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OBJECTIVE To study the toxicity of zinc oxide nanoparticles (ZnO-NPs) on murine macrophage Ana-1 cells and the mechanism.METHODS Ana-1 cells were incubated with ZnO-NP (2.5-160 mg· L-1).Cell viability was investigated by MTT assay.The integrity of cell membrane was investigated by acridine orange-ethidium bromide (AO-EB) staining.The intracellular uptake of ZnO-NP and the percentage of sub-G1 of Ana-1 cells were detected by flow cytometry.Zinc ions were determined by fluorescent probe.The change of cell viability was studied after chelating zinc ions with ethylene diamine tetraacetic acid (EDTA).RESULTS ZnO-NP 2.5,5,10 and 20 mg· L-1 decreased cell viability of Ana-1 cells (r=0.905,P＜0.05) in a concentration-dependent manner.The cell viability was decreased to 27.9％ after exposure to ZnO-NP 20 mg· L-1.Intracellular uptake of ZnO-NP was increased after Ana-1 cell incubated with ZnO-NP at concentrations ranging from 40 to 160 mg· L-1 (P＜0.05).There were obvious free zinc ions in the cells.EDTA 2.5 mmol· L-1 significantly increased the cell viability decreased by ZnO-NP 20 mg· L-1 (P＜0.05).Chelating free zinc ions significantly mitigated ZnO-NP induced cell toxicity (P＜0.05).CONCLUSION Cytotoxicity and apoptosis of Ana-1 cells induced by ZnO-NP might be related to intracellular uptake of ZnO-NP and release of zinc ions.
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Objective:To reveal the dynamic characteristics of the intracellular complement mRNA from mouse macrophage ANA-1 treated with LPS or IL-4. Methods:The polarization models of macrophage ANA-1 were established by treating with LPS(1μg/ml) and IL-4(20 ng/ml),respectively. After treating at 3,8,12 and 24 h,the total RNA were abstracted by Trizol lysis methods . The macrophage polarization were estimated by the expression of IL-1β, CCL2 and Arg-1 mRNA detected by Real-time fluorescent quantitative PCR. The intracellular complement C1q, C3, CfB and CRIg mRNA were quantitatively analyzed. Results: The mouse macrophage ANA-1 cells treated with LPS was polarized to M1 since the levels of IL-1β and CCL2 mRNA were up-regulated significantly,in which their 2-△△Ct value were up to 297. 0±31. 0 and 19. 9±3. 3 respectively at 12 h. On the other hand,the ANA-1 cells treated with IL-4 was polarized to M2 because the level of Arg-1 mRNA was obviously higher( the 2-△△Ct value of Arg-1 mRNA was up to 27.3±9.1 at 24 h)(P<0.05).The intracellular complement C1q,C3,CfB and CRIg mRNAs all were up-regulated in different polarized macrophages. The intracellular C1q and C3 mRNA in polarized M2 were significantly higher,in which the peak value of C1q and C3 were to 94. 9±12. 9 and 11. 3±2. 4 at 12 h,respectively(P<0. 05). Reversely,the CfB mRNA in polarized M1 increased obviously,in which its 2-△△Ct was to 61. 4±6. 2 at 12 h. In addition,the CRIg mRNA in both groups was only up-regulated at 24 h,in which the 2-△△Ct value was 6. 5±1. 8 in M1 and 10. 8±3. 2 in M2(P<0. 05). Conclusion: The macrophage ANA-1 cell polarization models were successfully established by treated with LPS or IL-4. The intracellular complement C1q,C3 and CRIg mRNA in polarized M2 were transcripted more than in M1. But the intracellular CfB mRNA in polarized M1 was up-regulated significantly. These results suggested that the dynamic characteristic of complement components in different polarized macrophage would be correlated with its fun-tions.
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Objective To study the protective effect of propofol on lipopolysaccharide (LPS)-induced Ana-1 cell damage.Methods The LPS-induced inflammation damage of Ana-1 cells was es-tablished as a model.Ana-1 cells were divided into control group(group C),LPS group(group L), LPS+propofol 10μmol/L group(group LP1),LPS+propofol 20μmol/L group(group LP2),LPS+propofol 40 μmol/L group(group LP3),and LPS+propofol 80 μmol/L group(group LP4),respec-tively.Cell survival rate was assessed using MTT,interleukin 1βand 6,and tumor necrosis factors (TNF-α)were determined.Results Survival rate of Ana-1 cells in group L was lower than that in group C,while it was significantly improved in group LP1,LP2,LP3,LP4 in a dose-dependent man-ner (P <0.01);IL-1β,IL-6,TNF-αrelease was significantly higher in group L than in group C;IL-1β,IL-6,TNF-αrelease significantly reduced in group LP1,LP2,LP3,LP4 (P <0.01)in a dose-dependent manner (P <0.01).Within the selected dose range in this experiment,group LP4 had the best protection effect.Conclusion Propofol can reduce inflammation damage of Ana-1 cells caused by LPS.
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Objective:To observe the effect of different concentration HMGB1 on the secretion of TNF-αand NO from Ana-1 infected with DEN2 and virus copy.Methods:DEN2 were proliferated and identified by conventional methods.The adherence of DEN2 to Ana-1 was observed by direct immunofluorescence and RT-PCR.The level of virus mRNA were detected by qRT-PCR.The concentration of TNF-αwas detected by ELISA.The concentration of NO was detected with Griess reagent.Results:Ana-1 was able to adhered for DEN2.Compared with DEN group,the inhibition ratio(%) of the level of virus mRNA in D-HMGB1-1 group,D-HMGB1-10 group,D-HMGB1-100 group,D-HMGB1-1000 group was 41.53 ±2.12,55.30 ±1.59,74.75 ±1.12,86.35 ±1.42.Compared with DEN group,the level of TNF-αand NO decreased in D-HMGB1 groups(P<0.05).Conclusion:HMGB1 can be effectively regulated of Ana-1 secreted inflammation factor of infected with DEN2,and inhibited DEN2 replication.