Objective: To investigate the effect of TNF on α-synuclein degradation and to explore the potential role of autophagy-lysosome pathway involved in this processs.

Background: In PD and other neurodegenerative disorders, protein misfolding and aggregation is a common pathological feature. Recently TNF have been revealed to contribute to amyloid beta precursor protein processing and beta amyloid production in Alzheimer’s disease. However, it remains poorly understood whether TNF affects α-SYN accumulation/aggregation.

Methods: The contents of TNF and IL-1β in the microglia culture supernatants were determined with ELISA.The cell viability was surveiedied by CCK-8.Western blot and PCR were used to detect the protein and transcriptional level of autophagy related protein.RFP-GFP-tandem fluorescent tagged LC3 (tf-LC3) transfection were emploied to evaluate the function of autophagy. LysoSensor Yellow/Blue dextran were used as a ratiometric pH probe.

Results: The content of IL-1β and TNF were both significantly higher after LPS treatment. α-synuclein levels of conditional media group was significantly higher; PC12 cell viability was concentration-dependent decreased by TNF-α. α-synuclein protein levels showed a concentration-dependent increased and then decreased, mRNA levels of α-synuclein and TNF-α concentrations had no correlation; After the proteasome inhibitor MG132 inhibited UPS pathway, the protein levels of α-synuclein did not change obviously, while using autophagy blocker CQ inhibited ALP, the protein levels of α-synuclein increased significantly. The protein levels of LC3-II and p62 appeared synchronization concentration-dependent increased after TNF-α treatment, and the mRNA p62 did not change. TNF treatment led to an obvious increase in the number of yellow when the cells were transfected with RFP-GFP-tandem fluorescent LC3.LysoTracker fluorescence intensity decreased, lysosomal pH rised; mTORC1 inhibitor PP242 allows lysosomal acidification, the up-regulation of TFEB and LAMP1, lysosomal pH decreased, promote degradation of α-synuclein.

Conclusions: In sum, our study demonstrates that the pro-inflammatory cytokine TNF reduced α-SYN degradation by disrupting lysosome function, and thus caused α-SYN accumulation in DA cells. Our findings reveal a novel effect of TNF on neurons and highlight a role of autophagy in neuroinflammation-related DA neuron degeneration in PD.

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MDS Abstracts - https://www.mdsabstracts.org/abstract/tnf-compromises-lysosome-acidification-and-reduces-synuclein-degradation-via-autophagy-in-dopaminergic-cells/