Mitochondrial fission and selective mitochondrial autophagy (mitophagy) form an essential axis of mitochondrial quality control that plays a critical role in the development of cardiac ischemia-reperfusion (IR) injury. However, the precise upstream molecular mechanism of fission/mitophagy remains unclear. Dual-specificity protein phosphatase1 (DUSP1) regulates cardiac metabolism, but its physiological contribution in the reperfused heart, particularly its influence on mitochondrial homeostasis, is unknown. Here, we demonstrated that cardiac DUSP1 was downregulated following acute cardiac IR injury. In vivo, compared to wild-type mice, DUSP1 transgenic mice (DUSP1TG mice) demonstrated a smaller infarcted area and the improved myocardial function. In vitro, the IR-induced DUSP1 deficiency promoted the activation of JNK which upregulated the expression of the mitochondrial fission factor (Mff). A higher expression level of Mff was associated with elevated mitochondrial fission and mitochondrial apoptosis. Additionally, the loss of DUSP1 also amplified the Bnip3 phosphorylated activation via JNK, leading to the activation of mitophagy. Increased mitophagy overtly consumed mitochondrial mass resulting into the mitochondrial metabolism disorder. However, the reintroduction of DUSP1 blunted Mff/Bnip3 activation and therefore alleviated the fatal mitochondrial fission/mitophagy by inactivating the JNK pathway, providing a survival advantage to myocardial tissue following IR stress. The results of our study suggest that DUSP1 and its downstream JNK pathway are therapeutic targets for conferring protection against IR injury by repressing Mff-mediated mitochondrial fission and Bnip3-required mitophagy.

Increased mitochondrial damage is related to the progression of a diet-induced nonalcoholic fatty liver disease. The aim of our study is to investigate the role of Sirtuin 3 (Sirt3) in treating nonalcoholic fatty liver disease with a focus on mitophagy and the ERK-CREB pathway. Our data indicated that Sirt3 was downregulated in liver tissue in response to chronic HFD treatment. Interestingly, re-introduction of Sirt3 protected hepatic function, attenuated liver fibrosis, alleviated the inflammatory response, and prevented hepatocyte apoptosis. Molecular investigations demonstrated that lipotoxicity was associated with an increase in mitochondrial apoptosis as evidenced by reduced mitochondrial potential, augmented ROS production, increased cyt-c leakage into the nucleus, and activated caspase-9 apoptotic signalling. Additionally, Sirt3 overexpression protected hepatocytes against mitochondrial apoptosis via promoting Bnip3-required mitophagy. Functional studies showed that Sirt3 reversed Bnip3 expression and mitophagy activity via the ERK-CREB signalling pathway. Blockade of the ERK-CREB axis repressed the promotive effects of Sirt3 on Bnip3 activation and mitophagy augmentation, finally negating the anti-apoptotic influences of Sirt3 on hepatocytes in the setting of high-fat-stress. Collectively, our data show that high-fat-mediated liver damage is associated with Sirt3 downregulation, which is followed by ERK-CREB pathway inactivation and Bnip3-mediated inhibition of mitophagy, causing hepatocytes to undergo mitochondria-dependent cell death. Based on this, strategies for enhancing Sirt3 activity and activating the ERK-CREB-Bnip3-mitophagy pathways could be used to treat nonalcoholic fatty liver disease.

Engineered carbonaceous nanomaterials (ECNs), including single-wall carbon nanotubes (SWCNTs), multi-wall carbon nanotubes (MWCNTs), graphene and graphene oxide (GO), are potentially hazardous to the lung. With incremental experience in the use of predictive toxicological approaches, seeking to relate ECN physicochemical properties to adverse outcome pathways (AOPs), it is logical to explore the existence of a common AOP that allows comparative analysis of broad ECN categories. We established an ECN library comprised of three different types of SWCNTs, graphene and graphene oxide (two sizes) for comparative analysis according to a cell-based AOP that also plays a role in the pathogenesis of pulmonary fibrosis. SWCNTs synthesized by Hipco, arc discharge and Co-Mo catalyst (CoMoCAT®) methods were obtained in their as-prepared (AP) state, following which they were further purified (PD) or coated with Pluronic (PF108) or bovine serum albumin (BSA) to improve dispersal and colloidal stability. Graphene oxide (GO) was prepared as two sizes, GO-small (S) and GO-large (L), while the graphene samples were coated with BSA and PF108 to enable dispersion in aqueous solution. In vitro screening showed that AP- and PD-SWCNTs, irrespective of the method of synthesis, as well as graphene (BSA) and GO (S and L) could trigger interleukin 1β (IL-1β) and transforming growth factor (TGF-β1) production in myeloid (THP-1) and epithelial (BEAS-2B) cell lines, respectively. Oropharyngeal aspiration in mice confirmed that AP-Hipco tubes, graphene (BSA), GO-S and GO-L could induce IL-1β and TGF-β1 production in the lung in parallel with lung fibrosis. Notably, GO-L was the most pro-fibrogenic material based on rapid kinetics of pulmonary injury. In contrast, PF108-dispersed SWCNTs and -graphene failed to exert fibrogenic effects. Collectively, these data indicate that the dispersal state and surface reactivity of ECNs play key roles in triggering a pro-fibrogenic AOP, which could prove helpful for hazard ranking and a proposed tiered testing approach for large ECN categories.

