Escherichia coli Braun Lipoprotein (BLP) exhibits endotoxemia – like pathology in Swiss albino mice

Abstract

The endotoxin lipopolysaccharide (LPS) promotes sepsis, but bacterial peptides also promote inflammation leading to sepsis. We found, intraperitoneal administration of live or heat inactivated E. coli JE5505 lacking the abundant outer membrane protein, Braun lipoprotein (BLP), was less toxic than E. coli DH5α possessing BLP in Swiss albino mice. Injection of BLP free of LPS purified from E. coli DH5α induced massive infiltration of leukocytes in lungs and liver. BLP activated human polymorphonuclear cells (PMNs) ex vivo to adhere to denatured collagen in serum and polymyxin B independent fashion, a property distinct from LPS. Both LPS and BLP stimulated the synthesis of platelet activating factor (PAF), a potent lipid mediator, in human PMNs. In mouse macrophage cell line, RAW264.7, while both BLP and LPS similarly upregulated TNF-α and IL-1β mRNA; BLP was more potent in inducing cyclooxygenase-2 (COX-2) mRNA and protein expression. Peritoneal macrophages from TLR2−/− mice significantly reduced the production of TNF-α in response to BLP in contrast to macrophages from wild type mice. We conclude, BLP acting through TLR2, is a potent inducer of inflammation with a response profile both common and distinct from LPS. Hence, BLP mediated pathway may also be considered as an effective target against sepsis.

Differential glycosylation of alpha-1-acid glycoprotein (AGP-1) contributes to its functional diversity

“…We previously observed that serum AGP-1 preferentially inhibits the TLR-4 agonist bacterial lipopolysaccharide (LPS), but not TLR-2 agonist Braun Lipoprotein (BLP) mediated inflammatory responses both in vivo and in vitro (24,25). We now show that the hepatocytederived serum AGP-1 (sAGP-1) is proinflammatory, while the neutrophil derived glycoforms (nAGP-1) primarily display antiinflammatory activity.…”

“…The labelled neutrophils (1x10 6 cells/well) were incubated with sAGP-1 or nAGP-1 (25, 50 and 100 µg/ml) in triplicate wells in twelve well cell culture plates (Nest Biotechnology Co. Ltd., China) pre-coated with 0.2% gelatin. Unbound neutrophils were removed by washing twice with HBSS/A and the adherent neutrophils were visualized and photographed at a magnification of 10x by fluorescent microscopy (Motic BA410 fluorescence microscope, Hong Kong; with Nikon DS-Qi2 camera, Japan) (25). The number of cells adhered in each well was determined by counting the cells adhered in 10 randomly chosen fields using ImageJ software and then calculating the average number of cells adhered per field.…”

“…Neutrophil migration was assessed using Transwell plates (Costar, Croning Inc., USA) with 5µm pore size inserts. 1x10 6 (200µl) freshly isolated neutrophils in M-199 medium were incubated in the upper chamber of the transwell and the chemoattractant, IL-8 (10 ng/ml) and/ or AGP-1 (serum and neutrophil glycoforms) (25,50 and 100 µg/ml) were introduced in the lower chamber. The number of neutrophils migrating to the lower chamber after incubation for 1 hour at 37° C with 5% CO 2 were counted with a hemocytometer and expressed as % neutrophil migrated using IL-8 as positive control (100% migration).…”

Class A CpG Oligonucleotide Priming Rescues Mice from Septic Shock via Activation of Platelet-Activating Factor Acetylhydrolase

“…Various inflammation-associated factors, including LPS, TNF-α, IL-1, nitric monoxide, PAF, Braun’s lipoprotein (BLP), and high-mobility group box 1 protein, promote inflammation leading to sepsis ( 39 , 40 ). It has been reported that anti-TNF antibody protects mice from LPS-induced lethal shock in a dose-dependent manner ( 41 ).…”

Modulation of inflammatory platelet-activating factor (PAF) receptor by the acyl analogue of PAF

“…Total lipids were extracted by using the Bligh and Dyer method (38), dried under nitrogen, and then reconstituted in methanol. A methanolic aliquot was used to quantify acyl-PAF, alkyl-PAF (C16: predominantly hexadecyl at the sn-1 position of GPC), lyso-PC, and lyso-PAF as described previously (16,43,44). Products or substrates present at the end of the third minute during platelet aggregation were measured relative to C16:0-d 4 -PAF (Cayman Chemical, Ann Arbor, MI) and are expressed as the mean number of nanomoles per assay from two independent experiments.…”