OBJECTIVETo study the protective effect of sufentanil preconditioning against myocardial ischemia-reperfusion (I/R) injury and the role of PI3K/Akt signaling pathway.

METHODSSixty male SD rats weighing 250-350 g were randomly divided into 5 equal groups, namely the sham-operated group, I/R group, sufentanil preconditioning group (Spc group), sufentanil preconditioning +PI3K inhibitor group (Spc+W group), and PI3K inhibitor group (W group). Myocardial I/R model was established by ligation of the anterior descending branch of the left coronary artery for 30 min followed by reperfusion for 120 min. Sufentanil was administered in 3 doses via the femoral vein before the occlusion, each at 1 µg/kg infused within 5 min at a 5-min interval. In Spc+W and W groups, PI3K inhibitor wortmannin (15 µg/kg) was given intravenously 5 min before sufentanil preconditioning and 35 min before ischemia, respectively. Heart rate and mean arterial pressure (MAP) were continuously monitored during I/R. At the end of reperfusion, blood samples were obtained to determine plasma activation of CK-MB and LDH. Acute infarct size was measured by triphenyltetrazolium chloride staining, and the myocardial tissues were obtained to detect the expression of phosphorylated Akt using Western blotting.

RESULTSPhosphorylated Akt expression was significantly up-regulated in I/R and Spc groups as compared with the sham group, and was significantly higher in Spc group than in I/R group. After reperfusion, sufentanil preconditioning significantly decreased myocardial infarct size (P<0.01) and lowered the levels of CK-MB (P<0.01) and LDH (P<0.01) compared with those in the I/R group. The I/R , Spc+W and W groups showed no significant differences in myocardial infarct size or the levels of CK-MB and LDH.

CONCLUSIONThe protective effect of sufentanil preconditioning against myocardium against I/R injury in rats may involve PI3K/Akt signaling pathway activation.

Animals ; Ischemic Preconditioning, Myocardial ; methods ; Male ; Myocardial Reperfusion Injury ; metabolism ; prevention & control ; Proto-Oncogene Proteins c-akt ; metabolism ; Rats ; Signal Transduction ; Sufentanil ; pharmacology

Objective To compare the similarities and differences of the dental pulp cells in protein expression under different conditions and to investigate the effect of recombinant human interleukin-1β ( rhIL -1 β) on the production of protein by human dental pulp cells (HDPC) in vitro.Methods HDPC were induced with rhIL-1 β for 12 hours and the dental pulp cell entire protein was separated by a 2-DE technique.The rhIL-1β induction and the normal dental pulp cell protein 2-DE atlas were established.Difference expression protein was confirmed by Image Master 2D Elite 5.0 software analysis.Results In induced dental pulp cells,39 protein spots were obviously different,of which 15 points were up-regulated,13 were new protein spots,7 points were down-regulated,4 points expressed only in the control group.Conclusions Two-dimensional gel electrophoresis can not only show the whole protein of human dental pulp cells are likely to be involved in the process of dental pulp cell damage.

OBJECTIVETo investigate the prevalence and natural outcome of late-life depression in the community and to analyze the risk-prediction models.

METHODSA community in Hang Zhou was selected as a trial. A total of 1 275 persons aged 60 or more in this community were screened by PHQ-9 questionnaire; SCID was used for interviewer to diagnostic interview the people whose PHQ-9 was more than 10 points, 50 % of those whose PHQ-9 was from 5 to 9 points and 5 % of those whose PHQ-9 was less than 5 points, then all those who accepted diagnostically interview were interviewed by PHQ-9 every 3 months in one year, and were diagnostic interviewed by SCID in the last month. Logistic regression analysis was used to explore depressive risk factors in 12 months.

RESULTSThere were 141 (11.1%) persons whose PHQ-9 score was more than 10 points, 298 (23.4%) whose PHQ-9 score were 5-9 points, and 836 (65.5%) whose PHQ-9 score were 0 to 4 points in the preliminary survey, 93 were major depressive disorder (MDD). The prevalence of late-life depression was 7.3%. Compared with the PHQ-9 score in one year, 17.6% of those with no depressive symptoms emerged depression; 50% of those who had depressive symptoms declined, 9% developed to significant depressive symptoms, and 41% did not change; 12% of those with significant depressive symptoms were found no depression, 24% reduced, and 64% still had depression. The significant predictors were the accumulation of disease, social support, educational level, daily capacity and baseline of depression.

