The choroid plexus epithelium as a novel player in the stomach-brain axis during Helicobacter infection.

Several studies suggest a link between shifts in gut microbiota and neurological disorders. Recently, we reported a high prevalence of Helicobacter suis (H. suis) in patients with Parkinson's disease. Here, we evaluated the effect of gastric H. suis infection on the brain in mice. One month of infection with H. suis resulted in increased brain inflammation, reflected in activation of microglia and cognitive decline. Additionally, we detected choroid plexus inflammation and disruption of the epithelial blood-cerebrospinal fluid (CSF) barrier upon H. suis infection, while the endothelial blood-brain barrier (BBB) remained functional. These changes were accompanied by leakage of the gastrointestinal barrier and low-grade systemic inflammation, suggesting that H. suis-evoked gastrointestinal permeability and subsequent peripheral inflammation induces changes in brain homeostasis via changes in blood-CSF barrier integrity. In conclusion, this study shows for the first time that H. suis infection induces inflammation in the brain associated with cognitive decline and that the choroid plexus is a novel player in the stomach-brain axis.

Synthesis and Validation of a Hydroxypyrone-Based, Potent, and Specific Matrix Metalloproteinase-12 Inhibitor with Anti-Inflammatory Activity In Vitro and In Vivo.

A hydroxypyrone-based matrix metalloproteinase (MMP) inhibitor was synthesized and assayed for its inhibitory capacity towards a panel of ten different MMPs. The compound exhibited selective inhibition towards MMP-12. The effects of inhibition of MMP-12 on endotoxemia and inflammation-induced blood-cerebrospinal fluid barrier (BCSFB) disruption were assessed both in vitro and in vivo. Similar to MMP-12 deficient mice, inhibitor-treated mice displayed significantly lower lipopolysaccharide- (LPS-) induced lethality compared to vehicle treated controls. Following LPS injection Mmp-12 mRNA expression was massively upregulated in choroid plexus tissue and a concomitant increase in BCSFB permeability was observed, which was restricted in inhibitor-treated mice. Moreover, an LPS-induced decrease in tight junction permeability of primary choroid plexus epithelial cells was attenuated by inhibitor application in vitro. Taken together, this hydroxypyrone-based inhibitor is selective towards MMP-12 and displays anti-inflammatory activity in vitro and in vivo.

Neutralizing TNFα restores glucocorticoid sensitivity in a mouse model of neutrophilic airway inflammation.

Asthma is a heterogeneous disorder, evidenced by distinct types of inflammation resulting in different responsiveness to therapy with glucocorticoids (GCs). Tumor necrosis factor α (TNFα) is involved in asthma pathogenesis, but anti-TNFα therapies have not proven broadly effective. The effects of anti-TNFα treatment on steroid resistance have never been assessed. We investigated the role of TNFα blockade using etanercept in the responsiveness to GCs in two ovalbumin-based mouse models of airway hyperinflammation. The first model is GC sensitive and T helper type 2 (Th2)/eosinophil driven, whereas the second reflects GC-insensitive, Th1/neutrophil-predominant asthma subphenotypes. We found that TNFα blockade restores the therapeutic effects of GCs in the GC-insensitive model. An adoptive transfer indicated that the TNFα-induced GC insensitivity occurs in the non-myeloid compartment. Early during airway hyperinflammation, mice are GC insensitive specifically at the level of thymic stromal lymphopoietin (Tslp) transcriptional repression, and this insensitivity is reverted when TNFα is neutralized. Interestingly, TSLP knockout mice displayed increased inflammation in the GC-insensitive model, suggesting a limited therapeutic application of TSLP-neutralizing antibodies in subsets of patients suffering from Th2-mediated asthma. In conclusion, we demonstrate that TNFα reduces the responsiveness to GCs in a mouse model of neutrophilic airway inflammation. Thus antagonizing TNFα may offer a new strategy for therapeutic intervention in GC-resistant asthma.

TNFR1-induced lethal inflammation is mediated by goblet and Paneth cell dysfunction.

