Study On The Activation Of Macrophage Induced By Lactobacillus Rhamnosus Against Salmonella Typhimurium

Macrophage(Mφ),as an important bridge between innate immunity and acquired immunity,could mediate innate and adaptive immunity to prevent and eliminate pathogen invasion.Mφcan also work as a shelter for certain intracellular pathogens(such as Salmonella)to escape the host’s immune defense.Lactic acid bacteria(LAB)are important representative of probiotics and played an important role in activating antigen-presenting cells such as Mφand dendritic cells(DC)owning its unique colonization position and probiotic function.Therefore,it is commonly used to prevent various intestinal diseases caused by intestinal pathogenic bacteria infection.Lactobacillus rhamnosus GG has a significant role in the prevention of inflammatory responses caused by Salmonella typhimurium(S.typhimurium).Therefore,S.Typhimurium was applied to establish enteritis model following by pre-administration with Lactobacillus rhamnosus GG(LGG)strain.The protective effect of LGG against S.Typhimurium was evaluated.(1)LGG can alleviate enteritis caused by S.Typhimurium infectionIn order to clarify the protective effect of LGG against S.Typhimurium infection,S.Typhimurium was used in this study to establish the enteritis model,and the model mice were pre-administrated with LGG to intervene in the enteritis model.The protection effects of LGG on intestinal enteritis were evaluated by measuring the growth rate and survival rate of mice,intestinal pathological changes,organ index,bacterial burden in tissue,and intestinal cytokine secretion.The results showed that LGG could increase the body weight and survival rate of mice infected with S.Typhimurium(P<0.001);intestinal pathological damage was significantly reduced,and pathological score was significantly decreased(P<0.05);The results showed that LGG could inhibit the invasion of S.Typhimurium to the spleen(P<0.001),liver(P<0.001)and mesenteric lymph nodes(MLN)(P<0.01);In addition,LGG could promote the secretion IL-10(P<0.05),inhibit the secretion of NO and IL-12(P<0.001),which demonstrated that LGG have the ability of alleviating intestinal inflammation.This result was also consistent with intestinal pathological change.Therefore,LGG had the protective effect in preventing S.Typhimurium infection and improving intestinal damage.(2)Study on LGG-induced macrophage activation and its mechanismIn order to further reveal the biological function of LGG-mediated Mφ,this experiment established C57B/L6 mouse bone marrow-derived macrophage(BMDMs)and intestinal inflammation in vitro.It analyzed the effects of LGG on the activation and phagocytosis of BMDMs,and explored the regulatory role of LGG on inflammatory signal pathways of NF-кB and iNOS.The expression levels of CD80~+and CD86~+of BMDMs activated by LGG were detected by flow cytometry.The expression of inducible nitric oxide synthase(iNOS)in BMDMs was analyzed by immunofluorescence.The phagocytic killing effect of BMDMs on S.typhimurium was detected by immunofluorescence.The secretion levels of cytokines NO,IL-10 and IL-12 were measured by ELISA.The expression levels of NF-κB and iNOS signaling molecules were detected by Real-time PCR and Western blot.The results showed that LGG could increase the expression of CD80~+(P<0.01)and CD86~+(P<0.001)molecules in BMDMs and promote the activation of Mφ.Immunofluorescence assay showed that LGG enhanced the phagocytosis of BMDMs(P<0.01)and the ability to kill S.typhimurium by increasing the expression level of iNOS protein in BMDMs.ELISA showed that LGG could inhibit the secretion of IL-12(P<0.001)and NO(P<0.001)and increase the secretion level of IL-10(P<0.001).Real-time PCR and Western blot showed that LGG could inhibit the activation and protein expression of NF-κB(P<0.05),iNOS(P<0.05).The above results indicate that LGG may alleviate intestinal inflammation and intestinal damage by inhibiting the expression of inflammatory signaling molecules.This study found that LGG enhances the ability of BMDMs to sterilize and phagocytose S.typhimurium by increasing the level of iNOS expression in BMDMs.It also inhibits the inflammatory signaling molecules of NF-кB and iNOS in BMDMs,alleviates S.typhimurium-induced enteritis and maintains the immune balance of animals.In conclusion,LGG can mediate macrophage activation to alleviate the intestinal inflammation against S.typhimurium.