infectious complications, including bacteria pneumonia, pulmonary fungus disease, CMV interstitial pneumonia and tuberculosis, some cases were caused by two pathogens, and 11 episodes were noninfectious complications, including late-onset noninfectious pulmonary complications (LONIPCs) (n=9), pulmonary edema (n=1) and interstitial pneumonia (n=1). The overall mortality was 12.9%. Graft-versus-host disease (GVHD) prophylaxis without MTX, severe acute GVHD and extensive chronic GVHD were high risk factors for pulmonary complications and advanced disease at transplantation, extensive chronic GVHD were significantly associated with the incidence of LONIPCs. CONCLUSION: Pulmonary disease is the main complication occurred in patients undergoing allo-PBSCT. It is of greatly importance to treat pathogens specifically and diagnose LONIPCs in its early stage.

[Unrelated donor peripheral blood stem cell transplantation for hematologic malignancies].

This study was aimed to explore feasibility and efficacy of unrelated donor peripheral blood stem cell transplantation (UD-PBSCT) in treatment of hematologic malignancies. Ten patients with hematologic malignancies underwent high resolution DNA based typing HLA-matched or 1 locus mismatched UD-PBSCT. Busulfan, cyclophosphamide, Ara-C, MeCCNU and antithymocyte globulin (ATG) were used for preconditioning regimen in all cases. All patients received mycophenolate mofetile, cyclosporin A and short-term methotrexate with CD25 antibody as the graft-versus-host disease (GVHD) prophylaxis. The results showed that rapid engraftment was observed in all cases who presented full donor chimerism at 28 days post transplantation by STR-PCR. The median time of neutrophil recovery > 0.5 x 10(9)/L, platelet recovery > 20 x 10(9)/L was 13, 17.5 days respectively after transplantation. The incidence of acute GVHD was 3 cases (one case with grade I was recovered from GVHD by himself, one case with grade III was cured, one case with grade VI was died). It is concluded that above-mentioned preconditioning regimen and GVHD prophylaxis are effective approaches for unrelated donor peripheral blood stem cell transplantation in hematopoietic malignancies.

[Comparison of rituximab plus CHOP regimen and CHOP regimen alone for treatment of newly diagnosed patients with diffuse large B-cell lymphoma].

BACKGROUND & OBJECTIVE: CHOP regimen is the standard treatment for patients with diffuse large B-cell lymphoma. Rituximab, an anti-CD20 monoclonal antibody, is effective in treating diffuse large B-cell lymphoma. This study was conducted to compare the efficacy of rituximab plus CHOP and CHOP alone on newly diagnosed patients with diffuse large B-cell lymphoma, and analyze their toxicities. METHODS: A total of 72 newly diagnosed patients with diffuse large B-cell lymphoma were divided into 2 groups prospectively with concurrent control: 34 received CHOP plus rituximab (375 mg/m2, 2 days before each course) (combination group), 38 received CHOP alone. Each course lasted 3 weeks. All cases were evaluated after 6 courses. RESULTS: The total response rate in combination group was 93.8% (30/32), among which complete remission was seen in 23 patients and partial remission was seen in 7 patientsû while the total response rate in CHOP group was 75.0% (27/36), among which complete remission was seen in 19 patients and partial remission was seen in 8 patients. The therapeutic efficacy was significantly better in combination group than in CHOP group (P<0.05). The 1-year progression-freely and overall survival rates were significantly higher in combination group than in CHOP group (81.2% vs. 52.8%, 93.8% vs. 75.0%, P<0.05). The major adverse events in combination group were infusion-related response which could be well tolerated, and hematological toxicities which were similar to those in CHOP group. CONCLUSIONS: Rituximab increases the therapeutic efficacy of CHOP regimen on newly diagnosed patients with diffuse large B-cell lymphoma, without a clinically significant increase in toxicity. Rituximab plus CHOP can be used as a first-line therapy of diffuse large B-cell lymphoma.

[Complications of successively double autologous hemopoietic stem cell transplants].

