LPS from bovine serum albumin drives TNF-α release during ex-vivo placenta perfusion experiments, contaminates the perfusion system but can be effectively removed by oxidative cleaning

Abstract Introduction The dual ex-vivo perfusion of human placental tissue is useful to study inflammatory pathways. We found significant TNF-α release in negative controls similar in concentration to lipopolysaccharide (LPS) stimulated placentas. The aim of the current study was to (i) identify sou...

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Bibliographic Details
Published in:	Placenta (Eastbourne) Vol. 35; no. 12; pp. 1095 - 1098
Main Authors:	Vasanthan, T, Rochow, N, Mian, F, Codini, T, DeFrance, B, Fusch, G, Samiee-Zafarghandy, S, Fusch, C
Format:	Journal Article
Language:	English
Published:	Kidlington Elsevier Ltd 01-12-2014 Elsevier
Subjects:	Acid–base depyrogenation Animals Biological and medical sciences Bovine serum albumin Cell Line Embryology: invertebrates and vertebrates. Teratology Female Fundamental and applied biological sciences. Psychology Internal Medicine Lipopolysaccharide Lipopolysaccharides - pharmacology Macrophages - cytology Macrophages - drug effects Macrophages - metabolism Mice Obstetrics and Gynecology Oxidative depyrogenation Placenta - cytology Placenta - drug effects Placenta - metabolism Placenta perfusion model Pregnancy Serum Albumin, Bovine - pharmacology Tumor Necrosis Factor-alpha - metabolism Lipopolysaccharide Bovine serum albumin Placenta perfusion model Oxidative depyrogenation Acid–base depyrogenation Bovine Fetal membrane Cytokine Ox Serum albumin Vertebrata Mammalia Placenta Ex vivo Perfusion Animal Artiodactyla Ungulata Tumor necrosis factor α Release
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Summary:	Abstract Introduction The dual ex-vivo perfusion of human placental tissue is useful to study inflammatory pathways. We found significant TNF-α release in negative controls similar in concentration to lipopolysaccharide (LPS) stimulated placentas. The aim of the current study was to (i) identify sources driving TNF-α release and (ii) develop an approach to control for it. Method (i) To determine sources leading to TNF-α release, solutions frequently circulated through the perfusion system and perfusion media with different bovine serum albumin (BSA) quality were exposed to mouse macrophage cell lines (RAW264.7) and subsequently measured for TNF-α expression. (ii) To assess memory effects and validate cleaning procedures, sham perfusion experiments were conducted either in the presence or absence of exogenous LPS, in new tubing that was contaminated, cleaned and analyzed for the effectiveness of LPS removal. Oxidative and acid–base cleaning were tested for their effectiveness to reduce LPS contamination. Results TNF-α release, observed in negative control experiments, was attributed to the use of LPS-contaminated BSA as well as inadequate cleaning of the perfusion system. Once introduced in the perfusion system, LPS accumulated and created a memory effect. Oxidative but not acid–base depyrogenation effectively reduced LPS levels to concentrations that were in accordance with FDA guidelines (<0.5 EU/mL) for medical equipment redeemed appropriate for re-use. Discussion LPS contamination of the placenta perfusion model could have confounding effects on experimental outcomes leading to misinterpretation of data. To circumvent LPS contamination LPS-free BSA and oxidative depyrogenation cleaning techniques should be implemented in future placental perfusion studies.
Bibliography:	ObjectType-Article-1 SourceType-Scholarly Journals-1 ObjectType-Feature-2 content type line 23
ISSN:	0143-4004 1532-3102
DOI:	10.1016/j.placenta.2014.10.005