OBJECTIVE:To study the restraining factors in the intravenous immune globulin(IVIG.)bacterial endotoxin test.METHODS:A series tests were designed;the sensitivity of limulus was checked and the limit of endotoxin was validated;the effects of vehicle and pH value on the IVIG endotoxin test results were observed so as to find out the chief restraining factors of this test method.RESULTS:Lower pH value has an inhibition effect on the in IVIG bacterial endotoxin test.CONCLUSION:Bacterial endotoxin test can replace the rabbit testing method in the pyrogen test of IVIG.When the condition of lower pH is changed.

Objective To investigate the effect of Alzheimer' s beta-amyloid (Aβ) on the production and the translocation in cytoplasm of retinoid receptor-α (RXRα). MethodsN2awt cells were treated with Aβ peptide or amyloid protein precursor(APP695) transfection. The nucleus were separated from the cytoplasm by kit. The quantity of RXRα in the nucleus and cytoplasm was detected by Westernblot. The translocation of RXRα in the nucleus and cytoplasm of above N2awt cells or of the cortex cells in the brains of Alzheimer' s disease (AD) patients and their normal control groups was observed by immune fluorescence. ResultsIn N2awt cells, the increasing APP or Aβ had no significant effect on the production of RXRα but resulted in RXRα exporting into cytoplasm, the ratio of RXRα in cytoplasm increased from 3.2％ (in control group) to 17.6％ (in APP+ group) and from 3.8％ (in control group) to 14.3％ (in Aβ + group) respectively; compared with normal cortex cells, the translocation of RXRα in the cytoplasm of the cortex cells in the brains of AD increased significantly. ConclusionAβ may affect RXRα exporting into cytoplasm.

Objective To investigate the effect of down-regulation of lysine specific demethylase 1 (LSD1) by shRNA on the apoptosis and cell cycle of human acute myelogenous leukemia cells.Methods The lentiviral vector-mediated LSD1-shRNA was transfected into human acute promyelocytic leukemia HL-60 cells and acute monocytic leukemia SHI-1 cells.The expressions of LSD1 mRNA and protein were examined by real time quantitative PCR and Western blot,respectively.The flow cytometry was applied to detect the apoptosis and cell cycle distribution after AnnexinV-PE/7-AAD and PI dying,respectively.Results The expressions of LSD1 mRNA and protein in HL-60 and SHI-1 LSD1-shRNA group were significantly decreased compared with the blank control group and the negative shRNA group (P ＜ 0.01,respectively).The apoptosis levels of HL-60 and SHI-1 cells were significantly increased after knockdown of LSD1 (P ＜ 0.01).Moreover,the cell cycle distribution in the G0/G1 phases was also significantly increased(P ＜ 0.01).Conclusion LSDI-shRNA promotes cell apoptosis and increases the percentage of cells in the G0/G1 phases of human acute myelogenous leukemia cells.

Small interfering ribonucleic acid (siRNA)-induced RNA degradation can inhibit viral infection, and has been investigated extensively for its efficacy as antiviral therapy. The potential therapeutic role of lentiviral-mediated short hairpin ribonucleic acid (shRNA) to Newcastle disease virus (NDV) replication in vivo has been explored less often. We constructed two recombinant lentiviral vectors containing shRNA against the phosphoprotein (P) of the NDV, RNAi-341 and RNAi-671. Recombinant shRNA lentivirus vectors were co-transfected into 293T cells, along with helper plasmids, to package the recombinant shRNA lentivirus. Lentivirus-based shRNAs were titrated and transduced into NDV-susceptible chicken embryo fibroblasts (CEFs) and chick embryos. Antiviral activity against the NDV strain was evaluated by virus titration and real-time reverse transcription-polymerase chain reaction. RNAi-341 and RNAi-671 strongly suppressed transient expression of a FLAG-tagged P fusion protein in 293T cells. RNAi-341 and RNAi-671 NDV reduced virus titers by 66.6-fold and 30.6-fold, respectively, in CEFs 16 h after infection. RNAi-341 and RNAi-671 reduced virus titers in specific pathogen-free chick embryos by 99% and 98%, respectively, 48 h after infection. Both shRNAs inhibited accumulation of not only P-gene mRNA, but also nucleocapsid, M-, F-, HN-, and L-gene mRNA. RNAi-341 silenced P-gene mRNA more potently than RNAi-671. These results suggest that shRNAs silencing the P gene had substantial antiviral properties and inhibited NDV replication in CEFs and chick embryos.
Animals ; Chick Embryo ; Chickens ; Down-Regulation ; Fibroblasts ; virology ; Gene Targeting ; Lentivirus ; genetics ; metabolism ; Newcastle Disease ; virology ; Newcastle disease virus ; genetics ; physiology ; Phosphoproteins ; genetics ; metabolism ; Poultry Diseases ; virology ; RNA Interference ; RNA, Small Interfering ; genetics ; metabolism ; Viral Proteins ; genetics ; metabolism ; Virus Replication