In order to get clinical information about safety and feasibility of successively double autologous hemopoietic stem cell transplants (SD-AHSCT) in malignant hematological disease patients, the complications and hematological reconstitution after SD-AHSCT in 20 patients were analyzed retrospectively. 20 patients with hematologic malignancies received autologous peripheral blood stem/progenitor cell transplantation at the first transplant, and then were given autologous bone marrow transplantation as the second transplant at 4-10 months. The results showed that all the patients tolerated mobilization and collection of peripheral blood stem/progenitor cells as well as bone marrow collection. All the patients got enough hematological stem/progenitor cells for SD-AHSCT and achieved hematological reconstitution after SD-AHSCT. The speed of hematological reconstitution was positively correlated with the transfused quantity of hematological stem/progenitor cells (r = 0.968). The hematological reconstitution after the first autologous hemopoietic stem cell transplant (AHSCT) was earlier than that of the second (P < 0.05). There was no statistical difference between the first and the second AHSCT for the incidence of skin or mucous membrane bleeding (P > 0.05). No patients occurred massive hemorrhage during SD-AHSCT. The quantity of platelet transfusion in the second AHSCT was larger than that in the first AHSCT (P < 0.01). The incidence of oral ulcer in the first AHSCT was significantly higher than that in the second (P < 0.01). No statistical difference between the first and the second AHSCT was there in infectious sites, infectious pathogens and infection incidence (P > 0.10). All the complications were improved or cured, and no patients died of SD-AHSCT complications. In conclusion, SD-AHSCT is safe and feasible, and worthy to be further popularized.

[Unrelated HLA-matched donor marrow transplantation for one case of myelodysplastic syndrome].

To explore feasibility and efficacy of unrelated HLA matched donor marrow transplantation in treatment of myelodysplastic syndrome, one case (male, 31 years old) of myelodysplastic syndrome-refractory cytopenia with multilineage dysplasia (MDS-RCMD) has been received unrelated HLA-matched donor transplantation. Busulfan, cyclophosphamide, Ara-C, MeCCNU and antithymocyte globulin (ATG) were used for preparative regimen. Mycophenolate mofetile, cyclosporin A and short-term methotrexate were used for graft-versus-host disease prophylaxis. The results showed that neutrophil of > 0.5 x 10(9)/L, platelet of 58 x 10(9)/L and hemoglobin of 114 g/L were observed at 10, 20 days and 3 months respectively post transplantation. Disease-free survival without GVHD was 9 months. In conclusion, unrelated HLA matched donor marrow transplantation is an effective approach for treatment of patients with MDS.

[Comparison of curative effect of autologous peripheral blood stem cell transplantation versus bone marrow transplantation for acute leukemia].

To compare the clinical outcome of autologous peripheral blood stem cell transplantation (APBSCT) and autologous bone marrow transplantation (ABMT) in treatment of patients with acute leukemia in first remission, 41 patients received APBSCT, 17 patients received unpurged ABMT and 30 patients received purged ABMT. The results showed that hematopoietic recovery was significantly earlier after APBSCT than that after purged or unpurged ABMT. The 3-year disease-free survival (DFS), relapse rate (RR) and transplant-related mortality (TRM) for all patients of 3 groups were 51.7%, 41.7% and 6.8%, respectively. DFS and RR were significantly influenced by disease types (ALL or AML) and intervals between diagnosis and CR(1) or CR(1) and transplant. The main causes of transplant-related death were infection and hemorrhage. After APBSCT, DFS, RR and TRM were 48.4%, 43.9% and 4.9%, respectively, and did not differ significantly from those found in unpurged ABMT (47.1%, 45.6% and 11.8%) or purged ABMT (66.5%, 29.6% and 6.7%). It is concluded that the clinical outcome of APBSCT is similar to unpurged or purged ABMT but APBSCT allows faster recovery of hematopoiesis and needs less transfusion support.

[The effect of separating red blood cells from bone marrow graft in vitro by methylcellulose].

