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Objective To observe the effect oflipopolysaccharide (LPS) on the cell form of BV-2 cells and the expressions of interleukin-6 (IL-6) and tumor necrosis factor-a (TNF-a) so as to detect the role of silence information regulator 1 (SIRT1) in regulation of LPS-induced proinflammatory cytokines production in activated BV-2 cells.Methods BV-2 cells were divided into control group (normal culture medium) and treatment groups; BV-2 cells in the treatment groups were subdivided into LPS treatment groups,Resveratrol+LPS treatment groups and Sirtinol+LPS treatment groups (cultured with different concentrations of LPS,SIRT1 activator Resveratrol or SIRT1 inhibitor Sirtinol,respectively).MTT assay was employed to identify the cell survival after the inducement.Based on the above MTT results,the cells were then grouped into the control group,LPS treatment group,Resveratrol+LPS treatment group and Sirtinol+LPS treatment group having suitable concentrations of LPS,Resveratrol and Sirtinol; then,the levels of IL-6 and TNF-a were measured with enzyme-linked immuno sorbent assay (ELISA) at 12 and 24 h after the inducement; and the expression of SIRT1 at 24 hafter the inducement was detected by Western blotting.Results As compared with those in the control group,the BV-2 cells in the LPS treatment group had increased cell number,hypertrophic cell body,and shorten cell processes.The cell survival rate increased with increased concentrations of LPS.As compared with those in the control group,the levels of IL-6 and TNF-a in LPS treatment group increased and level of SIRT 1 decreased with significant differences (P＜0.05).Significantly increased levels of IL-6and TNF-a and obviously decreased expression of SIRT1 in the Resveratrol+LPS treatment group were noted as compared with those in the LPS treatment group (P＜0.05); conversely,significantly decreased levels of IL-6 and TNF-a and obviously increased expression of SIRT1 in the Sirtinol+LPS treatment group were noted as compared with those in the LPS treatment group (P＜0.05).Conclusion LPS can change the morphology of BV-2 cells,and induce the levels ofproinflammatory cytokines; impairment of SIRT1 may contribute to such progress obviously.
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Objective To observe the effects of silent information regulator 1 (SIRT1) on toxicity of activated BV-2 to PC12 cells and the possible mechanisms.Methods BV-2 microglial cells and PC12 cells were routinely cultured in vitro; ELISA was used to measure to the levels of tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6) stimulated by lipopolysaccharide (LPS,1 μg/mL) in BV-2 cells.MTT assay was employed to identify the cell viability of PC12 cells injured by culture medium of activated BV-2 and determined the suitable concentrations of resveratrol (a potent SIRT1 activator,5,10,25,50 and 100 μmol/L) and nicotinamide (a known SIRT1 inhibitor,5,10,25 and 50 mmol/L).PC12 cells were divided into groups as follows:control group Ⅱ,LPS+BV-2 co-cultured group,resveratrol treatment group and nicotinamide treatment group (pretreated with resveratrol or nicotinamide for 2 h,and then subjected to culture medium of activated BV-2 cells,in the presence of resveratrol or sirtinol for 18 h); the cell viability was measured by OD value in MTT assay,and the expressions of SIRT1 and acetyl-p53 were detected by Western blotting.Results TNF-α and IL-6 secretions increased gradually at 6,12 and 24 h after LPS being induced BV-2,with significant difference between each two time points (P＜0.05).PC12 cell viability decreased in the LPS+BV-2 co-cultured group as compared with that in the control group Ⅰ,LPS treatment group and BV-2 supernate group (P＜0.05).The cell viability of cells in the 100 μmol/L resveratrol treatment group and 50 μmol/L niacinamide treatment group decreased as compared with that in the control group Ⅱ (P＜0.05),therefore,50 μmol/L resveratrol and 25 μmol/L nicotinamide were chosen in the next experiment.As compared with those in the control group Ⅲ,the cell viability and SIRT1 expression significantly decreased,and acetyl-p53expression significantly increased in the LPS+BV-2 co-cultured group (P＜0.05); as compared with those in the in the LPS+BV-2 co-cultured group,the cell viability and SIRT1 expression significantly increased,and acetyl-p53 expression significantly decreased in the 50 μmol/L resveratrol treatment group,and opposite results were noted in the 25 μmol/L nicotinamide treatment group (P＜0.05).Conclusion SIRT1 can inhibit toxicity of activated BV-2 to PC12 cells,the mechanism of which is partly via p53 activation.
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This paper introduces a kind of evaluation method in pipetting performance on new fully automated biochemistry analyzers by experiments. The performance of sample pipetting volume is confirmed by dye dilution method, the performance of reagent pipetting volume and dummy volume is done by weighing method. Meanwhile, researches and comparative researches on dummy volumes in different conditions have been made, providing valuable reference for clinical applications of automatic biochemistry analyzers.