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Title:   Author  item
                                            KIM, SUNG PHIL - Ajou University Of Koreaitem
                                            PARK, SUN OK - Str Biotech Co Ltditem
                                            LEE, SANG JONG - Str Biotech Co Ltditem
                                            SAM, SEOK HYUN - Ajou University Of Koreaitem
                                            Friedman, Mendel
  Submitted to: Journal of Agricultural and Food Chemistry
                                    Publication Type: Peer Reviewed Journal
                                    Publication Acceptance Date: 2/24/2014
                                    Publication  Date: 3/5/2014
                                    Citation: Kim, S., Park, S., Lee, S., Sam, S., Friedman, M. 2014. A polysaccharide isolated from the liquid culture of Lentinus edodes (shiitake) mushroom mycelia containing black rice bran protects mice against Salmonellosis through up-regulation of the Th1 immune reaction. Journal of Agricultural and Food Chemistry. 62:2384-2391. DOI: 10.1021/jf405223q.Interpretive Summary: In a recent publication (J. Agric. Food Chem. 2013, Online Nov. 8, DOI: 10.10.21/jf40317k) we describe the isolation and characterization of a new bioprocessed polysaccharide (BPP) from the mushroom mycelia culture of Lentinus edodes (Shiitake) with added black rice bran that protected mice against Salmonella lipopolysaccharide induced endotoxemia.  The protective effect was associated with stimulation of the immune system and amelioration of endotoxemia-induced pathological effects in the liver, lung, and kidney tissues.  Because the cause of endotoxemia (sepsis, septic shock) is largely associated with infections by Gram-negative bacteria such as Salmonella, it was of interest to find out whether BPP would also neutralize adverse effects of sublethal and lethal doses of Salmonella Typhimurium bacteria in infected mice via stimulation of the immune system.  To demonstrate this possibility, we evaluated the potential of the food-derived biopolymer to protect mice against Salmonella-induced infections.  Administration of BPP by intraperitoneal or by oral routs protected the mice against both against Salmonella-induced liver necrosis and lethality.  The protective effect was accompanied by stimulation of innate immune cells, as demonstrated by a series of chemical, biochemical, and in vivo assays.  Because the previous and present studies show that BPP can protect against both endotoxemia and infection, it has the potential to serve as a multi-functional food.  In addition, because reported studies on exposure to natural antimicrobials might be only partly effective in inactivating foodborne pathogens in different food categories, we suggest that it would be worthwhile to determine whether adding BPP to food would protect animals and humans against residual levels of pathogens and toxins produced by the pathogens after consumption via stimulation of the immune system.  This is a largely unexplored area of research in food safety.
                                Technical Abstract: The present study investigated the antibacterial effect of a bioprocessed polysaccharide (BPP) isolated from Lentinus edodes liquid mycelial culture supplemented with black rice bran against murine salmonellosis.  BPP was not bactericidal in vitro, but did, however stimulate uptake of the bacteria into RAW 264.7 murine macrophage cells, as indicated by increased colony-forming unit (CFU) counts of the contents of the lysed macrophages incubated with Salmonella Typhimurium for 30 and 60 min.  Two hours post-infection, the bacterial counts drastically increased in the macrophages, but 4 and 8 h post-infection BPP extract-treated cells showed lower bacterial counts than the vehicle (saline phosphate pH 7.4 buffer, PBS)-treated control.  BPP elicited altered morphology and markedly elevated inducible nitric oxide (NO) synthase (iNOS) mRNA and protein expression in the infected macrophage cells.  BPP also activated leukocytes in S. Typhimurium-infected mice, as determined by spleen lymphocyte proliferation and IFN-' levels in mice sera.  ELISA analysis on cytokine production by Th1 and Th2 immune cells from splenocytes of infected mice showed significant increases in the levels of the following Th1 cytokines: IL-1ß, IL-2, IL-6, and IL-12.  Histology assays of the livers of mice infected with a sublethal dose (1 × 104 CFU) of S. Typhimurium showed that BPP, administered daily through an intraperitoneal (ip) or oral route, protected against necrosis of the liver, a biomarker of in vivo salmonellosis.  The lifespan of mice similarly infected with a lethal dose of S. Typhimurium (1 × 105 CFU) was significantly extended by ip injection or oral administration of the BPP.  These results suggest that the activity of BPP against bacterial infection in mice occur mainly through the activation of macrophage-mediated immune response resulting from augmented Th1 immunity.  The significance of the results for microbial food safety and human health and further research needs are discussed.


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