Knockout of Micro-RNA-21 Gene in DLBCL OCI-Ly3 cells by TALEN Technique / 中国实验血液学杂志

<p><b>OBJECTIVE</b>To explore an efficient way to knockout microRNA genes in hemapoietic cell lines with a very low transfection efficiency, so as to facilitate the study of microRNA function in hematopoietic malignancies.</p><p><b>METHODS</b>TALE-nucleases was utilized to knockout the microRNA-21 gene in human diffuse large B-cell lymphoma cells (OCI-Ly3). The OCI-Ly3 single cell clones without expression of miR-21 were established through eGFP(+) enrichment, PCR screening, and microRNA quantification. Finally, the miR-21 changes of mutant clones were identified by sequencing.</p><p><b>RESULTS</b>Four miR-21-knockouted OCI-Ly3 single-cell-derived clones were established after 2 round transfection and screening. The miR-21 knockout efficiency was around 10/10(6) original cells. Sequencing the mutant clones indicated that miR-21 expression could be drastically reduced by simply altering sequences immediately adjacent to the microRNA duplex.</p><p><b>CONCLUSION</b>This strategy may be applied to knockout any microRNA of interest even in hemapoietic cell lines with very low transfection efficiency.</p>

Efficacy of CALLG2008 protocol in treatment of adult acute lymphoblastic leukemia: a single center analysis / 中国实验血液学杂志

CALLG2008 Protocol is sequential chemotherapy for adult acute lymphoblastic leukemia (ALL) established by Collaborative Group of adults acute lymphoblastic leukemia. It is emphasized that comprehensive treatment of adult ALL according to risk stratification is rather important. This study was purposed to evaluate the therapeutic efficacy of CALLG2008 for adult ALL. The clinical data of adult ALL patients of ≥ 14 years old diagnosed and treated by CALLG2008 Protocol were collected from May 1, 2009 to December 31, 2011 in Fujian Medical University Union Hospital, and the efficacy was analyzed. The results showed that 31 out of 33 cases of ALL achieved CR, the CR rate was up to 93.9%, the PR rate was 3.1%, and the total response rate was 97%. There were no uncontrolled severe toxicities, and no early deaths were observed. The overall survival (OS) at 1 year was only 66.7%,the relapse rate was 43.8% and the 1-year mortality was 33.3 %. This may be related with no-enough compliance, no-enough economical support and short follow-up time of the patients. The risk factor analysis showed that WBC level in newly diagnosed patients may influence the OS and relapse-free survival (RFS) of ALL. It is concluded that CALLG2008 protocol applied to adult ALL has a high remission quality and low mortality rate during the induction. The disease free survival (DFS) needs to be observed longer. It is essential to carry out MRD monitoring to determine the early recurrence and improving the long-term efficacy.

Efficacy of remission induction chemotherapy and prognostic analysis in elderly patients with acute myeloid leukemia / 中华血液学杂志

<p><b>OBJECTIVE</b>To explore the outcome of remission induction chemotherapy (IC) and prognostic in elderly patients with acute myeloid leukemia (AML).</p><p><b>METHODS</b>The clinical data of 156 AML patients older than 60 years in the Institute of Hematology, Union Hospital of Fujian Medical University from January 2003 to July 2010 were analyzed retrospectively. 104 patients received cytarabine-based regimens, including protocol DA,IA or CAG,while 52 patients received palliative treatment. The median survival time was compared between patients with and without IC. The prognostic factors were evaluated by using univariate and multivariate analyses.</p><p><b>RESULTS</b>145 (93%) cases were followed-up. The median survival time was 316 days in 96 IC patients, compared with 37 days in 49 PT patients (P < 0.01). Not receiving induction chemotherapy,high-risk karyotype,hyperleukocytosis (> or = 100 x 10(9)/L), Charlson Comorbidity Index (CCI) > or = 2 were adverse prognostic factors of the survival time with univariate analysis, and all were independent poor factors affecting the survival time with multivariate analysis.</p><p><b>CONCLUSIONS</b>IC can improve outcomes in elderly AML patients. The patients with hyperleukocytosis (> or = 100 x 10(9)/L) , high-risk karyotype, CCI > or = 2 and without receiving IC have poorer prognosis.</p>

Effects of baicalin on HL-60 cell xenografts in nude mice and its mechanism / 中国实验血液学杂志