CONCLUSIONThe prevalence of late-life depression was high. The rates of recognition, diagnosis and treatment were low. The natural outcome after a year did not relieve apparently. Specialist-community health partnership management model is one of the important ways to prevent and treat late-life depression.

Aged ; Aged, 80 and over ; Depression ; diagnosis ; epidemiology ; etiology ; Female ; Follow-Up Studies ; Humans ; Logistic Models ; Male ; Mass Screening ; Middle Aged ; Prevalence ; Prognosis ; Risk Factors ; Surveys and Questionnaires

OBJECTIVETo construct a lentiviral vector for delivering short hairpin RNA (shRNA) targeting PAX6 and investigate its effect on the proliferation of glioma U251 cells in vitro.

METHODSTwo small interfering RNA sequences targeting PAX6 gene were designed based on the reported sequence of PAX6 and annealed to form a double?stranded chain, which was inserted into a lentiviral vector to construct the recombinant lentiviral vector shRNA?PAX6. The recombinant vector was infected into U251 cells, and the expression of PAX6 mRNA and protein in the cells was detected by real?time PCR and Western blotting, respectively. The changes in the proliferation of U251 cells after the infection was assessed using MTT assay.

RESULTSDouble enzyme digestion of the lentiviral vector pLKD?CMV?G&NR?U6?shRNA yielded an 8208?bp fragment, and colony PCR and sequencing analysis confirmed successful construction of the lentiviral vector shRNA?PAX6. Infection of the cells with shRNA?PAX6 caused a significant reduction of the expressions of PAX6 mRNA and protein (P<0.05) and resulted in obviously increased proliferation of U251 cells (P<0.05).

CONCLUSIONWe successfully constructed the recombinant vector shRNA?PAX6 for silencing PAX6 gene. PAX6 gene silencing results in increased proliferation of U251 cells in vitro.

OBJECTIVETo observe the direct regulation of miR-127 on Bcl-6 and the effect of Bcl-6 in rescuing miR-127-induced cell cycle and cell growth inhibition.

METHODSThe 3'UTR and coding region of human bcl-6 gene were amplified by PCR and cloned into pcDNA3.0-Luc and pcDNA3.0-Flag vectors, respectively. Mutations were introduced into the seed sequences of the predicted miR-127 target sites within the Bcl-6 3'UTR using recombinant PCR. Luciferase assay was used to verify the direct targeted regulation of miR-127 on Bcl-6. In HepG2 cell models with overexpression or knockdown of miR-12, the changes of cell cycle and cell growth were investigated after transfection with the constructed vectors.

RESULTSThe recombinant plasmids were successfully obtained as confirmed by double digestion and sequence identification. Luciferase assay showed that in 293T and HepG2 cells, miR-127 inhibited the activation of wild-type Bcl-6 3'UTR reporter vector but not mutated Bcl-6 3'UTR vector. Overexpression of miR-127 induced cell cycle arrest at G(2)/M phase and suppressed the growth of HepG2 cells, and these effects were reversed by Bcl-6 overexpression.

CONCLUSIONWe successfully cloned wild-type and mutated 3'UTR reporter vectors and expression vector of bcl-6 gene and confirmed their biological functions.

3' Untranslated Regions ; Cell Cycle ; Cell Proliferation ; DNA-Binding Proteins ; genetics ; Genes, Reporter ; Genetic Vectors ; Hep G2 Cells ; Humans ; Luciferases ; MicroRNAs ; genetics ; Proto-Oncogene Proteins c-bcl-6 ; Transfection

OBJECTIVETo investigate the effect of bear bile powder and ursodesxy cholic acid (UDCA) on peripheral blood, bone marrow megakaryocyte and immune organs in mouse model with thrombocytopenia, so as to provide a reference for studying the curative effects of bear bile powder and its succedaneum on thrombocytopenic purpura (TP).

