| Objective: Endotoxin sepsis is a systemic inflammatory response syndrome caused by microbial infection,which has high mortality rate and it's an important cause of death in the intensive care unit.Endotoxin is protein complex that lipopolysaccharide(LPS)that the cell wall of Gram-negative bacteria and a trace protein,and has been considered as the main substance for inducing sepsis.Radix Isatidis,a traditional Chinese medicine,has the effects of clearing away heat and detoxifying,cooling blood and pharynx,and is widely used in clinical practice for anti-inflammatory,anti-viral,anti-bacterial and antipyretic.In recently years,the prevention and treatment of clearing heat and detoxifying herbs in endotoxin sepsis and the intrinsic mechanism has been gradually discovered.According to previous research reports,chloroform extract and methanol extract of R.Isatidis have the effectof anti-endotoxin,while in clinical application,the pharmacodynamic material basis of R.Isatidis are water-soluble components.Therefore,this study aimed to study the anti-endotoxin septic effect of the water extract of R.Isatidis and explore possible pharmacological molecular mechanism.Methods: Human cervical cancer He La cells and mouse peritoneal mononuclear macrophages RAW264.7 were used to study the in vitro activity and pharmacodynamic mechanism of the water extract of R.Isatidis.In vivo experiments,the protective effect of the water extract of R.Isatidis on the LPS-induced endotoxin-shock model mice was evaluated.In the experiment,the water extract of R.Isatidis was given the clinical equivalent dose,and the evaluated index was the survival rate(%)of mice.Real-time quantitative-polymerase chain reaction(q-PCR)and enzyme-linked immunosorbent assay(ELISA)were used to study the expression and release of nuclear factor-?B(NF-?B)downstream cytokine tumor necrosis factor-a(TNF-a)and interleukin-1?(IL-1?);expression and release of interferon TIR domain adaptor protein(TRIF)kinase downstream interferon regulatory factor 3(IRF3)regulatory gene interferon-?(IFN-?)and IFN-? induced genes.Results: The water extract of R.Isatidis significantly improved the survival rate of LPS-induced endotoxin-septic mice: all mice in the LPS model group died within 2 days;the survival rate of the water extract of R.Isatidis prevention group reached 80%,and it can effectively retarded the body temperature decline trend of LPS-induced endotoxin mice;reduced the liver inflammatory cell infiltration and lung tissue damage,and significantly inhibit the overexpression of inflammatory cytokines Nos2 and IL-6 m RNA in liver and lung tissues.The results of mechanism study showed that the water extract of R.Isatidis had inhibitory effects on the expression of TNF-a and IL-1? m RNA only at 4 h,but no significant inhibition at 1 h and 8 h.By measuring the expression and release of IRF3 downstream gene IFN-? and the expression of its induced genes at different time points,the results showed that the water extract of R.Isatidis significantly inhibited the m RNA expression and protein release of IFN-? at different time points.And the m RNA expression of IFN-?-induced genes CXCL9,CXCL10,CXCL11 were significantly inhibited.In addition.In vitro experiments,the water extract of R.Isatidis effect on NF-?B active and gene expression of LPS-induced RAW264.9 cells were consistent with in vivo experiments.Conclusion: The water extract of R.Isatidis has protective effect on LPS-induced endotoxin mice,which regulates the transcriptional level and release of IFN-?,a downstream cytokine of IRF3,blocks the IFN-?/STAT interferon signaling pathway,and inhibits the expression of IFN-?-induced inflammatory cytokines CXCL9,CXCL10,and CXCL11,thereby protecting the organ damage of mice and reduces the body damage caused by endotoxin.This study reveals the anti-endotoxin effect of R.Isatidis and the mechanism of clearing away heat and toxic materials,which has great significance for the clinical drug guidance of R.Isatidis. |
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