Engineered nanomaterials (ENMs) continue to attract significant attentions because they have novel physicochemical properties that can improve the functions of products that will benefit human lives. However, the physicochemical properties that make ENMs attractive could interact with biological systems and induce cascades of events that cause toxicological effects. Recently, there are more studies suggesting inflammasome activation may play an important role in ENMs-induced biological responses. Inflammasomes are a family of multi-protein complexes and are increasingly recognized as major mediators of host immune system. Among these, NLRP3 inflammasome is the most studied one that could directly interact with ENMs to generate inflammatory responses. In this review, we aim to link the ENM physicochemical properties to NLRP3 inflammasome activation. The understanding of the mechanisms of ENMs-NLRP3 inflammasome interaction will provide us strategies for safer nanomaterial design and therapy.

We demonstrate through PdO doping that creation of heterojunctions on Co3O4 nanoparticles can quantitatively adjust band-gap and Fermi energy levels to study the impact of metal oxide nanoparticle semiconductor properties on cellular redox homeostasis and hazard potential. Flame spray pyrolysis (FSP) was used to synthesize a nanoparticle library in which the gradual increase in the PdO content (0–8.9%) allowed electron transfer from Co3O4 to PdO to align Fermi energy levels across the heterojunctions. This alignment was accompanied by free hole accumulation at the Co3O4 interface and production of hydroxyl radicals. Interestingly, there was no concomitant superoxide generation, which could reflect the hole dominance of a p-type semiconductor. Although the electron flux across the heterojunctions induced upward band bending, the Ec levels of the doped particles showed energy overlap with the biological redox potential (BRP). This allows electron capture from the redox couples that maintain the BRP from −4.12 to −4.84 eV, causing disruption of cellular redox homeostasis and induction of oxidative stress. PdO/Co3O4 nanoparticles showed significant increases in cytotoxicity at 25, 50, 100, and 200 μg/mL, which was enhanced incrementally by PdO doping in BEAS-2B and RAW 264.7 cells. Oxidative stress presented as a tiered cellular response involving superoxide generation, glutathione depletion, cytokine production, and cytotoxicity in epithelial and macrophage cell lines. A progressive series of acute pro-inflammatory effects could also be seen in the lungs of animals exposed to incremental PdO-doped particles. All considered, generation of a combinatorial PdO/Co3O4 nanoparticle library with incremental heterojunction density allowed us to demonstrate the integrated role of Ev, Ec, and Ef levels in the generation of oxidant injury and inflammation by the p-type semiconductor, Co3O4.

