OBJECTIVETo explore the effect of down-regulation of insulin-like growth factor binding protein 7 (IGFBP7) on the proliferation and invasiveness of leukemia cell line U937 cells.

METHODSThree pairs of double-strand siRNA targeting IGFBP7 gene were transfected into SMMC7721 cells to select the most efficient one for U937 cells. qRT-PCR and Western blot were used to detect the expression of IGFBP7 in U937 cells after transiently transfected with siRNA of IGFBP7. Cell proliferation, adhesion, trans-endothelial migration and invasion were performed in transfected cells and control groups.

RESULTSAfter transfected with siRNA of IGFBP7 in U937 cells, the ability of cell proliferation was significantly decreased at 24 h (0.580 ± 0.159) compared to that of parental cells and scramble negative control (1.049 ± 0.274, 0.946 ± 0.195, respectively) (P < 0.01). Adhesion of U937 cells transfected with IGFBP7 gene specific siRNA to ECV304 cells was significantly lower than that of the control groups (0.247 ± 0.031 vs 0.406 ± 0.023 and 0.395 ± 0.011) (P < 0.01). Transendothelial membrane of U937 cells into the bottom of the 24-well plate for experimental group were less than those in the control groups \[(0.387 ± 0.021)×10(5) vs (1.017 ± 0.031)×10(5) and (0.908 ± 0.027)×10(5)\]. Cells adherent to the matrigel for experimental group were less than those in the control groups \[(0.197 ± 0.098)×10(5) vs (0.493 ± 0.067)×10(5) and (0.469 ± 0.083)×10(5)\]. The difference was significant (P < 0.01).

CONCLUSIONIGFBP7 gene plays a contributing role in leukemogenesis involving in leukemic cells' proliferation and interaction with endothelial cells through adhesion, invasion and migration.

Cell Proliferation ; Down-Regulation ; Humans ; Insulin-Like Growth Factor Binding Proteins ; genetics ; metabolism ; Leukemia, Myeloid, Acute ; metabolism ; pathology ; Transfection ; U937 Cells

Diagnosis, Differential ; Fatty Liver ; diagnosis ; Humans ; Liver Neoplasms ; diagnosis ; Magnetic Resonance Imaging ; Male ; Middle Aged ; Tomography, X-Ray Computed

This study was purposed to culture murine compact bone-derived mesenchymal stem cell (MSC) and analyze the immunological and trilineage differentiation potential. Tibia and femur were extracted. Bone marrow cells were flushed out and compact bone fragments were digested with collagenase. The digested cells were cultured in 6-well plates. The immunophenotype, immunosuppressive function and trilineage differentiation potential were analysed by flow cytometry, mixed lympocyte reaction and Oil red O, von Kossa and alcian blue straining, respectively. The results indicated that the pure compact bone MSC could be isolated with in 3 weeks. The resulting MSC had trilineage differentiation potential and immunosuppressive effect on mixed lymphocyte reaction. The count per minute (CPM) value in control group of BALB/c T cells cocultured with irradiated C57BL/6 T cells was (2.56 ± 0.31) × 10(4), while CPM values of mixed lymphocyte cocultured with C57BL/6 compact bone MSC at ratios of 100:1 and 10:1 were (0.47 ± 0.12) × 10(4) and (0.28 ± 0.09) × 10(4). The CPM value of control group was higher than those of MSC cocultured group (P < 0.001). Compact bone-MSC had an immunosuppressive effect on mixed lymphocyte reaction in a dose dependent manner. It is concluded that murine compact bone has rich MSC and the primary MSC is contaminated with less hematopoietic cells. Murine compact bone-MSC have immunosuppressive effect on mixed lymphocyte reaction and trilineage differentiation potential. Compact bone-MSC have promising experimental study value.
Animals ; Bone Marrow Cells ; cytology ; immunology ; Bone and Bones ; cytology ; Cells, Cultured ; Female ; Immunophenotyping ; Lymphocyte Culture Test, Mixed ; Mesenchymal Stromal Cells ; cytology ; immunology ; Mice ; Mice, Inbred BALB C ; Mice, Inbred C57BL

