alpha-synuclein and Ischemia

The present study aimed to investigate the mechanism of injured neurons caused by ischemia/reperfusion in the induction of microglia activation. Spinal neurons were prepared and exposed to ischemic/reperfused conditions. The α-synuclein protein levels in these cells were analyzed by western blot, immunofluorescence, or enzyme-linked immunosorbent assay. Ischemia/reperfusion exposure led to elevated α-synuclein protein expression and release. Furthermore, when cocultured with injured neurons or supernatants from injured neurons, nitric oxide generation, H2O2 production, and tumor necrosis factor-α expression were promoted in microglia. Nevertheless, this effect was impeded by pretreatment of the α-synuclein antibody in the supernatants from injured neurons. Moreover, toll-like receptor 2 (TLR2) rather than TLR3 or TLR4 mediated microglia activation by α-synuclein. This process involved p38 MAPK and NF-κB activation, the inhibition of which resulted in reduced NADPH oxidase 2 (Nox2) in microglia. In conclusion, ischemia/reperfusion-injured neurons could express and release increased levels of α-synuclein and cause microglia activation through TLR2 both in vitro and in vivo.

Topics: alpha-Synuclein; Animals; Animals, Newborn; Antibodies; Cells, Cultured; Coculture Techniques; Embryo, Mammalian; Enzyme-Linked Immunosorbent Assay; Gene Expression Regulation; Hydrogen Peroxide; Ischemia; Microglia; Neurons; Nitric Oxide; Rats; Rats, Sprague-Dawley; Reperfusion Injury; Spinal Cord; Time Factors; Toll-Like Receptor 2; Tumor Necrosis Factor-alpha

We have previously demonstrated that alpha-synuclein (Snca) gene ablation reduces brain arachidonic acid (20:4n-6) turnover rate in phospholipids through modulation of endoplasmic reticulum-localized acyl-CoA synthetase activity. Although 20:4n-6 is a precursor for prostaglandin (PG), Snca effect on PG levels is unknown. In the present study, we examined the effect of Snca ablation on brain PG level at basal conditions and following 30s of global ischemia. Brain PG were extracted with methanol, purified on C(18) cartridges, and analyzed by LC-MS/MS. We demonstrate, for the first time, that Snca gene ablation did not affect brain PG mass under normal physiological conditions. However, total PG mass and masses of individual PG were elevated approximately 2-fold upon global ischemia in the absence of Snca. These data are consistent with our previously observed reduction in 20:4n-6 recycling through endoplasmic reticulum-localized acyl-CoA synthetase in the absence of Snca, which may result in the increased 20:4n-6 availability for PG production in the absence of Snca during global ischemia and suggest a role for Snca in brain inflammatory response.

Topics: alpha-Synuclein; Analysis of Variance; Animals; Chromatography, Liquid; Gene Expression Regulation; Ischemia; Male; Mice; Mice, Knockout; Prostaglandins; Tandem Mass Spectrometry