This study was aimed to investigate the effects of baicalin on HL-60 cell xenografts in nude mice in vivo and explore its mechanism. Xenograft tumor model of HL-60 cells in nude mice was established, which was divided randomly into 6 groups: negative control group (injection of 5% NaHCO(3)), 25, 50 and 100 mg/kg baicalin groups, combination group (50 mg/kg baicalin + 2 mg/kg VP16) and positive control group (VP16 4 mg/kg). The nude mice with HL-60 cell xenografts were treated with drugs via intraperitoneal injection daily. After treatment for 14 days average weigh and inhibitory rate of transplanted tumor stripped from 5 nude mice in each group were calculated, and the ultrastructure change of xenografts cells were tested by transmission electron microscopy. Histopathologic examination was used to observed the change of main organs in nude mice. The expression of signaling molecular PI3K/Akt proteins extracted from xenografts was detected by Western blot. The effects of baicalin on overall survival time in nude mice with HL-60 cell xenografts were evaluated. The results showed that baicalin could inhibit the growth of transplanted tumors in dose-dependent manner. There were more necrotic and apoptotic cells in mice of baicalin-treated groups and combination group than that in mice of negative control group. Baicalin could inhibit the proliferation of HL-60 cells in vivo by down-regulating the PI3K/Akt/mTOR signal pathway, where the expressions of p-Akt, mTOR and p-mTOR proteins decreased compared with negative control group, and no significant difference of Akt expression was found between different groups. Compared with negative control group, the median survival time of mice in combination group was more prolongated (P < 0.05). It is concluded that baicalin can inhibit growth and induce apoptosis of HL-60 cell xenografts in nude mice, and prolong median survival time of nude mice. The possible mechanisms may be related to inhibition of Akt activity and down-regulation of the PI3K/Akt/mTOR signal pathway. The combination of baicalin and VP16 shows a synergistic effect on inhibiting growth of HL-60 cell xenografts in nude mice.

Two novel splicing variants of eIF4E obtained by electronic cloning technique combined with RT-PCR / 中国实验血液学杂志

In order to clone the full-length cDNA of a novel EST which is probably related to acute leukemia relapse and to analyse the sequences, the electronic cloning technique combined with RT-PCR was used to clone the full-length cDNA, and the sequences were analyzed by bioinformatics. The results showed that the two novel splicing variants of eIF4E named as splicing variant 1 and 2 of eIF4E were obtained. Bioinformatics analysis showed that variant 1 and 2 exhibited 84% and 47% similarity to eIF4E mRNA, and were localized on eIF4E locus on chromosome 4. The lengths of 2 variants are 1 904 bp and 3 393 bp, encoding 245 amino acids and 132 amino acids respectively. BLAST results showed that the both variants mentioned above contains seven exons. Among them, the sequences of the three exons at 5' end of variant 1, variant 2 and eIF4E mRNA were different from each other. Protein BLAST showed that they are partially different from eIF4E protein. It is concluded that the two novel splicing variants of eIF4E were cloned, and their relation to acute leukemia relapse needs to be further investigated.

Effects of baicalin on proliferation and apoptosis of adriamycin-resistant human leukemia HL-60/ADR cells / 中国实验血液学杂志

This study was purposed to explore the effects of baicalin on proliferation and apoptosis of adriamycin-resistant human myeloid leukemia cell line HL-60/ADR. HL-60/ADR cells were in vitro cultured and its proliferation inhibition was detected by MTT assay. The cell apoptosis was tested by Annexin V FITC/PI double staining analysis, DNA fragmentation and TUNEL labeling method. The expressions of c-myc and bcl-2 mRNA were detected by RT-PCR, and the protein expressions of C-MYC, BCL-2, caspase-3 precursor (procaspase-3), PARP and BAD were determined by Western blot. The results showed that baicalin could remarkably inhibited the HL-60/ADR cell proliferation, the cell doubling time was 48 hours, with an IC50 value of 28 micromol/L. Apoptosis occurred in dose dependent manner (20, 40, 80 micromol/L), and cell apoptosis in earlier and later stages could be detected by Annexin V FITC/PI double staining analysis, DNA fragmentation and TUNEL labeling method. The expressions of c-myc and bcl-2 mRNA in baicalin-treated cells decreased in a time-dependent manner (12, 24, 48 hours). Meanwhile, protein expressions of C-MYC, BBL-2, procaspase-3 and PARP (116 kD) were down-regulated in a time-dependent manner, while the expression of PARP (85 kD) and BAD were up-regulated. It is concluded that the baicalin efficiently induces proliferative inhibition and apoptosis in HL-60/ADR cells. All of above related genes and proteins may be involved in these processes.

Differentiation-inducing effects of baicalin on HL-60 leukemia cells / 中国病理生理杂志

AIM:To investigate the differentiation-inducing effects of baicalin on HL-60 leukemia cells.METHODS:The effect of baicalin on differential induction in AML cell line HL-60 was evaluated by cellular morphology,clone formation assay,CD11b and CD33 expression and NBT assay.RESULTS:Baicalin inhibited the proliferation of HL-60 cells.It enhanced the expression of CD11b on HL-60 cells,also increased the expression of CD33.HL-60 cells showed differentiation morphology after the drug treatment examined by Wright-Gimesa staining and NBT assay.CONCLUSION:Baicalin possessed differentiation-inducing effects on HL-60 cells.