Activation of murine microglial cells by muramyl dipeptide alone and in combination with Toll-like receptor agonists

Abstract

Meningitis, meningoencephalitis and sepsis are severe diseases causing many deaths all over the world every year. Escherichia coli (E. coli) is the most common Gram-negative bacterium causing neonatal meningitis and sepsis but also causes meningitis in old and immunocompromised people. Especially immunocompromised patients carry a high risk of developing infections e.g. in the central nervous system (CNS) caused by different pathogens, such as bacteria, viruses and fungi. One cause of this increased susceptibility to CNS infections might be a decreased local immune defense. Microglial cells, the resident immune cells of the brain, constitutively express Toll-like receptors (TLRs) and nucleotide-binding oligomerization domain (NOD)-like receptors, which are known to trigger innate immune responses against microbial infection upon pathogen recognition. This study aimed at three major questions: to detect whether stimulation of microglial cells with (i) the Nod2 ligand muramyl dipeptide (MDP) or (ii) the viral TLR3 agonist poly(I:C) affects phagocytosis and intracellular killing of E. coli K1, and (iii) to detect if there is synergism between MDP and the tested TLR agonists on the mentioned parameters. Therefore, primary cultures of murine microglia were stimulated with MDP or a TLR agonist alone as well as in combinations. Phagocytic activity was determined after 30 and 90 min of incubation with E. coli K1. To analyze the ability of microglia to kill ingested E. coli, bacteria were quantified at different time points after phagocytosis. In the current work, I was able to demonstrate for the first time that MDP and poly(I:C) alone increased phagocytosis and intracellular killing of E. coli K1 in murine microglia. In comparison, activation through the TLR system caused a stronger increase of phagocytosis than stimulation of the Nod2 system alone. Most notably, upon co-stimulation, the Nod2 and TLR systems can synergize to enhance both the phagocytic and bactericidal activities of microglial cells. In conclusion, microglial innate immune responses to invading E. coli K1 are enhanced by stimulation with the TLR3 agonist and Nod2 receptor alone as well as by a dual stimulation with poly(I:C) and MDP. These findings underline the cooperative action of innate immune receptor/signalling systems in fighting infectious threats. These results indicate that more studies should be investigated to find out whether a pre-stimulation with a Nod2 receptor agonist and a TLR agonist could improve CNS resistance to infections in immunocompromised patients and could therefore help the patients to develop mechanisms of resistance against lethal pathogens.