Background
Parkinson's disease (PD) is characterized by intraneuronal α-synuclein aggregation called Lewy bodies and progressive dopaminergic neurodegeneration. Toll-like receptor (TLR) signaling is a major pathway mediating inflammation. The molecular link on how neuroinflammation upregulates neuronal TLRs and induces accumulation of α-synuclein aggregates to drive synucleinopathy remains to be determined.
Conclusion
Activated microglia-derived neuroinflammation induced neuronal TLR2/4-p38/JNK activation to perturb autophagy, causing accumulation of α-synuclein aggregates and neurite shortening. Targeting neuronal TLR2/4 pathway might be a mechanistic-based therapy for neurodegenerative disease, such as PD.
Methods
Genetic variants of TLRs in PD patients were genotyped and the multiplex cytokines, sRAGE, and HMGB1were assessed. A coculture system was employed to measure α-synuclein aggregates and neurite shortening by confocal microscope. The expression of TLR2/4 and autophagy flux was detected by western blot and immunofluorescence.
Objective
Despite conditioned medium from microglia and TLR agonists were utilized to study their effects on neuronal cells, a Transwell coculture system, comprising lipopolysaccharide-activated microglia on top and retinoic acid-differentiated SH-SY5Y cells at the bottom more mimicking in vivo neuroinflammation, was employed to elucidate the mechanism of activated microglia on neuronal cells.
Results
PD patients showed higher plasma levels of proinflammatory cytokines and genetic TLR4 variant, c.896 A > G (p. D299G). Elevated proinflammatory cytokines in coculture medium was also seen. Phosphorylation and aggregation of α-synuclein, shortening of neurite, upregulation of TLR2/4 expression, activation of downstream p38 and JNK, and dampening of autophagic flux were seen in SH-SY5Y cells cocultured with activated microglia. Those were prevented by inhibiting TLR2/4 and p38/JNK signaling.