TDP43-Mediated Microglial Activation through NF-κB, AP-1 and NLRP3 Inflammasome (P02.172)

OBJECTIVE: To define the effects of TDP43 on microglia and whether they contribute to the underlying mechanisms of neurotoxicity in amyotrophic lateral sclerosis (ALS). BACKGROUND: Increasing evidence supports the role of microglial activation on motoneuron death and disease progression in ALS. Transactive response DNA-binding protein 43 (TDP43) has been identified as a critical disease-related protein in ALS. However, the mechanisms whereby TDP43 contribute to neurotoxicity remain unknown. DESIGN/METHODS: The effects of TDP43 on microglia were determined by using primary cultures. Microglial activation and toxicity was determined by measuring NOX2, TNF-α and IL-1β using quantitative RT-PCR and ELISA assays. Western blot was performed to detect IκBα and p65 in NF-κB pathway. Active Caspase 1 in microglia was examined using fluorescent probe FAM-YVAD-FMK. RESULTS: TDP43 proteins, either wild-type (WT), 25kD C-terminal fragments, or mutant forms, were able to activate microglia by up-regulating NOX2 (major source of superoxide free radical) and cytotoxic cytokine production (TNF-α and IL-1β). PMBS, a LPS inhibitor, did not block TDP43-induced microglial activation, indicating that potential LPS contamination in the protein preparation can be excluded. TDP43 treatment resulted in activation of NF-κB pathway in microglia by decreasing IκBα protein levels, and subsequently enhancing p65 phosphorylation. The microglial response to TDP-43 was dependent on the CD14 receptor; CD14 blocking antibodies suppressed the TDP43-mediated NF-κB activation and pro-inflammatory cytokine production in microglia. AP-1 transcription factor was also involved in TNF-α and IL-1β production, which appeared to contribute to translational regulation. NLRP3 was upregluated, and active Caspase 1 was increased in TDP43-treated microglia, indicating that TDP43 also activated NLRP3 inflammasome in microglia. CONCLUSIONS: TDP-43 interacted with CD14 receptor and induced microglial cytotoxic activation through NF-κB/AP-1 pathways and NLRP3 inflammasome. This study suggests that efforts to block TDP43-induced microglial cytotoxicity may offer a therapeutic option for ALS. Supported by: NIH, Muscular Dystrophy Association. Disclosure: Dr. Zhao has nothing to disclose. Dr. Bell has nothing to disclose. Dr. Beers has nothing to disclose. Dr. Liao has nothing to disclose. Dr. Henkel has nothing to disclose. Dr. Baloh has nothing to disclose. Dr. Appel has received personal compensation for activities with Neuraltus Pharmaceuticals, Inc. as a member of their Scientific Advisory Board.