Objective To investigate the effects of shRNA interference (RNAi) targeting the histone lysine specific demethylase 1 (LSD1) on the proliferation and apoptosis in acute leukemia (AL) cells. Methods LSD1 shRNA vectors were constructed and transduced into HL-60 and SHI-1 AL cell lines. Inhibitory efficiency of LSD1 was detected by real-time quantitative PCR (RT-qPCR) and Western blot respectively. Cell proliferation was assessed by methyl thiazolyl tetrazolium (MTT), and cell apoptosis was measured by flow cytometry. Results After interference of LSD1, the expression levels of LSD1 mRNA and protein in HL-60 and SHI-1 cells (mRNA: 0.242 ±0.023, 0.207 ±0.006; Protein: 0.256 ±0.015, 0.486 ±0.042) were decreased compared with blank control group without transfection process (mRNA: 1.021 ±0.178, 1.039 ±0.395; Protein:0.552 ±0.026, 0.754 ±0.060) and empty vector negative control group (shNC group) (mRNA: 0.935 ±0.136, 1.016±0.203;Protein: 0.500±0.026, 0.750±0.049) (P<0.05). The levels of their cell proliferation (absorbance value: 0.712±0.010, 0.549±0.007) were inhibited compared with blank control group (absorbance value:0.823±0.010, 0.625±0.005) and shNC group(absorbance value: 0.818±0.019, 0.621±0.003) (P< 0.05). While cell apoptosis rates were increased [(32.80 ±1.35) %, (23.49 ±1.40) %] compared with blank control group [(8.08 ±0.62) %, (7.28 ±1.17) %] and shNC group [(8.08 ±0.62) %, (7.28 ±1.17) %] (P< 0.05). Conclusions Lentivirus-mediated shRNA interferencing LSD1 can inhibit cells ' proliferation and promote apoptosis of HL-60 and SHI-1 AL cell lines, indicating that LSD1 may be a potential biological molecular marker and a new treatment target for AL.

Aim Toclarifytheeffectofacteosideon proliferation of neural stem cells (NSCs ) from adult mice,as well as the involved signaling pathway.Meth-ods NSCswereisolatedfromthesubventricularzone (SVZ)of adult C57BL/6 mice,then identified by im-munofluorescence staining with Nestin,the marker of NSCs.NSCs were exposed to acteoside (5,10,20,40μmol·L-1 )in absence of mitogen(EGF/bFGF)for 24 h.We employed CCK8 assay to detect NSCs viability and BrdU staining to identify NSCs proliferation.We performed Western blot to quantify the expression level ofp-AktinducedbyacteosideonNSCs.Results With-out mitogen,acteoside increased NSCs proliferation by activating p-Akt,which can be blocked by LY294002, the inhibitor of PI3K/AKT signaling pathway.Conclu-sion ActeosidepromotestheproliferationofNSCsfrom adult mice by activating PI3K/AKT pathway.

Objective To systematically review the evidence for the effect of vitamin D supplementation on insulin resistance and blood glucose in people who are obese or people with abnormal glucose metabolism. Methods We searched databases including Pubmed, Elsevier, Web of Science, and WANFANG Database etc. for randomized controlled trials comparing vitamin D or analogues with placebo. We extracted data on insulin resistance and blood glucose, including homeostasis model assessment of insulin resistance( HOMA-IR), fasting blood glucose, HbA1C (% ). A1l data were analyzed using Review Manager 5. 0. Results Nine studies involving 867 participants were included. The results of meta-analysis showed that: ( 1 ) For people who are obese and with abnormal glucose metabolism, meta-analysis showed a small improvement in HOMA-IR(SMD -0. 34,95% CI -0. 61 to -0. 06, P<0. 05) and a small effect on fasting glucose (SMD -0. 41 mmol/ L, 95% CI -0. 68 to -0. 15, P<0. 05),while such effects were not seen in people who are obese but with normal blood glucose. (2) No serious adverse events were associated with the administration of vitamin D. Conclusion vitamin D supplementation may be benefit for improving insulin resistance and fasting blood glucose in people who are obese and with abnormal glucose metabolism, but has no effect on obese people with normal blood glucose.

