title: NFκB-dependent increase of kynurenine pathway activity in human placenta: inhibition by sulfasalazine.
abstract: Catabolism of tryptophan via the kynurenine pathway is up-regulated in the human placenta by infection, resulting in the release of pro-inflammatory and neuroactive metabolites into the fetal circulation. In this study we determined if activation of NFκB is involved in the inflammation-induced increase of kynurenine pathway activity in the human placenta. Placentae obtained after elective caesarian section at 37-40 weeks gestation (n=8), and explants (35-40 mg) prepared from terminal villi were incubated under standard conditions in the presence of 10 μg/ml LPS for 24 or 48 h; duplicates of each explant were incubated either with or without 5mM sulfasalazine added to the medium. Expression of mRNAs for key kynurenine-forming enzymes, indoleamine 2,3-dioxygrenase (IDO) and tryptophan 2,3-doalxygenase (TDO) and the inflammatory cytokines TNFα and IL6 was studied by RT-PCR. Kynurenine output by explants was measured in samples in the incubation medium by absorbance at 363nm after separation from other metabolites using an HPLC technique. Expression of IDO, TDO, TNFα and IL6 mRNAs was increased with LPS treatment, a response mitigated by the presence of sulfasalazine (P<0.01, P<0.01, P=0.03 &P=0.04). Kynurenine output into the culture medium increased with LPS treatment but this was also prevented by sulfasalazine at 24h (mean ± SEM; 412.1 ± 40 vs. 147.7 ± 48.9 nM/mg, P=0.01) and 48 h (636 ± 39.1 vs. 135.5 ± 29.8 nM/mg, P=0.001, respectively). Sulfasalazine inhibited the LPS induction of both the kynurenine pathway and pro-inflammatory cytokines in the placenta, implicating NFκB in the LPS effect. Direct measurement of NFκB activity showed that sulfasalazine decreased NFκB activation under both control and LPS-treated conditions. These observations show that kynurenine pathway activity in the human placenta is increased by a NFκB dependent pathway, and suggests a new therapeutic strategy for the management of pregnancies with in utero infection.
authors.affiliation: Department of Obstetrics and Gynaecology, Centre for Women's Health Research, Monash Institute of Medical Research, Monash University, Clayton 3168, Australia.
authors.forename: P
authors.initials: P
authors.lastname: Dharane Neé Ligam
authors.forename: U
authors.initials: U
authors.lastname: Manuelpillai
authors.forename: E
authors.initials: E
authors.lastname: Wallace
authors.forename: D W
authors.initials: DW
authors.lastname: Walker
mesh_terms: D000894:Anti-Inflammatory Agents, Non-Steroidal / Q000494:pharmacology*; D002467:Cell Nucleus / Q000187:drug effects / Q000378:metabolism; D016207:Cytokines / Q000235:genetics / Q000378:metabolism*; D005260:Female; D018507:Gene Expression Regulation, Developmental / Q000187:drug effects; D006801:Humans; D050503:Indoleamine-Pyrrole 2,3,-Dioxygenase / Q000235:genetics / Q000378:metabolism; D007375:Interleukin-1 / Q000235:genetics / Q000378:metabolism; D015850:Interleukin-6 / Q000235:genetics / Q000378:metabolism; D007737:Kynurenine / Q000378:metabolism*; D008070:Lipopolysaccharides / Q000633:toxicity; D016328:NF-kappa B / Q000235:genetics / Q000502:physiology*; D009924:Organ Culture Techniques; D010920:Placenta / Q000187:drug effects* / Q000378:metabolism*; D011247:Pregnancy; D011251:Pregnancy Complications, Infectious / Q000188:drug therapy; D012333:RNA, Messenger / Q000378:metabolism; D012460:Sulfasalazine / Q000494:pharmacology*; D014364:Tryptophan / Q000378:metabolism; D014366:Tryptophan Oxygenase / Q000235:genetics / Q000378:metabolism; D014409:Tumor Necrosis Factor-alpha / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism
chemical_list: D000894:Anti-Inflammatory Agents, Non-Steroidal; D016207:Cytokines; C508600:IL6 protein, human; D050503:Indoleamine-Pyrrole 2,3,-Dioxygenase; D007375:Interleukin-1; D015850:Interleukin-6; D008070:Lipopolysaccharides; D016328:NF-kappa B; D012333:RNA, Messenger; D014409:Tumor Necrosis Factor-alpha; D007737:Kynurenine; D012460:Sulfasalazine; D014364:Tryptophan; D014366:Tryptophan Oxygenase