Tumor necrosis factor (TNF) is a powerful activator of the immune system and a well-validated target for treatment of autoimmune diseases. Injection of TNF induces systemic lethal inflammation characterized by hypothermia, induction of multiple cytokines, and extensive damage to multiple organs. Previously, we reported that TNF-induced lethal inflammation is strictly TNFR1(P55)-dependent. We also uncovered a crucial role for P55 expression levels in intestinal epithelial cells (IECs), in which P55+/+ expression is sufficient to sensitize to TNF lethality in an otherwise fully protected P55+/- background. Here, we investigated the molecular mechanism that drives TNF toxicity in IECs. Unexpectedly, we found that the degree of TNF-induced enterocyte damage and apoptosis in IECs is equally strong in TNF-sensitive P55+/+ mice and TNF-resistant P55+/- mice. Our results suggest that P55+/+-induced signaling causes goblet and Paneth cell dysfunction, leading to severe epithelial barrier dysfunction. As a result, intestinal permeability and systemic bacterial spread are induced, causing lethal systemic inflammation. In conclusion, we identified P55-induced goblet and Paneth cell dysfunction as a crucial mechanism for TNF-induced systemic and lethal inflammation.

Determining differentially expressed miRNAs and validating miRNA--target relationships using the SPRET/Ei mouse strain.

Micro RNAs (miRs) are involved in many biological processes. The challenge of identifying genes influenced by miRs is evidenced by the relatively few validated miR-target interactions. In this work, we used the Mus spretus SPRET/Ei strain as an in vivo system to identify new miR-target relations. Mus spretus diverged from Mus musculus over one million years ago, making it genetically and phenotypically divergent. SPRET/Ei mice are resistant to inflammation and several cancers, making them attractive for different research fields. Their phenotype is unique and is considerably different from that of almost all other laboratory mouse strains. We exploited the characteristics of SPRET/Ei mice as a tool to identify miR-target relationships. Hepatic genes and miRs differentially expressed between C57BL/6 and SPRET/Ei mice at basal levels were identified with an Affymetrix microarray and a multiplex qPCR, respectively. A total of 955 genes and 38 miRs were identified as differentially expressed. Increased miR expression might result in downregulation of its target mRNA and vice versa. Subsequently, we used our miR and mRNA data to identify possible in vivo miR-target interactions. Ingenuity pathway analysis (IPA) analysis revealed 380 possible miR-target interactions. Five miRs were selected for experimental validation by in vivo overexpression of the miRs. This resulted in the confirmation of six previously unknown miR-target interactions: miR-146a, Zdhhc2; miR-150, Elovl3, Kcnk5, and Nrd1d2; miR-155, Camta1; and miR-592, Steap2. In conclusion, we show that SPRET/Ei mice can be used as a platform for miR-target identification in vivo, and we used this platform to identify and experimentally confirm miR-target interactions.

Pro-inflammatory effects of matrix metalloproteinase 7 in acute inflammation.

Matrix metalloproteinase 7 (MMP7) is a member of the MMP family. In the small intestine, MMP7 is responsible for activating α-defensins, which are broad-spectrum anti-microbial peptides produced by the Paneth cells. We report that MMP7(-/-) mice are resistant to LPS-induced lethality and that this resistance is correlated with reduced levels of systemic cytokines. LPS induced the upregulation and activation of MMP7 in the small intestine, degranulation of the Paneth cells, and induction of intestinal permeability in MMP7(+/+) mice. In MMP7(-/-) mice, both LPS-induced intestinal permeability and consequent bacterial translocation to the mesenteric lymph nodes were reduced. Based on gene expression analysis and evaluation of intestinal damage, we attribute the protected state of MMP7(-/-) mice to reduced intestinal inflammation. Interestingly, we found that different α-defensins, namely Crp1 (DEFA1) and Crp4 (DEFA4), can stimulate IL-6 release in macrophages and ileum explants in a TLR4 independent way. We conclude that absence of MMP7 protects mice from LPS-induced intestinal permeability and lethality, and suggest that MMP7-activated α-defensins, in addition to their previously recognized bactericidal and anti-inflammatory roles, may exhibit pro-inflammatory activities in the intestine by activating macrophages and amplifying the local inflammatory response in the gut, leading to intestinal leakage and subsequent increase in systemic inflammation.