title: A Helminth Protease Inhibitor Modulates the Lipopolysaccharide-Induced Proinflammatory Phenotype of Microglia in vitro.
abstract: The aim of this study was to examine whether the natural protease inhibitor Av-cystatin (rAv17) of the parasitic nematode Acanthocheilonema viteae exerts anti-inflammatory effects in an in vitro model of lipopolysaccharide (LPS)-activated microglia.
  
  
  
  Primary microglia were harvested from the brains of 2-day-old Wistar rats and cultured with or without rAv17 (250 nM). After 6 and 24 h the release of nitric oxide (Griess reagent) and TNF-α (ELISA) was measured in the supernatant. Real-time PCR was performed after 2, 6 and 24 h of culture to measure the mRNA expression of IL-1β, IL-6, TNF-α, COX-2, iNOS and IL-10. To address the involved signaling pathways, nuclear NF-x0138;B translocation was visualized by immunocytochemistry. Morphological changes of microglia were analyzed by Coomassie blue staining. Differences between groups were calculated using one-way ANOVA with Bonferroni's post hoc test.
  
  
  
  Morphological analysis indicated that LPS-induced microglial transformation towards an amoeboid morphology is inhibited by rAv17. Av-cystatin caused a time-dependent downregulation of proinflammatory cytokines, iNOS and COX-2 mRNA expression, respectively. This was paralleled by an upregulated expression of IL-10 in resting as well as in LPS-stimulated microglia. Av-cystatin reduced the release of NO and TNF-α in the culture supernatant. Immunocytochemical staining demonstrated an attenuated translocation of NF-x0138;B by Av-cystatin in response to LPS. In addition, Western blot analysis revealed a rAv17-dependent reduction of the LPS-induced ERK1/2-pathway activation.
  
  
  
  The parasite-derived secretion product Av-cystatin inhibits proinflammatory mechanisms of LPS-induced microglia with IL-10, a potential key mediator.
authors.affiliation: Department of Trauma Surgery, University of Kiel, Kiel, Germany.
authors.forename: Peter
authors.initials: P
authors.lastname: Behrendt
authors.forename: Philipp
authors.initials: P
authors.lastname: Arnold
authors.forename: Max
authors.initials: M
authors.lastname: Brueck
authors.forename: Uta
authors.initials: U
authors.lastname: Rickert
authors.forename: Richard
authors.initials: R
authors.lastname: Lucius
authors.forename: Susanne
authors.initials: S
authors.lastname: Hartmann
authors.forename: Christian
authors.initials: C
authors.lastname: Klotz
authors.forename: Ralph
authors.initials: R
authors.lastname: Lucius
mesh_terms: D063486:Acanthocheilonema; D000818:Animals; D000831:Animals, Newborn; D002478:Cells, Cultured; D006376:Helminths; D018836:Inflammation Mediators / Q000037:antagonists & inhibitors / Q000378:metabolism*; D008070:Lipopolysaccharides / Q000633:toxicity*; D017628:Microglia / Q000187:drug effects / Q000378:metabolism*; D010641:Phenotype; D011480:Protease Inhibitors / Q000302:isolation & purification / Q000494:pharmacology*; D051381:Rats; D017208:Rats, Wistar
chemical_list: D018836:Inflammation Mediators; D008070:Lipopolysaccharides; D011480:Protease Inhibitors