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Lactobacillus rhamnosus LGG and LC705 effects on human primary macrophages

ABSTRACT: Analysis of human primary macrophages after live Lactobacillus rhamnosus GG (LGG) or LC705 stimulation for 6h and 24h. The results reveal novel mechanisms for probiotics-induced activation of the healthy human innate immune system. Macrophages are phagocytic cells of the innate immune system that perform sentinel functions to initiate appropriate responses to surrounding stimuli. Macrophages that reside at mucosa encounter ingested bacteria. Our understanding of the responses elicited by nonpathogenic bacteria in human innate immune system is limited. Lactobacillus are nonpathogenic bacteria commonly used in food and as supplements with health-promoting probiotic potential. In this study, we have utilized global gene expression profiling to compare the responses of human primary macrophages to two closely related, well-characterized L. rhamnosus strains LGG and LC705. Our results demonstrate that live LGG and LC705 induced quantitatively different gene expression in macrophages. A gene ontology analysis revealed functional similarities and differences in responses to LGG and LC705. Both LGG and LC705 induced IL-1b production in macrophages that required caspase-1 activity. LC705 but not LGG induced a strong type I IFN-dependent gene activation that correlated with the ability of LC705 to prevent influenza A virus replication and production of viral proteins in macrophages.

ORGANISM(S): Homo sapiens

PROVIDER: GSE20940 | GEO | 2013/03/18

SECONDARY ACCESSION(S): PRJNA124419

REPOSITORIES: GEO

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Project description:Analysis of human primary macrophages after live Lactobacillus rhamnosus GG (LGG) or LC705 stimulation for 6h and 24h. The results reveal novel mechanisms for probiotics-induced activation of the healthy human innate immune system. Macrophages are phagocytic cells of the innate immune system that perform sentinel functions to initiate appropriate responses to surrounding stimuli. Macrophages that reside at mucosa encounter ingested bacteria. Our understanding of the responses elicited by nonpathogenic bacteria in human innate immune system is limited. Lactobacillus are nonpathogenic bacteria commonly used in food and as supplements with health-promoting probiotic potential. In this study, we have utilized global gene expression profiling to compare the responses of human primary macrophages to two closely related, well-characterized L. rhamnosus strains LGG and LC705. Our results demonstrate that live LGG and LC705 induced quantitatively different gene expression in macrophages. A gene ontology analysis revealed functional similarities and differences in responses to LGG and LC705. Both LGG and LC705 induced IL-1b production in macrophages that required caspase-1 activity. LC705 but not LGG induced a strong type I IFN-dependent gene activation that correlated with the ability of LC705 to prevent influenza A virus replication and production of viral proteins in macrophages. Differentiated 7d human primary macrophages from 18 healthy individuals were stimulated with either live L. rhamnosus GG (LGG) or LC705 for 6 h and 24 h in RPMI medium containing penicillin and streptomycin. As a control, macrophages were stimulated with the medium only. The experiment was performed three times, each time with cells from six different individuals (samples 1-3). For each experiment, macrophages were stimulated separately as described, and macrophages from different donors were pooled after each stimulation experiment. Extracted RNA was hybridized to AffymetrixM-BM-. U133-plus2.0 GeneChipM-BM-. arrays.

Project description:Background: Sepsis-induced organ failure and high mortality are largely ascribed to the failure of bacterial clearance from the infected tissues. Recently, probiotic bacteria-released extracellular vesicles (BEVs) have been implicated as critical mediators of intercellular communication which are widely involved in the regulation inflammatory response. However, their functional role in macrophage phagocytosis during sepsis has never been explored. Methods: BEVs were collected from three different strains of probiotics including Lactobacillus plantarum WCFS1 (LP WCFS1), Lactobacillus rhamnosus Gorbach-Goldin (LGG), and Escherichia coli Nissle 1917 (EcN), or from LGG cultured under three pH conditions (pH5-acid, pH6.5-standard, pH8-akaline) through differential centrifugation, filtering, and ultracentrifugation of their culture supernatants. In vitro phagocytosis was measured in Raw264.7 and bone marrow-derived macrophages using pHrodo red E. coli BioParticles. The in vivo therapeutic effects of BEVs were tested using a feces-intraperitoneal-injection (FIP) model of polymicrobial sepsis. Results: LGG-derived EVs (BEVLGG) were the best among these three probiotics BEVs in stimulating macrophages to take up bacteria. Furthermore, BEVLGG collected from pH8 culture condition (BEVpH8) exhibited the strongest capacity of phagocytosis, compared with BEVpH5 and BEVpH6.5. Treatment of septic mice with BEVpH8 significantly prolonged animal survival, increased bacterial clearance from the blood, peritoneal lavage, and multiple organs, decreased serum levels of pro-inflammatory cytokines, alanine aminotransferase (ALT, marker of liver injury) and creatinine (Cr, marker of kidney injury), in comparison with control-treated septic mice. Mechanistically, RNA-seq and bioinformatic analysis identified that the FPR1/2 signaling was remarkably activated, along with its downstream pathways (PI3K-Akt-MARCO and NADPH-ROS) in BEVpH8-treated macrophages, compared with control cells. Accordingly, pre-addition of Boc2, a specific antagonist of FPR1/FPR2, to macrophages significantly attenuated BEVpH8-mediated phagocytosis, compared to controls. Conclusions: This study demonstrates that LGG-derived BEVs may have therapeutic effects against sepsis-induced organ injury and mortality through enhancing FPR1/2-mediated macrophage phagocytosis.

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Lactobacillus rhamnosus LGG and LC705 effects on human primary macrophages

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