B. pertussis 165 LPS

Lipopolysaccharides are anchored to the outer membrane of Gram-negative bacteria by a hydrophobic moiety known as lipid A, which potently activates the host innate immune response. Lipid A of Bordetella pertussis, the causative agent of whooping cough, displays unusual structural asymmetry with respect to the length of the acyl chains at the 3 and 3 positions, which are 3OH-C10 and 3OH-C14 chains, respectively. Both chains are attached by the acyltransferase LpxA, the first enzyme in the lipid A biosynthesis pathway, which, in B. pertussis, has limited chain length specificity. However, this only partially explains the strict asymmetry of lipid A. In attempts to modulate the endotoxicity of B. pertussis lipid A, here we expressed the gene encoding LpxA from Neisseria meningitidis, which specifically attaches 3OH-C12 chains, in B. pertussis. This expression was lethal, suggesting that one of the downstream enzymes in the lipid A biosynthesis pathway in B. pertussis cannot handle precursors with a 3OH-C12 chain. We considered that the UDP-diacylglucosamine pyrophosphohydrolase LpxH could be responsible for this defect as well as for the asymmetry of B. pertussis lipid A. Expression of meningococcal LpxH in B. pertussis indeed resulted in new symmetric lipid A species with 3OH-C10 or 3OH-C14 chains at both the 3 and 3 positions, as revealed by MS analysis. Furthermore, co-expression of meningococcal lpxH and lpxA resulted in viable cells that incorporated 3OH-C12 chains in B. pertussis lipid A. We conclude that the asymmetry of B. pertussis lipid A is determined by the acyl chain length specificity of LpxH.

Background: Oestrogen has anti-inflammatory property in obesity. However, the mechanism is still not defined. Objective: To investigate the effect of oestrogen on LPS- induced monocyte chemoattractant protein-1 (MCP-1) production in adipocytes. Methods: Lipopolysaccharides (LPS) was used to imitate inflammatory responses and monocyte chemotactic protein-1 (MCP-1) was selected as an inflammatory marker to observe. 17 beta-Estradiol (E-2), SB203580 (SB), pyrrolidine dithiocarbamate (PDTC), pertussis toxin (PTX), wortmannin (WM), p65 siRNA and p38 MAPK siRNA were pre-treated respectively or together in LPS- induced MCP-1. Then p38 MAPK and NF-kappa B cascade were silenced successively to observe the change of each other. Lastly, oestrogen receptor (ER) alpha agonist, ER beta agonist and ER antagonist were utilised. Results: LPS-induced MCP-1 largely impaired by pre-treatment with E-2, SB, PDTC or silencing NF-kappa B subunit. E-2 inhibited LPS- induced MCP-1 in a time- and dose-dependent manner, which was related to the suppression of p65 translocation tonucleus. Furthermore, LPS rapidly activated p38 MAPK, while E-2 markedly inhibited this activation. It markedly attenuated LPS- stimulated p65 translocation to nucleus and MCP-1 production by transfecting with p38 MAPK siRNA or using p38 MAPK inhibitor. The oestrogen's inhibitory effect was mimicked by the ER alpha agonist, but not by the ER beta agonist. The inhibition of E-2 on p38 MAPK phosphorylation was prevented by ER antagonist. Conclusions: E-2 inhibits LPS-stimulated MCP-1 in adipocytes. This effect is related to the inhibition of p38 MAPK/NF-kappa B cascade, and ER alpha appears to be the dominant ER subtype in these events. (C) 2016 Asia Oceania Association for the Study of Obesity. Published by Elsevier Ltd. All rights reserved.