Journal Article

Manifestation of lipopolysaccharide-induced tolerance in neuro-glial primary cultures of the rat afferent somatosensory system

Nürnberger, Franz, Leisengang, Stephan, Murgott, Jolanta, Roth, Joachim, Rummel, Christoph, Ott, Daniela, Gerstberger, Rüdiger

2021

Overview

Objective Bacterial lipopolysaccharide (LPS) may contribute to the manifestation of inflammatory pain within structures of the afferent somatosensory system. LPS can induce a state of refractoriness to its own effects termed LPS tolerance. We employed primary neuro-glial cultures from rat dorsal root ganglia (DRG) and the superficial dorsal horn (SDH) of the spinal cord, mainly including the substantia gelatinosa to establish and characterize a model of LPS tolerance within these structures. Methods Tolerance was induced by pre-treatment of both cultures with 1 µg/ml LPS for 18 h, followed by a short-term stimulation with a higher LPS dose (10 µg/ml for 2 h). Cultures treated with solvent were used as controls. Cells from DRG or SDH were investigated by means of RT-PCR (expression of inflammatory genes) and immunocytochemistry (translocation of inflammatory transcription factors into nuclei of cells from both cultures). Supernatants from both cultures were assayed for tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6) by highly sensitive bioassays. Results At the mRNA-level, pre-treatment with 1 µg/ml LPS caused reduced expression of TNF-α and enhanced IL-10/TNF-α expression ratios in both cultures upon subsequent stimulation with 10 µg/ml LPS, i.e. LPS tolerance. SDH cultures further showed reduced release of TNF-α into the supernatants and attenuated TNF-α immunoreactivity in microglial cells. In the state of LPS tolerance macrophages from DRG and microglial cells from SDH showed reduced LPS-induced nuclear translocation of the inflammatory transcription factors NFκB and NF-IL6. Nuclear immunoreactivity of the IL-6-activated transcription factor STAT3 was further reduced in neurons from DRG and astrocytes from SDH in LPS tolerant cultures. Conclusion A state of LPS tolerance can be induced in primary cultures from the afferent somatosensory system, which is characterized by a down-regulation of pro-inflammatory mediators. Thus, this model can be applied to study the effects of LPS tolerance at the cellular level, for example possible modifications of neuronal reactivity patterns upon inflammatory stimulation.

• Publisher: Springer International Publishing,Springer Nature B.V
• Subject: Allergology / Animals / Astrocytes / Bioassays / Biomedical and Life Sciences / Biomedicine / CCAAT-Enhancer-Binding Protein-beta - metabolism / CCAAT/enhancer-binding protein / Cells, Cultured / Cytokines - genetics / Cytokines - metabolism / Dermatology / Dorsal horn / Dorsal root ganglia / Ganglia / Ganglia, Spinal - cytology / Gene expression / Immunocytochemistry / Immunology / Immunoreactivity / Inflammation / Interleukin 10 / Interleukin 6 / Lipopolysaccharides / Lipopolysaccharides - pharmacology / Macrophages / Microglial cells / Neuroglia - drug effects / Neuroglia - metabolism / Neurology / Neuronal-glial interactions / NF-kappa B - metabolism / NF-κB protein / Nuclear transport / Original Research Paper / Pain / Pharmacology/Toxicology / Polymerase chain reaction / Pretreatment / Rats / Rats, Wistar / Rheumatology / Sensory neurons / Somatosensory system / Spinal cord / Spinal Cord Dorsal Horn - cytology / Stat3 protein / STAT3 Transcription Factor - metabolism / Stimulation / Substantia gelatinosa / Thermal resistance / Transcription factors / Translocation / Tumor necrosis factor-TNF / Tumor necrosis factor-α