Schistosome Eggs Impair Protective Th1/Th17 Immune Responses Against Salmonella Infection

Countries with a high incidence of helminth infections are characterized by high morbidity and mortality to infections with intracellular pathogens such as Salmonella. Some patients with Salmonella-Schistosoma co-infections develop a so-called “chronic septicemic salmonellosis,” with prolonged fever and enlargement of the liver and spleen. These effects are most likely due to the overall immunoregulatory activities of schistosomes such as induction of Tregs, Bregs, alternatively activated macrophages, and degradation of antibodies. However, detailed underlying mechanisms are not very well investigated. Here, we show that intraperitoneal application of live Schistosoma mansoni eggs prior to infection with Salmonella Typhimurium in mice leads to an impairment of IFN-γ and IL-17 responses together with a higher bacterial load compared to Salmonella infection alone. S. mansoni eggs were found in granulomas in the visceral peritoneum attached to the colon. Immunohistological staining revealed IPSE/alpha-1, a glycoprotein secreted from live schistosome eggs, and recruited basophils around the eggs. Noteworthy, IPSE/alpha-1 is known to trigger IL-4 and IL-13 release from basophils which in turn is known to suppress Th1/Th17 responses. Therefore, our data support a mechanism of how schistosomes impair a protective immune response against Salmonella infection and increase our understanding of helminth-bacterial co-infections

Black garlic exhibited hepatoprotective effect against monosodium glutamate-induced hepatotoxicity in animal model

Monosodium glutamate (MSG) is commonly used as a flavor-enhancing agent in foods, and studies have demonstrated its toxic effects in animal models. Black garlic is known for its antioxidant and anti-inflammatory properties; however, there is a lack of studies on the potential hepatoprotective effect of black garlic ethanol extract (BGE) against MSG-induced hepatotoxicity in rats. The aim of this study was to investigate the hepatoprotective effects of ethanol extract of black garlic against MSG-induced liver damage in animal model. Twenty-five male Wistar rats were randomly assigned to five groups (n=5): negative control, MSG only, and MSG with three different doses of BGE. The MSG only and MSG with BGE groups were orally administered with 8 mg/kg MSG daily. After MSG treatment, the MSG with BGE groups received BGE orally at daily doses of 200, 400, or 600 mg/kg body weight for 16 consecutive days. Subsequently, the levels of serum liver enzymes aspartate aminotransferase (AST), alanine aminotransferase (ALT), interferon-gamma (IFN-γ), and cyclooxygenase-2 (COX-2) were measured. Our data indicated that the group treated with 200 mg/kg BGE had significant lower levels of AST and ALT significantly compared to the MSG-only group. The MSG-treated group had higher levels of the inflammatory markers COX-2 and IFN-γ, which were lowered by administration of 200 mg/kg BGE. In contrast, higher doses of BGE led to greater levels of COX-2 and IFN-γ compared to those in the MSG-only group. This study suggested that BGE might have hepatoprotective effects at low dose, potentially mitigating MSG-induced liver damage. However, the higher dose of black garlic extract did not alleviate inflammation, as shown by the higher levels of COX-2 and IFN-γ

PAIN REDUCTION AFTER WET AND DRY CUPPING THERAPIES: ROLES OF α2β1 INTEGRIN AND μ-OPIOID RECEPTOR IN ANIMAL MODELS

Abstract Background: Wet cupping therapy (WCT) and dry cupping therapy (DCT) are complementary therapies in pain management. Stretching keratinocyte by cupping seems to stimulate the expression of α2β1-integrin. Its benefit in pain reduction could be mediated by the expression of μ-opioid receptor (MOR). Objective: This study aims to determine pain thresholds, α2β1-integrin and MOR after WCT and DCT. Method: Thirty-two adult male Wistar rats were divided into negative control, positive control, DCT, and WCT groups. In DCT group and WCT group, cupping was conducted at left and right paralumbar region 48h after CFA injection. Hot plate test was conducted 24h after therapy to assess pain threshold and immunohistochemistry were conducted from the skin for α2β1-integrin and the spinal cord for MOR. Result: WCG showed the highest expression of α2β1-integrin in the keratinocytes and MOR in the lamina II spinal cord than the other groups. On immunohistochemical examination, the expression of α2β1 integrin showed positive cells (keratinocytes) with brown stain on dry cupping and wet cupping. Significant differences were shown between negative control group showed significant differences with positive control group, dry cupping therapy group, and wet cupping therapy group. Conclusion: WCT is more effective than DCT in reducing pain. The benefit of both therapies in reducing pain in rats could be mediated by the expression of α2β1-integrin and MOR

