Prion diseases are neurodegenerative disorders characterized by the accumulation of misfolded prion protein, spongiform changes in the brain, and brain inflammation as a result of the wide-spread activation of microglia. Autophagy is a highly conserved catabolic process for the clearance of cytoplasmic components, including protein aggregates and damaged organelles; this process also eliminates pathological PrPSc as it accumulates during prion infection. The NALP3 inflammasome is a multiprotein complex that is a component of the innate immune system and is responsible for the release of pro-inflammatory cytokines. Our previous study showed that the neurotoxic prion peptide PrP106-126 induces NALP3 inflammasome activation and subsequent IL-1β release in microglia. Autophagy is involved in the regulation of the immune responses and inflammation in many diseases including neurodegenerative diseases. However, the relationship between autophagy and NALP3 inflammasome in prion diseases has not been investigated. In this study, we demonstrated that the processing and release of mature IL-1β is significantly enhanced by the inhibition of autophagy. Conversely, gene-silencing of the NALP3 inflammasome promotes autophagy. Suppression of TRIF or TLR4 by siRNA attenuated PrP106-126-induced autophagy, which is indicating that the TLR4-TRIF signaling pathway is involved in PrP106-26-induced autophagy. Caspase 1 directly cleaved TRIF to diminish TLR-4-TRIF mediated autophagy. Our findings suggest that the inhibition of autophagy by NALP3 inflammasome is probably mediated by activated Caspase-1-induced TRIF cleavage. This is the first study reporting that the NALP3 inflammasome complex negatively regulates autophagy in response to PrP106-126 stimulation in microglia, and partly explains the mechanism of autophagy inhibition by Caspase-1 in PrP106-126-induced BV2 cell activation. Our findings suggest that autophagy up-regulation and inhibition of Caspase-1 may protect against prion-induced neuroinflammation and accelerate misfolded protein degradation and are potential therapeutic approaches for prion diseases.

SummaryMitochondrial malfunction is a universal and critical step in the pathogenesis of many neurodegenerative diseases including prion diseases. Dynamin‐like protein 1 (DLP1) is one of the key regulators of mitochondrial fission. In this study, we investigated the role of DLP1 in mitochondrial fragmentation and dysfunction in neurons using in vitro and in vivo prion disease models. Mitochondria became fragmented and redistributed from axons to soma, correlated with increased mitochondrial DLP1 expression in murine primary neurons (N2a cells) treated with the prion peptide PrP106–126 in vitro as well as in prion strain‐infected hamster brain in vivo. Suppression of DLP1 expression by DPL1 RNAi inhibited prion‐induced mitochondrial fragmentation and dysfunction (measured by ADP/ATP ratio, mitochondrial membrane potential, and mitochondrial integrity). We also demonstrated that DLP1 RNAi is neuroprotective against prion peptide in N2a cells as shown by improved cell viability and decreased apoptosis markers, caspase 3 induced by PrP106–126. On the contrary, overexpression of DLP1 exacerbated mitochondrial dysfunction and cell death. Moreover, inhibition of DLP1 expression ameliorated PrP106–126‐induced neurite loss and synaptic abnormalities (i.e., loss of dendritic spine and PSD‐95, a postsynaptic scaffolding protein as a marker of synaptic plasticity) in primary neurons, suggesting that altered DLP1 expression and mitochondrial fragmentation are upstream events that mediate PrP106–126‐induced neuron loss and degeneration. Our findings suggest that DLP1‐dependent mitochondrial fragmentation and redistribution plays a pivotal role in PrPS c‐associated mitochondria dysfunction and neuron apoptosis. Inhibition of DLP1 may be a novel and effective strategy in the prevention and treatment of prion diseases.

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Bisphenol A (BPA) is toxic to the reproductive and nervous system, even carcinogenetic in humans and animals. However, few studies focused on effects of BPA on the intestinal tract. Here, we detected BPA‐induced injuries on intestinal mucosa and explored a reliable approach to counteract BPA effects. C57BL/6J mice were gavage BPA or BPA accompanied with ingestion of 4% (w/w) of glutamine for 4‐wks. In vitro, IEC‐6 cells were treated with 0.4 mmol/L BPA for 6 hours mimicking acute injury and 0.2 mmol/L BPA for 12 hours followed with or without the inclusion of 4 mmol/L glutamine for 12 hours to determine cell renewal, mitochondrial function and ROS‐JNK/MAPK pathway upon moderate BPA exposure. As results, BPA exposure caused severe intestinal injury, and disturbed intestinal epithelial cell proliferation and apoptosis, accompanied with mitochondrial malfunction and activated JNK/MAPK pathway as well. Notably, glutathione metabolism was implicated in BPA‐induce injury. Glutamine could well rescue cell renewal and mitochondrial function from BPA exposure‐induced injuries. In conclusion, we demonstrated impaired effect of BPA exposure on intestinal functions, which could be well counteracted by glutamine partly via restoring mitochondrial function and normalizing ROS‐JNK/MAPK pathway. Thereby, we provided a novel application of glutamine to rescue intestinal injury.

An experiment was conducted to 1) compare the regression and fat-free diet methods for estimating total or basal endogenous losses of fat (ELF) and fatty acids (ELFA) and true digestibility (TD) or standardized digestibility (SD) of fat and fatty acids in growing pigs and 2) compare these estimated values at the end of the ileum and over the entire intestinal tract. Ten barrows (initial body weight: 45.1 ± 2.8 kg) were surgically fitted with a T-cannula in the distal ileum, and allotted to 1 of 5 experimental diets in a 3-period Youden Square design. A fat-free diet was formulated using cornstarch, soy protein isolate and sucrose. Four oil-added diets were formulated by adding 4 levels of soybean oil (2, 4, 6 and 8%) to the fat-free basal diet at the expense of cornstarch. All diets contained 26% sugar beet pulp and 0.40% chromic oxide. Results indicated that here were no differences between true ileal digestibility (TID) of fat and true total tract digestibility (TTTD) of fat when pigs were fed soybean oil. The TID of C18:0 and total saturated fatty acids were greater than TTTD (P < 0.05). The total ELF at the end of the ileum was not different from that over the entire intestinal tract. In addition, total endogenous losses of C18:0 and total saturated fatty acids were greater for the entire intestinal tract than at the end of the ileum, whereas total endogenous losses of C18:2 and total unsaturated fatty acids were just the opposite. Similar results were observed for basal ELF and ELFA. As inclusion level of soybean oil increased, apparent digestibility of fat and fatty acids increased linearly (P < 0.05) except for apparent ileal digestibility of C18:0. However, SD of fat and fatty acids were not influenced by inclusion level of soybean oil. Estimation of ELF and ELFA observed by the regression and fat-free diet methods did not differ when measured at the end of the ileal or total tract. There were no differences between estimations of TD and SD of fat and fatty acids for soybean oil. Collectively, estimation of TD or SD of fat can be measured over the entire intestinal tract, whereas the ileal analysis method should be used to determine the ELF, ELFA and TD or SD of fatty acids. Correcting apparent digestibility for basal ELF and ELFA can accurately estimate SD values of fat and fatty acids.