Microglial TIR-domain-containing adapter-inducing interferon-β (TRIF) deficiency promotes retinal ganglion cell survival and axon regeneration via nuclear factor-κB.
ABSTRACT TIR-domain-containing adapter-inducing interferon-β (TRIF) is the sole downstream adaptor of Toll-like receptor (TLR)3, which is one of the major signaling pathways in immune cells leading to neuroinflammation in the central nervous system. Overexpression of TRIF may lead to activation of inflammatory responses, and contribute to pathophysiological progression in both acute and chronic neurodegenerative retinal diseases. In the present study, was aimed to elucidate the contributions of TRIF to optic nerve (ON) regeneration and retinal ganglion cell (RGC) survival following injury to the ON, a widely studied model of central nervous system injury and of degenerative diseases such as glaucoma.
We used retrograde labeling with a fluorochrome, hydroxystilbamidine (Fluorogold) to evaluate RGC survival, and immunostaining with growth-associated protein-43 to evaluate axon regeneration in an ON crush model. Changes in microglial cytokines following RGC injury was examined with ELISA and real-time PCR. In vivo studies were carried out in wild-type and trif-/- mice. A Transwell co-culture system and migration test were used to mimic the crosstalk between microglia and RGCs. TRIF-associated downstream adaptors were determined by western blotting.
Compared with wild-type (WT) mice, TRIF knockout (KO) mice displayed a robust ability to regenerate axons 3 or 7 days after nerve injury. In addition, RGC survival was considerably higher in trif-/- than in WT mice. ON lesion induced less microglial activation in trif-/- than in WT mice. and more WT microglia distorted and migrated toward the foramen opticum. In the transwell system, few trif-/- microglia migrated through the membrane when stimulated by the performed lesion on RGC axons in a transwell system. Inactivation of microglial cells in trif-/- mice was associated with reduced production of inflammatory cytokines, as detected with real-time RT-PCR and ELISA. Furthermore western blot analysis showed that activation of known downstream effectors of TRIF, including TBK1, IKKε and NF-κB, were significantly inhibited by TRIF deficiency.
Our results indicate that TRIF deficiency promotes ON axon regeneration by attenuating microglial activation and consequently reducing the release of harmful cytokines via NF-κB inactivation.
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ABSTRACT: Stimulation of mouse astrocytes and microglia in enriched culture by lipopolysaccharide (LPS) results in the production of a cytotoxic factor that is functionally similar to macrophage-derived tumor necrosis factor-alpha (TNF). This cytotoxic factor kills murine L929 cells, and is neutralized by a monoclonal anti-mouse TNF. The production of this factor is inhibited by pretreatment of cells by actinomycin D. Microglia are more sensitive to LPS and produce much higher activity of cytotoxicity in lower concentration of LPS than astrocytes. These results suggest that microglia are the major producers of TNF in the brain and may play a role in certain pathological conditions in the brain.Brain Research 08/1989; 491(2):394-7. · 2.73 Impact Factor
Article: Immunohistochemical localization of a macrophage-specific antigen in developing mouse retina: phagocytosis of dying neurons and differentiation of microglial cells to form a regular array in the plexiform layers.[show abstract] [hide abstract]
ABSTRACT: In the developing mouse retina degenerating neurons can be observed initially in the ganglion cell layer followed by a phase of cell death in the inner nuclear layer. Using an immunohistochemical method to localize the mouse macrophage specific antigen F4/80, we show that macrophages migrate from the vascular supply overlying the developing retina and phagocytose the degenerating neurons. The macrophages subsequently differentiate to become the microglia of the retina and form a regularly spaced distribution across the retina in the inner and outer plexiform layers. These experiments provide strong evidence for the mesodermal origin of central nervous system microglia.The Journal of Cell Biology 08/1983; 97(1):253-7. · 10.26 Impact Factor