It has need to separate red blood cells (RBC) from marrow graft in ABO group unmatched BMT and auto-BMT with purging tumor cells, the separating effect of methylcellulose was observed. The mixture of 0.5% methylcellulose and bone marrow was laid up in an open transfusion system, and then sedimentation of RBC was performed in the transfusion tube. The separating results of 18 marrow grafts showed that the recovery rates of mononuclear cells and CD34(+) cells were (83.8 +/- 55.2)% and (90.3 +/- 7.2)%, respectively. RBC residual rate was (4.3 +/- 1.5)%. The yield of CFU-GM was (60.8 +/- 22.4)/2 x 10(5) MNC, and there was no difference to [(69.8 +/- 23.4)/2 x 10(5) MNC] yielded from same marrow samples, separated by Ficoll-Hypaque separation. It is concluded that this method could be used for bone marrow transplantation.

[Comparative study on activated immunocytes of human bone marrow and peripheral blood by cytokines].

To study immunophenotype and cytotoxicity of the immunocytes in bone marrow and peripheral blood after activation by combined cytokines, mononuclear cells (MNC) of bone marrow and peripheral blood were activated by IFN-gamma, IL-1, IL-2 and McAb-CD3 in vitro. The cell amount and morphology during culture were observed. Cytochemical staining and immunophenotype analysis were done before and after culture in two groups of MNC. Cytotoxicity was tested by MTT method. The results showed that the cell number of two groups increased obviously in culture (P < 0.05), while the peripheral blood mononuclear cells increased more markedly (P < 0.05). The cytochemical staining showed POX decrease, but PAS increase in two groups. The positive ratios of CD3(+), CD56(+) and CD38(+) cells in two groups increased obviously after culture (P < 0.05), but there was no significant difference between those two groups. CD3(+) CD56(+) cells increased obviously in peripheral blood mononuclear cells activated by cytokines (P < 0.05), but CD3(+) CD56(+) cells did not increase in bone marrow mononuclear cells. There was no significant difference between two groups' cytotoxicity. It was concluded that IFN-gamma, IL-1, IL-2 and McAb-C D3 increased cell number and cytotoxicity of both bone marrow and peripheral blood mononuclear cells that can be used in cell immunotherapy.

[Change of adhesion molecule expression on CD34(+) cells from bone marrow and peripheral blood during mobilization with combination of chemotherapy and G-CSF].

To explore the dynamic change of CD34(+) cell expressing adhesion molecules in bone marrow and peripheral blood during mobilization with combination of chemotherapy and G-CSF and its clinical significance, mononuclear cells of bone marrow and peripheral blood from malignant hematopathy cases before and after mobilization with G-CSF were labeled by CD45-CY-Chrome, PE conjugated anti-CD34, and FITC conjugated anti-CD44, anti-CD49d, anti-CD62L and anti-CXCR4. For three-color fluorescence analysis by flow cytometry was performed on a FACScalibur. Also the relationship between the number of subpopulations in different expressions of adhesion molecules infused and the time of recovery in different blood cells after transplantation was evaluated. Results showed that a significantly lower expression of CD44(+) and CD49d(+) on CD34(+) cells in bone marrow after mobilization compared to that before mobilization, whereas great higher expression of CD44(+), CD49d(+), anti-CD62L(+) and lower of anti-CXCR4(+) in peripheral blood were observed after mobilization. No significant relations were found between expression of different adhesion molecules on CD34(+) cells infused and the time of reconstitution in blood cells after transplantation. It was concluded that this mobilizing regimen could downregulate the expressions of CD44, CD49d, CD62L, and anti-CXCR4 on CD34(+) cells in bone marrow, it may related to mobilization of CD34(+) cells from marrow to blood, and homing of blood CD34(+) cells into marrow.