FUN14 domain-containing protein 1 (Fundc1)-dependent mitophagy, mainly activated by ischemic/hypoxic preconditioning, benefits acute myocardial reperfusion injury and chronic metabolic syndrome via sustaining mitochondrial homeostasis. Mitochondrial fission plays a pathogenic role in ischemic acute kidney injury (AKI) through perturbation of mitochondrial quality and activation of mitochondrial apoptosis. The aim of our study was to explore the role of Fundc1 mitophagy in ischemia preconditioning (IPC)-mediated renoprotection. Proximal tubule-specific Fundc1 knockout (Fundc1PTKO) mice were subjected to ischemia reperfusion injury (IRI) and IPC prior to assessment of renal function, mitophagy, mitochondrial quality control, and Drp1-related mitochondrial fission. Following exposure to IPC, Fundc1 mitophagy was activated through post-transcriptional phosphorylation at Ser17. Interestingly, IRI-mediated renal injury, inflammation, and tubule cell death were mitigated by IPC whereas proximal tubule-specific Fundc1 knockout (Fundc1PTKO) mice abolished IPC-offered renoprotection. Mechanistically, IRI-evoked mitochondrial damage was improved by IPC whereas Fundc1 deficiency provoked mitochondrial abnormality, manifested by impaired mitochondrial quality and hyperactivated Drp1-dependent mitochondrial fission. Interestingly, Fundc1 deficiency-associated mitochondrial dysfunction was reversed by pharmacological inhibition of mitochondrial fission. In vivo, Fundc1 deletion-caused renal injury, severe pro-inflammatory response, and tubule cell death could be nullified by way of knockout Drp1 on Fundc1PTKO background. Finally, we also revealed that IPC triggered Fundc1 mitophagy activation through UNC-51-like kinase 1 (Ulk1) and Ulk1 ablation interrupted IPC-mediated Fundc1 activation and thus attenuated IPC-induced renoprotection. Fundc1 mitophagy, primarily driven by IPC, confers resistance to AKI through reconciliation of mitochondrial fission, implicating the therapeutic potential of targeting mitochondrial homeostasis for AKI.

Engineered nanomaterials (ENMs) including multiwall carbon nanotubes (MWCNTs) and rare earth oxide (REO) nanoparticles, which are capable of activating the NLRP3 inflammasome and inducing IL-1β production, have the potential to cause chronic lung toxicity. Although it is known that lysosome damage is an upstream trigger in initiating this pro-inflammatory response, the same organelle is also an important homeostatic regulator of activated NLRP3 inflammasome complexes, which are engulfed by autophagosomes and then destroyed in lysosomes after fusion. Although a number of ENMs have been shown to induce autophagy, no definitive research has been done on the homeostatic regulation of the NLRP3 inflammasome during autophagic flux. We used a myeloid cell line (THP-1) and bone marrow derived macrophages (BMDM) to compare the role of autophagy in regulating inflammasome activation and IL-1β production by MWCNTs and REO nanoparticles. THP-1 cells express a constitutively active autophagy pathway and are also known to mimic NLRP3 activation in pulmonary macrophages. We demonstrate that, while activated NLRP3 complexes could be effectively removed by autophagosome fusion in cells exposed to MWCNTs, REO nanoparticles interfered in autophagosome fusion with lysosomes. This leads to the accumulation of the REO-activated inflammasomes, resulting in robust and sustained IL-1β production. The mechanism of REO nanoparticle interference in autophagic flux was clarified by showing that they disrupt lysosomal phosphoprotein function and interfere in the acidification that is necessary for lysosome fusion with autophagosomes. Binding of LaPO4 to the REO nanoparticle surfaces leads to urchin-shaped nanoparticles collecting in the lysosomes. All considered, these data demonstrate that in contradistinction to autophagy induction by some ENMs, specific materials such as REOs interfere in autophagic flux, thereby disrupting homeostatic regulation of activated NLRP3 complexes.

The safe implementation of nanotechnology requires nanomaterial hazard assessment in accordance with the material physicochemical properties that trigger the injury response at the nano/bio interface. Since CuO nanoparticles (NPs) are widely used industrially and their dissolution properties play a major role in hazard potential, we hypothesized that tighter bonding of Cu to Fe by particle doping could constitute a safer-by-design approach through decreased dissolution. Accordingly, we designed a combinatorial library in which CuO was doped with 1–10% Fe in a flame spray pyrolysis (FSP) reactor. The morphology and structural properties were determined by XRD, BET, Raman spectroscopy, HRTEM, EFTEM and EELS, which demonstrated a significant reduction in the apical Cu-O bond length while simultaneously increasing the planar bond length (Jahn-Teller distortion). Hazard screening was performed in tissue culture cell lines and zebrafish embryos to discern the change in the hazardous effects of doped vs. non-doped particles. This demonstrated that with increased levels of doping, there was a progressive decrease in cytotoxicity in BEAS-2B and THP-1 cells, as well as an incremental decrease in the rate of hatching interference in zebrafish embryos. The dissolution profiles were determined and the surface reactions taking place in Holtfreter’s solution were validated using cyclic voltammetry (CV) measurements to demonstrate the Cu+/Cu2+ and Fe2+/Fe3+ redox species playing a major role in the dissolution process of pure and Fe doped CuO. Altogether, a safe-by-design strategy was implemented for the toxic CuO particles via Fe doping and has been demonstrated for their safe use in the environment.