Objective To investigate the effect and mechanism of cell differentiation agent Ⅱ (CDA-Ⅱ) on the differentiation of human acute mycloid leukemia (APL) HL-60 cells.Methods The cell morphology and differentiation was detected by Wright-Giemsa staining,the expression of cell surface differentiation antigen CD11b of HL-60 was detected by flow cytometry,the differentially expressed genes were screened by gene expression profile chip (NimbleGen).Results The result of Wright-Giemsa staining showed that CDA-Ⅱ induced HL-60 differentiation towards mature stages in a time-dependent manner.After treated with CDA-Ⅱ,the percentage of CD11b-positive HL-60 cells significantly increased in a time-and dose-dependent manner.The result of gene expression profiling indicated differentially expressed genes including 113 up-regulation genes and 140 down-regulation genes.The up-regulation expression genes involved in six pathways including mineral absorption,complement and coagulation cascades,down-regulation expression genes involved in nine pathways including pyrimidine metabolism,RNA polymerase,purine metabolism and so on.Conclusion CDA-Ⅱ can induce HL-60 differentiation and make gene differentially expressed.The data provide the feasibility of CDA-Ⅱ in differentiation induction therapy for APL.

ObjectiveTo investigate the effect of artesunate on the expression of vascular endothlial growth factors(VEGF)and VEGFR in SHI-1 cell line.MethodsEnzyme-linked immunosorbent assay analysis was performed to detect the amount of VEGF in culture supernatants of SHI-1 cell in the condition of artesunate or not. The expression of VEGFR1 and VEGFR2 in SHI-1 cell in the condition of artesunate or not were detected by flow cytometry.ResultsWithout artesunate,the concentration of VEGF in the culture supernatant of SHI-1 cell were (980.3±2.2) pg/ml in 24 h and (982.4±2.3) pg/ml in 48 h. The expression of VEGFRI in SHI-1 cell were (6.40±3.11) ％ in 24 h and (6.45±2.85) ％ in 48 h. The expression of VEGFR2 in SHI-1 cell were (13.90±2.26) ％ in 24 h and (13.95±1.96) ％ in 48 h. With artesunate at 5, 10, 20 ng/ml, the concentration of VEGF in culture supematant of SHI-1 cell were (234.6±1.8)pg/ml, (114.9±1.6)pg/ml, (108.8±1.5) pg/ml in 24 h and (62.3±1.7) pg/ml, (60.9±1.6) pg/ml, (32.7±1.7) pg/ml in 48 h, respectively. The levels of VEGF in SHI-1 cells treated with artesunate at different concentrations decreased significantly (P ＜0.05).There was significant difference between 24 hours group and 48 hours group(P ＜0.05).The expression of VEGFR1 in SHI-1 cell were (4.30±2.21) ％, (4.20±1.37) ％, (3.90±1.86) ％ in 24 h and (3.80±2.87) ％, (3.60±1.73) ％, (3.00±1.82) ％ in 48 h, respectively. The expression of VEGFR1 in SHI-1 cell treated with artesunate at different concentrations were not significantly different (P ＞0.05). No significant difference between 24 hours group and 48 hours group was observed (P ＞0.05). VEGFR2 expression of SHI-1 cell were(4.40±1.15) ％, (3.10±0.68) ％, (1.10±0.72) ％ in 24 h and (3.00±1.68) ％, (2.20±0.93) ％, (0.60±0.92) ％ in 48 h, respectively. The results indicated that the expression of VEGFR2 in SHI-1 cells treated with artesunate at different concentrations reduced significantly (P ＜0.05),but there was no significant difference between 24 h group and 48 h group (P ＞0.05). ConclusionThe concentration of VEGF in SHI-1 cell was high, and artesunate can down-regulate the expression of VEGF and VEGFR2,but the effect of artesunate on the VEGFR1 was not significant.

Objective To observe the changes of neural blood flow when oppressed with different pressures and at different sites in rat's sciatic nerve.MethodsThe sciatic nerve compression model of rat with pressure and time controlled simultaneously was established.The changes of neural blood flow were observed by oppressing the sciatic nerve with five grades pressures using Gasbag.The rats were randomly divided into the distal compression group and proximal compression group,and the changes of neural blood flow were observed in each group.ResultsThe neural blood flow changed significantly(P<0.05) when the sciatic nerve was oppressed with different pressures and it had the decreasing tendency with the pressures increasing.The neural blood flow of each group decreased obviously(P<0.01).The effect of oppressing distal nerve on neural blood flow was more obvious than that of oppressing proximal nerve(P<0.01).ConclusionMechanical compression can influence the neural blood flow obviously.The distal vessels of sciatic nerve are the major sources of nerve blood supply.