Effect Of DcR3 On Therapeutic And Microbiota Of Spetic Mice

Decoy receptor 3(DcR3)is the only soluble receptor of the tumor necrosis factor receptor superfamily(TNFRSF),can regulating the inflammatory response.It is reported that DcR3 can be used for early diagnosis and monitoring of sepsis as a biomarker.Exogenous DcR3 and its analogues have the ability to fight injury in models of infection or inflammatory disease,such as reducing the accumulation of pro-inflammatory cytokines and neutrophils induced by Fas L in lung tissue.Sepsis as a life-threatening organ dysfunction caused by the host’s maladjusted response to infection,its characteristics is persistent excessive inflammation and immunosuppression.Effective control of excessive inflammation is the key way to improve survival rate,which is a scientific issue in sepsis treatment.The treatment of DcR3 in inflammatory diseases is widely reported,such as Rheumatoid arthritis(RA)and inflammatory bowel disease(IBD).However,the studies of DcR3 treatment in sepsis is limited.Therefore,exploring the anti-inflammatory mechanism of DcR3 in sepsis is helpful for the clinical treatment of sepsis.(1)In this study,DcR3 protein was expressed and purification in vitro through the construction DcR3 expression vector.The sepsis mouse model induced by Cecal Lpuncture(CLP)is used for this study,intraperitoneal injection of DcR3(1.0 mg/kg.d)significantly improved the adverse signs of mice with sepsis,such as chills,white eyes,huddle and low body temperature.DcR3 significantly increased the survival rate of mice with sepsis(P < 0.05);DcR3 reduced the expression levels of pro-inflammatory cytokines such as IL-1β、IL-6 and TNF-ɑ in serum(P < 0.05);DcR3 reduced the mRNA and protein levels of IL-1β、IL-6 and TNF-ɑ in heart、liver、spleen、lung and kidney tissues(P < 0.05).The HE results showed that DcR3 could improve the degree of inflammatory cell infiltration and pathological changes in lung,liver and heart tissue of sepsis mice(P <0.05);DcR3 can reduced the expression levels of lymphocytes,monocytes、neutrophils and white blood cell in peripheral blood of mice(P < 0.05);Flow cytometry showed that DcR3 can also down-regulated the expression levels of activated NK cells and B220cells(P < 0.05);RT-q PCR results showed that DcR3 reduced the mRNA expression levels of Elane、Ms4a3、Cd63、Ece2、Gan、Prtn3 and Timp1 in sepsis mice,and significantly increased Pik3c2 b gene mRNA expression level;Western blot(WB)results showed that DcR3 increased the protein expression levels of pro-caspase-3,pro-caspase-8 and cleaved caspase-8(P < 0.05),and decreased the protein expression levels of My D88、p-ikbɑ、cleaved caspase-3 and TNFSF10(P < 0.05).(2)To study how DcR3 effect enteric microorganisms and celiac microbe in sepsis mice by metagenomic technology.The results showed that the diversity of Enteric microorganisms decreased significantly after CLP.Futhermore,the abundance of Bacillus cereus、Streptococcus alactolyticus、Klebsiella pneumoniae and Morganella morganii is increased in sepsis.DcR3 treatment significantly restored enteric species diversity of sepsis mice,and significantly reduced the abundance of pathogenic bacteria.The expression of IL-1β、IL-6、TNF-ɑ、activated NK cells、Acetic acid and Propionic acid were positively correlated with the relative abundance of Morganella and Klebsiella,and negatively correlated with the abundance of uncultured＿bacterium＿g＿Lachnospiraceae(P < 0.001).The detection results of short-chain fatty acids(SCFAs)content in enteric microorganisms showed that the contents of Acetic acid、Propionic acid and Butyric in sepsis mice were significantly increased.However,the content of Butyric acid in sepsis is decreased significantly.After DcR3 treatment,the contents of Acetic acid and Propionic acid in enteric microorganisms significantly reduced,and Butyric acid content increased.Metagenomic analysis showed that Bacillus cereus 、 Streptococcus alactolyticus and Escherichia coli increased in sepsis mice.After DcR3 treatment,the abundance of Bacillus cereus decreased(P < 0.01).Bacillus、Morganella and Klebsiella was positively correlated with IL-1β 、 IL-6、 TNF-ɑ and activated NK cells in celiac microbe,negative correlated with correlated with Parabacteroides.In conclusion,intraperitoneal injection of DcR3 can significantly improve the survival rate of sepsis mice by inhibiting the inflammatory response、down-regulating the expression of pro-inflammatory factors、inhibiting the expression level of inflammatory cells 、reducing the expression of TNFSF10 protein、reducing the abundance of harmful microorganisms and increasing the abundance of beneficial bacteria.The results of this study are expected to provide an important theoretical and experimental basis for the application of DcR3 in the clinical treatment of sepsis.