A novel compound targeting microglia activation

Abstract

Microglia, the resident macrophages of the brain, monitor the central nervous system (CNS) homeostasis. Alterations in homeostasis activate microglia in a variety of ways. Upon a patho-logical stimulus microglia change their appearance from a ramified into an amoeboid morphol-ogy, increase their phagocytic activity, and release pro-inflammatory cytokines and nitric oxide (NO). This response is involved in the pathological process in acute and chronical neuroinflam-matory disease such as stroke, multiple sclerosis, or schizophrenia. A dysregulated microglial activation can improve or exacerbate the outcome of those diseases. Therefore, it is of therapeu-tic interest to identify new compounds, which modulate the activation process of microglia spe-cifically. In this thesis, I identified a novel compound which inhibits specifically the induced NO release in microglia and macrophages independent of the applied pathological stimulus. We screened a library of 16544 compounds for their ability to interfere with lipopolysaccharide (LPS) induced NO release in the microglial cell line BV-2 and verified the results in the primary cultured neo-natal microglia. The compound showed a dose dependent reduction of NO after stimulation with LSP, interferon gamma (IFNγ), and polyinosinic:polycytidylic acid (polyIC) without being cyto-toxic. The LPS-induced release of proinflammatory cytokines TNFα, IL1β, and IL6 was not al-tered in the presents of the compound. I could show that the basal and LPS activated phagocy-tosis in microglia remained untouched, while it showed some influence on microglial migration. Furthermore, I could show that the compound does not interfere with the transcriptional regu-lation of the inducible nitric oxide synthase (iNOS), the enzyme responsible for the induced NO release in microglia and macrophages, and that it does decrease the NO release in already acti-vated microglia. The compound has a unique protein-like structure, composed of 4 proteinogenic and artificial amino acids. This chemical structure is new to the field of nitric oxide synthase (NOS). Its cell based IC50 value of 104 nM is below those of commonly used NOS inhibitors. In a prove of concept experiment, I could show that the application of this compound improves the outcome of stroke in mice.