Multigenerational Influences of the Fut2 Gene on the Dynamics of the Gut Microbiota in Mice

The FUT2 gene encodes an α-1,2-fucosyltransferase responsible for the expression of ABO histo-blood-group antigens on mucosal surfaces and bodily secretions. Individuals who carry at least one functional allele are known as "secretors," whereas those homozygous for loss-of-function mutations are known as "non-secretors." Non-secretor individuals are more susceptible to chronic inflammatory disorders such as Crohn's Disease, which may be mediated by alterations in the microbiota. Here, we investigated the dynamics of microbial community assembly with respect to genotype using a Fut2-deficient mouse model, taking the genotype of the maternal lineage over two generations into account. We found strong differences in community assembly of microbial communities over time, depending on the Fut2 genotype of the host and that of their progenitors. By applying network analyses, we further identified patterns of specialization and stabilization over time, which are influenced by the host and parental genotype during the process of community development. We also show genotype- and breeding-dependent patterns of community susceptibility to disturbance in a novel in silico approach integrating ecological- and network analysis. Our results indicate that it may be important to investigate the influence of Fut2 genotype in a familial context in order to fully understand its role in the etiology of chronic inflammatory disorders. © 2017 Rausch, Künzel, Suwandi, Grassl, Rosenstiel and Baines

Image_1_p38MAPK/MK2 signaling stimulates host cells autophagy pathways to restrict Salmonella infection.tif

Autophagy plays an important role in recognizing and protecting cells from invading intracellular pathogens such as Salmonella. In this work, we investigated the role of p38MAPK/MK2 in modulating the host cell susceptibility to Salmonella infection. Inhibition of p38MAPK or MK2 led to a significant increase of bacterial counts in Salmonella infected mouse embryonic fibroblasts (MEFs), as well as in MK2-deficient (Mk2-/-) cells. Furthermore, western blot analysis showed that Mk2-/- cells have lower level of LC3 lipidation, which is the indicator of general autophagy compared to Mk2-rescued cells. In Mk2-/- cells, we also observed lower activated TANK-binding kinase-1 phosphorylation on Ser172 and p62/SQTM1-Ser403 phosphorylation, which are important to promote the translocation of p62 to ubiquitinated microbes and required for efficient autophagy of bacteria. Furthermore, immunofluorescence analysis revealed reduced colocalization of Salmonella with LC3 and p62 in MEFs. Inhibition of autophagy with bafilomycin A1 showed increased bacterial counts in treated cells compared to control cell. Overall, these results indicate that p38MAPK/MK2-mediated protein phosphorylation modulates the host cell susceptibility to Salmonella infection by affecting the autophagy pathways.</p

Image_3_p38MAPK/MK2 signaling stimulates host cells autophagy pathways to restrict Salmonella infection.tif

Autophagy plays an important role in recognizing and protecting cells from invading intracellular pathogens such as Salmonella. In this work, we investigated the role of p38MAPK/MK2 in modulating the host cell susceptibility to Salmonella infection. Inhibition of p38MAPK or MK2 led to a significant increase of bacterial counts in Salmonella infected mouse embryonic fibroblasts (MEFs), as well as in MK2-deficient (Mk2-/-) cells. Furthermore, western blot analysis showed that Mk2-/- cells have lower level of LC3 lipidation, which is the indicator of general autophagy compared to Mk2-rescued cells. In Mk2-/- cells, we also observed lower activated TANK-binding kinase-1 phosphorylation on Ser172 and p62/SQTM1-Ser403 phosphorylation, which are important to promote the translocation of p62 to ubiquitinated microbes and required for efficient autophagy of bacteria. Furthermore, immunofluorescence analysis revealed reduced colocalization of Salmonella with LC3 and p62 in MEFs. Inhibition of autophagy with bafilomycin A1 showed increased bacterial counts in treated cells compared to control cell. Overall, these results indicate that p38MAPK/MK2-mediated protein phosphorylation modulates the host cell susceptibility to Salmonella infection by affecting the autophagy pathways.</p

cGAS-STING-TBK1-IRF3/7 induced interferon-β contributes to the clearing of non tuberculous mycobacterial infection in mice.