To investigate the effects of 3-hydroxy-3-methylglutaryl coenzyme A reductase inhibitor simvastatin (SV) on proliferation, apoptosis and the PI3K/AKT signaling pathway in human acute monocytic leukemia cell line SHI-1. SHI-1 cells were incubated with different concentrations of SV (5, 10, 15 µmol/L). Otherwise, SHI-1 cells without any treatment were used as control. Cells in different groups were collected at 24, 48 and 72 h after incubation for further detection. MTT method was used to assay the growth inhibition rate and flow cytometry was used to detect the early stage apoptosis ratio. The human PI3K-AKT Signaling Pathway RT(2) Profiler(TM) PCR Array was used to detect the expression of 84 genes involved in PI3K-AKT signaling. The results indicated that the SV inhibited the proliferation and inducted the apoptosis of SHI-1 cells in time- and dose-dependent manners significantly. The growth inhibition rates of SHI-1 cells treated with 15 µmol/L SV for 24, 48 and 72 h were 26.82, 47.09 and 63.92, respectively; and their early stage apoptosis ratios were 5.75, 13.25 and 15.59, respectively. Compared with the control group, expression levels of 39 genes were changed in the group of 15 µmol/L SV at 48 h, among them 26 genes were down-regulated and 13 genes were up-regulated. It is concluded that the SV can inhibit proliferation and induce apoptosis of SHI-1 cells, and the mechanism may be associated with the changes of gene expression level in PI3K-AKT signaling pathway regulated by SV.
Apoptosis ; drug effects ; Cell Line, Tumor ; Cell Proliferation ; drug effects ; Down-Regulation ; Gene Expression Regulation, Leukemic ; Humans ; Leukemia, Monocytic, Acute ; metabolism ; Phosphatidylinositol 3-Kinases ; metabolism ; Proto-Oncogene Proteins c-akt ; metabolism ; Signal Transduction ; drug effects ; Simvastatin ; pharmacology

OBJECTIVETo assess the prevalence of several tyrosine kinases (TKs) gene mutations including c-Kit, FLT3 and JAK2 V617F in core binding factor related acute myeloid leukemia (CBF-AML), and analyze their impact on clinical characteristics and prognosis.

METHODSMutations of c-Kit, FLT3-ITD and FLT3-TKD were detected by genomic DNA PCR and sequencing, and JAK2 V617F mutation screening by allele-specific PCR in 58 newly diagnosed CBF-AML patients [28 AML with inv(16) and 30 with t(8;21)], and analyze the patients clinical characteristics and prognoses.

RESULTSc-Kit aberrations were detected in 32.8% cases, including 6 cases mutated in exon 8 (mutKIT8) and 13 mutated in exon 17 (mutKIT17). MutKIT8 was more prominent in inv(16) than in t(8;21) patients (21.4% vs 0, P = 0.009). Only 2 cases had FLT3-ITD and 7 (12.1%) FLT3-TKD mutations. The result of JAK2 V617F mutation screenings in these CBF-AML patients was negative. The frequency of receptor tyrosine kinases(RTK) mutations was 46.6% and only one case had two kinds of missense mutations (mutKIT8 & TKD(+)). Median age of onset was higher for mutKIT17 than for wide-type c-Kit (wtKIT) patients (55 vs 31, P = 0.003). c-Kit mutations were significantly associated with decreased overall survival (OS) and continuous complete remission (CCR) rates (P = 0.053, and 0.048 respectively), and so did more for exon17 mutated patients reduced (P = 0.005, and 0.013 respectively). FLT3-TKD mutation showed no effects on prognosis of CBF-AML patients.

CONCLUSIONSRTK mutations are common in patients with CBF-AML. c-Kit mutations frequently and JAK2V617F mutation rarely appear in CBF-AML. c-Kit mutations, especially mutKIT17 confers higher relapse risk and poorer prognosis.

