In vivo Analysis of Neutrophil Infiltration during LPS-induced Peritonitis

Bio Protoc. 2016 Oct 5;6(19):e1945. doi: 10.21769/BioProtoc.1945.


Bacterial lipopolysaccharide (LPS) is present in the outer membrane of Gram-negative bacteria and functions as pathogen-associated molecular pattern (PAMP) (Whitfield and Trent, 2014). LPS therefore is a potent activator of inflammatory responses leading to cytokine release and neutrophils recruitment. The lipid A moiety of LPS activates the complex consisting of the LPS binding protein (LBP), CD14, MD-2 and Toll-like receptor 4 (TLR4) and the non-canonical inflammasome-linked caspases-4, 5 and 11, which in turn activate the canonical NLRP3 inflammasome (Shi et al., 2014; Hagar et al., 2013; Kayagaki et al., 2013; Hoshino et al., 1999; Poltorak, 1998; Nagai et al., 2002; Park et al., 2009; Ratsimandresy et al., 2013). In particular, the cytokine interleukin (IL)-1β produced in response to inflammasome activation has a crucial role in neutrophil recruitment through promoting neutrophil adhesion and migration (McDonald et al., 2010).This protocol allows studying of inflammatory response induced by LPS that affect neutrophil infiltration by tracking myeloperoxidase (MPO) activity in vivo (de Almeida et al., 2015).