[A Simplified Method for Cryopreservation of Peripheral Blood Stem Cells]

To simplify the traditional method for cryopreservation of peripheral blood stem cells(PBSCs) at -196 degrees C with rate-controlled freezing with 10% dimethyl sulfoxide(DMSO), the simplified method was carried out by freezing the cells to -80 degrees C in low-temperature freezer with the combination of 5% DMSO, 3% hydroxyethyl starch(HES) and 4% human serum albumin(HSA) as cryoprotectant. PBSCs were cryopreserved by different methods. Cell viability and recovery rate of mononuclear cells (MNC), CFU-GM and CD34(+) cells were compared. It was observed that the higher MNC and CFU-GM recovery rates were achieved and without agglutination with the simplified method. It was also found with this simplified method, satisfactory recovery rates of CFU-GM and CD34(+) cells could be obtained when PBSCs were preserved at -80 degrees C as long as 24 months. There was no difference observed in parameters of cryopreserved PBSCs thawed at 37 degrees C and 20 degrees C. After the cells being exposed to 5% DMSO at room temperature for 1 hour, the cell viability decreased from 93.2% to 84.5%, however, the CFU-GM recovery rate was not decreased. It is concluded that the simplified cryopreservation technique is better and simpler than the traditional crypreservation method, will be useful for institutions without rate-controlled freezing facility. Moreover, this method diminishes the amount of DMSO infused into patients, thus decreasing its toxicity.

[Morphology and Immunophenotype Study of Bone Marrow Mononuclear Cells after Activated by Different Cytokines In Vitro]

Bone marrow cells in cultures were divided into four groups and cultured with various cytokines in vitro. These four groups are: control, IL-2 group, CD3-AK group, and CIK group. The morphological (cell volume, nucleus/plasm) changes of bone marrow cells in culture were observed. Immunophenotype analysis (CD34, CD38, CD3, CD56) were done before and after culture in all groups. Cytotoxicity against fresh acute leukemia cells were detected by modified MTT methods. The cell volume became larger with increased nucleus/plasm ratio in IL-2 group, CD3-AK group and CIK group. The plasm filled with PAS positive granules in most of cells in CD3-AK group and CIK group. The positive ratio of CD3, CD56, CD38 in CD3-AK or CIK group increased markedly after culture (P < 0.05), but no significant difference between the two groups. The CD56(+) cell increased in IL-2 group. CD34(+) cells decreased in all groups and there were no significant differences among those four groups. The cytotoxicity to fresh leukemia cells: CD3-AK group and CIK group > IL-2 group > control group. There was no significant difference between CD3-AK group and CIK group. This experiment showed different effect on bone marrow cells by different cytokine combination. The cytokine combination of CD3-AK group or CIK group can make immunocytes of bone marrow proliferating and retained certain amount of stem/progenitor cells.

[Activation and Cytotoxicity of Bone Marrow Immunocytes by Using Various Cytokines]

The study aimed to explore the changes of the immunocyte quantities and cytotoxicity in bone marrow, and how many hematopoietic progenitor cells retained after the bone marrow cells were activated by different combinations of various cytokines. Bone marrow cells were divided into four groups and were cultured in vitro: (1) Control: no cytokines were added. (2) IL-2 group: bone marrow cells were activated by IL-2. (3) CD3-AK group: bone marrow cells were activated by IL-2 and CD3-McAb. (4) CIK group: IFN-gamma, IL-1, IL-2 and CD3-McAb were added. CFU-GM assay and CD3 phenotype detection were performed before and after activating culture in all groups. The changes of cell quantities during culture and cytotoxicity of cultured cells were tested. CD3 positive cells markedly increased in both CD3-AK and CIK groups. The cell numbers and cytotoxicity of CD3-AK and CIK groups were higher than those of control or IL-2 group obviously after culture (P < 0.05). CFU-GM were decreased in all groups after culture and there had no significant difference among four groups. The combination of IL-2 and CD3-McAb not only stimulates the proliferation of marrow immunocytes and increases their cytotoxicity but retains enough hematopoietic progenitor cells as well. This combination of cytokines can be used to purge autologous bone marrow in vitro.