Adolescent ; Adult ; Aged ; Core Binding Factors ; DNA Mutational Analysis ; Female ; Humans ; Janus Kinase 2 ; genetics ; Leukemia, Myeloid, Acute ; diagnosis ; etiology ; genetics ; Male ; Middle Aged ; Mutation ; Prognosis ; Protein-Tyrosine Kinases ; genetics ; Proto-Oncogene Proteins c-kit ; genetics ; Young Adult ; fms-Like Tyrosine Kinase 3 ; genetics

To investigate the adjuvant effect of granulocyte macrophage colony stimulating factor (GM-CSF) in Flaviviridae virus DNA vaccines. After DNA immunization, the antibody levels of serum from mice were detected by ELISA and indirect immunofluorescence assay. Co-immunization of GM-CSF suppressed the immune responses induced by DV1 and DV2 candidate vaccines whereas enhanced the immune response induced by HCV C and E1 DNA vaccines. As genetic adjuvant for DNA vaccines, GM-CSF might display complex diversity on the immune responses: an augmentation or suppression due to different immunogens. Therefore, GM-CSF should be used with some cautions in clinic.
Adjuvants, Immunologic ; administration & dosage ; Animals ; Antibodies, Viral ; immunology ; DNA, Viral ; administration & dosage ; genetics ; immunology ; Dengue ; immunology ; prevention & control ; virology ; Dengue Vaccines ; administration & dosage ; genetics ; immunology ; Dengue Virus ; genetics ; immunology ; Female ; Granulocyte-Macrophage Colony-Stimulating Factor ; administration & dosage ; immunology ; Hepacivirus ; genetics ; immunology ; Hepatitis C ; immunology ; prevention & control ; virology ; Humans ; Immunization ; Mice ; Mice, Inbred BALB C ; Vaccines, DNA ; administration & dosage ; genetics ; immunology ; Viral Vaccines ; administration & dosage ; genetics ; immunology

Objective To investigate the influence of calcium elevation on oxida tive stress in human lens epithelial cells (HLEC) SRA01/04.Method The cells (2 x 103 cells/well) which in the period of logarithmic phase were seeded into 96-well plates with three replicates for the two groups;and in the experimental group,SRA01/04 cells were exposed to a CaCI2 concentration gradient (3 mmol · L-1,5 mmol · L-1,7 mmol · L-1,9 mmol · L-1,11 mmol · L-1,13 mmol · L-1,15 mmol · L-1,17 mmol · L-1,19 mmol · L-1) for 0 h,12 h,24 h,36 h;while the cells in the control group were cultured in complete 1640 medium.Cell counting kit-8 (CCK-8) assay was used to measure cell viability.The levels of intracellular superoxide dismutase (SOD),glutathione (GSH) content and oxidized glutathione (GSSG) / total glutathione (T-GSH) were determined by using the microplate-reader method with the commercial total/oxidized glutathione and sod quantification kit.Results At first,the survival rate of SRA01/04 cells treated with 3 mmol · L-1,5 mmol · L-1,7 mmol· L-1 CaCL2 for 24 h showed a significant decrease with the increase of CaCl2 concentration by CCK-8 assays,but gradually increased when the concentration increased to 9 mmol · L-1,and the difference approached statistical significance (P ＜ 0.05).Meanwhile,there was significant difference in the viability of the control group (0.592 + 0.055) and cells exposed to 15 mmol · L-1 CaCI2 (0.293 + 0.02) (t =7.811,P ＜0.05).Cell treatment with 15 mmol· L-1 CaC12 for 24 h was the most appropriate condition for HLEC apoptosis,followed by the appearance of nuclear fragmentation and dissolution,enhanced intracellular SOD viability (t =-6.417,P ＜ 0.05),decreased T-GSH content (t =13.816,P ＜ 0.05),and increased ratio of GSSG/T-GSH (t =-4.396,P ＜ 0.05) when compared with the control group,and the differences were statistically significant.Conclusion Intracellular calcium elevation can inhibit the cell viability and increase the levels of SOD and GSSG in HLEC to aggravate the intracellular oxidative damage.

