phalloidine and 2-phenyl-4-4-5-5-tetramethylimidazoline-1-oxyl-3-oxide

Activated microglial cells are an important pathological component in brains of patients with neurodegenerative diseases. The purpose of this study was to investigate the effect of He-Ne (632.8 nm, 64.6 mW/cm2) low-level laser therapy (LLLT), a non-damaging physical therapy, on activated microglia, and the subsequent signaling events of LLLT-induced neuroprotective effects and phagocytic responses.. To model microglial activation, we treated the microglial BV2 cells with lipopolysaccharide (LPS). For the LLLT-induced neuroprotective study, neuronal cells with activated microglial cells in a Transwell™ cell-culture system were used. For the phagocytosis study, fluorescence-labeled microspheres were added into the treated microglial cells to confirm the role of LLLT.. Our results showed that LLLT (20 J/cm2) could attenuate toll-like receptor (TLR)-mediated proinflammatory responses in microglia, characterized by down-regulation of proinflammatory cytokine expression and nitric oxide (NO) production. LLLT-triggered TLR signaling inhibition was achieved by activating tyrosine kinases Src and Syk, which led to MyD88 tyrosine phosphorylation, thus impairing MyD88-dependent proinflammatory signaling cascade. In addition, we found that Src activation could enhance Rac1 activity and F-actin accumulation that typify microglial phagocytic activity. We also found that Src/PI3K/Akt inhibitors prevented LLLT-stimulated Akt (Ser473 and Thr308) phosphorylation and blocked Rac1 activity and actin-based microglial phagocytosis, indicating the activation of Src/PI3K/Akt/Rac1 signaling pathway.. The present study underlines the importance of Src in suppressing inflammation and enhancing microglial phagocytic function in activated microglia during LLLT stimulation. We have identified a new and important neuroprotective signaling pathway that consists of regulation of microglial phagocytosis and inflammation under LLLT treatment. Our research may provide a feasible therapeutic approach to control the progression of neurodegenerative diseases.

Topics: Actins; Analysis of Variance; Animals; Animals, Newborn; Brain; Cells, Cultured; Chromones; Cyclic N-Oxides; Cytokines; Cytotoxicity Tests, Immunologic; Enzyme Inhibitors; Free Radical Scavengers; Gene Expression Regulation; Humans; Imidazoles; Lipopolysaccharides; Low-Level Light Therapy; Mice; Mice, Inbred C57BL; Microglia; Microscopy, Confocal; Morpholines; Myeloid Differentiation Factor 88; Neuroblastoma; Nitric Oxide; Phagocytosis; Phalloidine; Proto-Oncogene Proteins pp60(c-src); rac1 GTP-Binding Protein; Signal Transduction; Statistics as Topic; Time Factors; Transfection; Tyrosine