RESEARCH ARTICLE A presumed antagonistic LPS identifies distinct functional organization of TLR4 in mouse microglia Christin D € oring 1 | Tommy Regen 1,2 | Ulla Gertig 1 | Denise van Rossum 1,3 | Anne Winkler 1 | Nasrin Saiepour 1 | Wolfgang Br € uck 1 | Uwe-Karsten Hanisch 1,4 | Hana Janova 1,5 1 Institute of Neuropathology, University Medical Center G€ ottingen, G€ ottingen 37075, Germany 2 Institute of Molecular Medicine, University of Mainz, Mainz 55131, Germany 3 Sartorius-Stedim Biotech GmbH, G€ ottingen 37079, Germany 4 Paul-Flechsig-Institute for Brain Research, University of Leipzig, Leipzig 04103, Germany 5 Clinical Neuroscience, Max-Planck-Institute of Experimental Medicine, G€ ottingen 37075, Germany Correspondence Hana Janova, Clinical Neuroscience, Max-Planck-Institute of Experimental Medicine, Hermann-Rein-Straße 3, 37075 G€ ottingen, Germany. E-mail: janova.hana@seznam.cz Abstract Microglia as principle innate immune cells of the central nervous system (CNS) are the first line of defense against invading pathogens. They are capable of sensing infections through diverse recep- tors, such as Toll-like receptor 4 (TLR4). This receptor is best known for its ability to recognize bacterial lipopolysaccharide (LPS), a causative agent of gram-negative sepsis and septic shock. A putative, naturally occurring antagonist of TLR4 derives from the photosynthetic bacterium Rhodo- bacter sphaeroides. However, the antagonistic potential of R. sphaeroides LPS (Rs-LPS) is no universal feature, since several studies suggested agonistic rather than antagonistic actions of this molecule depending on the investigated mammalian species. Here we show the agonistic versus antagonistic potential of Rs-LPS in primary mouse microglia. We demonstrate that Rs-LPS effi- ciently induces the release of cytokines and chemokines, which depends on TLR4, MyD88, and TRIF, but not CD14. Furthermore, Rs-LPS is able to regulate the phagocytic capacity of microglia as agonist, while it antagonizes Re-LPS-induced MHC I expression. Finally, to our knowledge, we are the first to provide in vivo evidence for an agonistic potential of Rs-LPS, as it efficiently triggers the recruitment of peripheral immune cells to the endotoxin-challenged CNS. Together, our results argue for a versatile and complex organization of the microglial TLR4 system, which specifically translates exogenous signals into cellular functions. Importantly, as demonstrated here for microglia, the antagonistic potential of Rs-LPS needs to be considered with caution, as reactions to Rs-LPS not only differ by cell type, but even by function within one cell type. KEYWORDS chemokines, cytokines, inflammation, monocytes, MyD88, neutrophils, Rs-LPS, TRIF 1 | INTRODUCTION Microglia are the innate immune cells of the central nervous system (CNS) parenchyma constituting a distinct population of long-lived tissue- resident macrophages (Bruttger et al., 2015; Goldmann et al., 2016). They constantly screen their environment (Hanisch & Kettenmann, 2007) and react towards the slightest disturbance in order to maintain CNS homeostasis and functionality (Nimmerjahn, Kirchhoff, & Helmchen, 2005). Upon activation, microglia can execute diverse functions, including cytokine and chemokine production (Regen et al., 2011), attraction of peripheral immune cells to the CNS, phagocytosis of infectious agents (Ribes et al., 2009) and endogenous material (Regen et al., 2011; van Rossum, Hilbert, Strassenburg, Hanisch, & Bruck, 2008) as well as MHC I surface expression as prerequisite for antigen presentation (Regen et al., 2011). Thereby, some of these functions are organized pan-populational, whereas others reside within subpopulations of microglia, indicating a certain degree of heterogeneity within the microglia population (Gertig & Hanisch 2014; Hanisch 2013; Scheffel et al., 2012). Microglial activation can be triggered through a broad range of cell surface and intracellular receptors, among them the family of pattern recognition receptors (PRR), which sense highly conserved structures Uwe-Karsten Hanisch and Hana Janova contributed equally to this work. Glia. 2017;1–10. wileyonlinelibrary.com/journal/glia V C 2017 Wiley Periodicals, Inc. | 1 Received: 8 November 2016 | Revised: 25 March 2017 | Accepted: 30 March 2017 DOI: 10.1002/glia.23151