Objective: To evaluate the safety and clinical efficiency of holmium laser enucleation of the prostate (HoLEP) in the treatment of small-volume BPH (SBPH) complicated by severe lower urinary tract symptoms (LUTS). METHODS: We retrospectively analyzed the clinical data on 82 cases of SBPH with severe LUTS treated by HoLEP from January 2017 to December 2018. The patients were aged (65.5 ± 7.6) years, with a mean prostate volume of <40 ml, a total IPSS of 24.8 ± 4.6, a QOL score of 5.2 ± 0.8, the maximum urinary flow rate (Qmax) of (7.6 ± 3.7) ml/s, and a mean PSA level of (1.8 ± 1.4) μg/L. RESULTS: All the operations were successfully completed, the mean operation time averaging (30.2 ± 5.0) min, enucleation time (26.7 ± 5.6) min and comminution time (3.5 ± 1.1) min, and the enucleated tissue weighing (20.3 ± 4.9) g. After surgery, the bladders were irrigated for (3.5 ± 1.9) h, with (3.0 ± 1.7) L of rinse solution, and catheterization lasted (24.8 ± 9.7) h. Histopathology revealed moderate or severe lymphocytic infiltration in 69 cases (84.1%). At 6 months after operation, significant improvement was observed in the IPSS, QOL, Qmax and PSA level compared with the baseline (P < 0.05). To date, no urethral stricture-related reoperation was ever necessitated. CONCLUSIONS HoLEP is safe and effective for the treatment of SBPH complicated by severe LUTS and can be employed after adequate preoperative evaluation of the patient.《.
Humans ; Lasers, Solid-State ; Lower Urinary Tract Symptoms/surgery* ; Male ; Prostate/surgery* ; Prostatic Hyperplasia/surgery* ; Quality of Life ; Retrospective Studies

Hydrogen peroxide (H

【Objective】To explore the effects of overexpressed senescence marker protein 30 （SMP30） on cell proliferation and antioxidative activity in human lens epithelial cell（HLEC）line SRA01/04 under high-calcium mediated oxidative stress. 【Methods】There were 3 groups in this experiment：SMP30 overexpressed group （OE，experimental group），NCOE group （negative control group） and SRA01/04 group （blank control group）. OE and NCOE lentiviral vectors were used to transfect SRA01/04 respectively. A high- calcium- mediated- stress cell model was established by culturing cells with medium containing 15 mmol/L CaCl2 for 24 h. BrdU assay was used to measure cell proliferation. SOD assay kit and GSSG/T- GSH assay kit were used to detect the level of intracellular oxidative stress. 【Results】Green fluorescence protein could be observed in all transfected cell groups under fluorescence microscope and the transfection efficiency was close to 80% ，suggesting that OE cell model was constructed successfully. Under the high calcium culture conditions，the activity of relative cell proliferation and SOD in OE group［（3.89 ± 0.20）and（47.5 ± 4.3 U/mg）］were significantly higher than that in NCOE group［（2.82 ± 0.34）and（30.6 ± 4.2 U/mg）］and SRA01/04 group［（2.96 ± 0.25）and（26.8 ± 1.5 U/mg）］，the ratio of GSSG/T-GSH in OE group（2.36 ± 0.51）was significantly lower than that in NCOE group（16.36 ± 2.48）and SRA01/04 group（20.12 ± 2.54）（n=3，P<0.05）；there was no significant difference between NCOE group and SRA01/04 group （n=3，P>0.05）. 【Conclusions】Overexpression of SMP30 increased the activity of cell proliferation and SOD，but decreased the ratio of GSSG/T- GSH in SRA01/04 cell（HLEC），indicating that SMP30 may alleviate the progression of high-calcium-mediated oxidative cell damage and possess the cytoprotective functions in HLEC.