Ciesielska, A. | Search bioRxiv

bioRxiv2024

Dynamic fibroblast-immune interactions shape wound healing after brain injury

Fibroblasts coordinate the response to tissue injury, directing organ regeneration versus scarring. In the central nervous system (CNS), fibroblasts are uncommon cells enriched at tissue borders, and their molecular, cellular, and functional interactions after brain injury are poorly understood. Here we define the fibroblast response to sterile brain damage across time and space. Early pro-fibrotic myofibroblasts infiltrated CNS lesions and were functionally and spatially organized by fibroblast TGF{beta} signaling, pro-fibrotic macrophages and microglia, and perilesional brain glia that activated TGF{beta} via integrin v{beta}8. Early myofibroblasts subsequently transitioned into a variety of late states, including meningeal and lymphocyte-interactive fibroblasts that persisted long term. Interruption of this dynamic fibroblast-macrophage-glial coordination impaired brain wound healing and the resolution of neuroinflammation, disrupted generation of late de novo CNS lymphocyte niches, and increased mortality in a stroke model. This work highlights an unexpected role of fibroblasts as coordinate regulators of CNS healing and neuroinflammation after brain injury.

immunology↗

bioRxiv2023

Flotillins affect LPS-induced TLR4 signaling by modulating the trafficking and abundance of CD14

Lipopolysaccharide induces a strong pro-inflammatory reaction of macrophages upon activation of Toll-like 4 receptor (TLR4) with the assistance of CD14 protein. Considering a key role of plasma membrane rafts in CD14 and TLR4 activity and the significant impact exerted on that activity by endocytosis and intracellular trafficking of the both LPS acceptors, it seemed likely that the pro-inflammatory reaction could be modulated by flotillins. Flotillin-1 and -2 are scaffolding proteins associated with plasma membrane rafts and also with endo-membranes, affecting both the plasma membrane dynamics and intracellular protein trafficking. To verify the above hypothesis, a set of shRNA was used to down-regulate flotillin-2 in Raw264 cells, which were found to also become deficient in flotillin-1. The flotillin deficiency inhibited strongly the TRIF-dependent endosomal signaling of LPS-activated TLR4, and to a lower extent also the MyD88-dependent one, without affecting the cellular level of TLR4. In contrast, the depletion of flotillins down-regulated the CD14 mRNA level and the total cellular content of CD14 protein, and decreased the amount of CD14 on the cell surface. The constitutive CD14 endocytosis remained unchanged but CD14 recycling was enhanced via EEA1-positive early endosomes and golgin-97-positive trans-Golgi network, likely to compensate for the depletion of the cell-surface CD14. Notably, a paucity of surface CD14 in resting cells can inhibit TLR4 signaling after the stimulation of cells with LPS. In conclusion, we have shown here that flotillins modulate the pro-inflammatory response of macrophages to LPS by affecting the abundance of CD14.

immunology↗