Type I interferons (IFN-I), such as IFN-α and IFN-β are important messengers in the host response against bacterial infections. Knowledge about the role of IFN-I in infections by nontuberculous mycobacteria (NTM) is limited. Here we show that macrophages infected with pathogens of the Mycobacterium avium complex produced significantly lower amounts of IFN-β than macrophages infected with the opportunistic pathogen M. smegmatis. To dissect the molecular mechanisms of this phenomenon, we focused on the obligate pathogen Mycobacterium avium ssp paratuberculosis (MAP) and the opportunistic M. smegmatis. Viability of both bacteria was required for induction of IFN-β in macrophages. Both bacteria induced IFN-β via the cGAS-STING-TBK1-IRF3/7-pathway of IFN-β activation. Stronger phosphorylation of TBK1 and higher amounts of extracellular bacterial DNA in the macrophage cytosol were found in M. smegmatis infected macrophages than in MAP infected macrophages. After intraperitoneal infection of mice, a strong Ifnb induction by M. smegmatis correlated with clearance of the bacteria. In contrast, MAP only induced weak Ifnb expression which correlated with bacterial persistence and increased number of granulomas in the liver. In mice lacking the type I interferon receptor we observed improved survival of M. smegmatis while survival of MAP was similar to that in wildtype mice. On the other hand, treatment of MAP infected wildtype mice with the IFN-I inducer poly(I:C) or recombinant IFN-β impaired the survival of MAP. This indicates an essential role of IFN-I in clearing infections by MAP and M. smegmatis. The expression level of IFN-I is decisive for transient versus persistent NTM infection

Systemic and Mucosal Immune Reactivity upon Mycobacterium avium ssp. paratuberculosis Infection in Mice.

Mycobacterium avium ssp. paratuberculosis (MAP) is the cause of Johne's disease, an inflammatory bowel disorder of ruminants. Due to the similar pathology, MAP was also suggested to cause Crohn's disease (CD). Despite of intensive research, this question is still not settled, possibly due to the lack of versatile mouse models. The aim of this study was to identify basic immunologic mechanisms in response to MAP infection. Immune compromised C57BL/6 Rag2-/- mice were infected with MAP intraperitoneally. Such chronically infected mice were then reconstituted with CD4+ and CD8+ T cells 28 days after infection. A systemic inflammatory response, detected as enlargement of the spleen and granuloma formation in the liver, was observed in mice infected and reconstituted with CD4+ T cells. Whereby inflammation in infected and CD4+CD45RBhi T cell reconstituted animals was always higher than in the other groups. Reconstitution of infected animals with CD8+ T cells did not result in any inflammatory signs. Interestingly, various markers of inflammation were strongly up-regulated in the colon of infected mice reconstituted with CD4+CD45RBlo/int T cells. We propose, the usual non-colitogenic CD4+CD45RBlo/int T cells were converted into inflammatory T cells by the interaction with MAP. However, the power of such cells might be not sufficient for a fully established inflammatory response in the colon. Nevertheless, our model system appears to mirror aspects of an inflammatory bowel disease (IBD) like CD and Johne's diseases. Thus, it will provide an experimental platform on which further knowledge on IBD and the involvement of MAP in the induction of CD could be acquired

Pathometagenomics reveals susceptibility to intestinal infection by Morganella to be mediated by the blood group-related B4galnt2 gene in wild mice

ABSTRACTInfectious disease is widely considered to be a major driver of evolution. A preponderance of signatures of balancing selection at blood group-related genes is thought to be driven by inherent trade-offs in susceptibility to disease. B4galnt2 is subject to long-term balancing selection in house mice, where two divergent allele classes direct alternative tissue-specific expression of a glycosyltransferase in the intestine versus blood vessels. The blood vessel allele class leads to prolonged bleeding times similar to von Willebrand disease in humans, yet has been maintained for millions of years. Based on in vivo functional studies in inbred lab strains, it is hypothesized that the cost of prolonged bleeding times may be offset by an evolutionary trade-off involving susceptibility to a yet unknown pathogen(s). To identify candidate pathogens for which resistance could be mediated by B4galnt2 genotype, we here employed a novel “pathometagenomic” approach in a wild mouse population, which combines bacterial 16S rRNA gene-based community profiling with histopathology of gut tissue. Through subsequent isolation, genome sequencing and controlled experiments in lab mice, we show that the presence of the blood vessel allele is associated with resistance to a newly identified subspecies of Morganella morganii, a clinically important opportunistic pathogen. Given the increasing importance of zoonotic events, the approach outlined here may find useful application in the detection of emerging diseases in wild animal populations