METHODSThe mouse model with thrombocytopenia indued by cytosine arabinoside (Ara-C) was established, a total of 70 mice were randomly divided into normal group, model group, prednisone group, bear bile (middle and high dose) powder group and UDCA (middle and high dose) group. From the first day of making model mice in the each group, 0.4 ml/(20 g·d) corresponding drug was administered by infusion. At day 10 after treatment the peripheral blood, spleen and thymus organ index, the number of bone marrow megakaryocyte in each group were compared.

RESULTScompared with the normal group, the Plt, WBC and megakaryocyte counts in model group decreased, the spleen index increased obviously (P<0.05), but the WBC count returned to normal by 10 days; after treatment, compared with model group, the Plt, WBC and megakaryocyte counts of treated groups increased, spleen index decreased significantly (P<0.05), but the WBC count in prednisone group decreased, which in bear bile powder (high) group and UDCA (high) group were particularly significant.

CONCLUSIONThe bear bile powder and UDCA have been confirmed to have therapeutical effect on thrombocytopenia models induced by Ara-C, UDCA can substitute bear bile powder as a treatment drug for thrombocytopenic purpura.

Animals ; Bile ; Bone Marrow ; Bone Marrow Cells ; Cytarabine ; Disease Models, Animal ; Megakaryocytes ; Mice ; Spleen ; Thrombocytopenia

This study was aimed to elucidate the expression of costimulatory molecule CD80 and CD86 in HL-60 cells induced by proteasome inhibitor MG132 and its effect on allogeneic mixed lymphocyte reaction. Acute myelocytic leukemia cell line HL-60 and chronic myelocytic leukemia cell line K562 were cultured. The viability of the cells was measured by flow cytometry. Proteasome inhibitor MG132 at the concentrations of 2 or 3 µmol/L was used to stimulate the HL-60 cell cultured for 24 h and 48 h respectively, and the Annexin V/7-AAD staining and flow cytomotry were used to detect the apoptosis of the HL-60 cells. HL-60 and K562 cells were treated with 1 µmol/L MG132 for 24 h and 48 h respectively, then CD80 and CD86 antibodies were added, finally the expression of CD80 and CD86 was analysed by flow cytomery. The mRNA expression of CD86 in the HL-60 cells treated with 1 µmol/L MG132 was detected by RT-PCR. HL-60 and K562 cells were treated by 1 µmol/L MG132 and then underwent irradiation of 75 Gy (60)Co to kill the cells with their antigenicity preserved. Peripheral blood mononuclear cells (PBMNCs) of healthy volunteers, as reactive cells, were isolated and inoculated into the (60)Co irradiated HL-60 cells of different concentrations, as stimulating cells, CCK-8 was added and then the A value of absorbance was measured at the wave length of 450 nm in an enzyme labeling instrument. The results showed that the cell viability of the HL-60 cells treated with 1 µmol/L MG132 for 24 h an d 48 h was 92.95% and 85.87% respectively. The apoptotic rates of the HL-60 cells treated with MG132 increased in dose-and time-dependent manner. High-concentration of MG132 directly killed HL-60 cells. Before MG132 treatment K562 cells did not express CD86, but the CD86 expression of the HL-60 cells was up-regulated time-dependently after MG132 treatment (P < 0.01). The mRNA expression of CD86 in the HL-60 treated with MG132 was up-regulated time-dependently (P < 0.01). CCK-8 test showed that the proliferation level of PBMNC gradually increased along with the concentration of HL-60 cells treated with MG132 and reached its peak when the concentration of the HL-60 cells was 1×10(5) (P < 0.01). No remarkable proliferation of PBMNC was observed in the K562 groups no matter if the HL-60 cells had been treated with MG132. It is concluded that the high concentration of MG132 can directly kill HL-60 cells, low-concentration of MG132 can induce the expression of costimulatory molecule CD86 in HL-60 cells, also can improve the proliferation of PBMNC.
Apoptosis ; B7-2 Antigen ; immunology ; Cell Survival ; Flow Cytometry ; HL-60 Cells ; Humans ; K562 Cells ; Leukocytes, Mononuclear ; drug effects ; Leupeptins ; pharmacology ; Lymphocyte Culture Test, Mixed ; Proteasome Inhibitors ; pharmacology ; Reverse Transcriptase Polymerase Chain Reaction ; Up-Regulation