NAMPT-derived NAD+ fuels PARP1 to promote skin inflammation through parthanatos cell death

Several studies have revealed a correlation between chronic inflammation and nicotinamide adenine dinucleotide (NAD+) metabolism, but the precise mechanism involved is unknown. Here, we report that the genetic and pharmacological inhibition of nicotinamide phosphoribosyltransferase (Nampt), the rate-limiting enzyme in the salvage pathway of NAD+ biosynthesis, reduced oxidative stress, inflammation, and keratinocyte DNA damage, hyperproliferation, and cell death in zebrafish models of chronic skin inflammation, while all these effects were reversed by NAD+ supplementation. Similarly, genetic and pharmacological inhibition of poly(ADP-ribose) (PAR) polymerase 1 (Parp1), overexpression of PAR glycohydrolase, inhibition of apoptosis-inducing factor 1, inhibition of NADPH oxidases, and reactive oxygen species (ROS) scavenging all phenocopied the effects of Nampt inhibition. Pharmacological inhibition of NADPH oxidases/NAMPT/PARP/AIFM1 axis decreased the expression of pathology-associated genes in human organotypic 3D skin models of psoriasis. Consistently, an aberrant induction of NAMPT and PARP activity, together with AIFM1 nuclear translocation, was observed in lesional skin from psoriasis patients. In conclusion, hyperactivation of PARP1 in response to ROS-induced DNA damage, fueled by NAMPT-derived NAD+, mediates skin inflammation through parthanatos cell death.

Prognostic and Immunological Role of Key Genes of Ferroptosis in Pan-Cancer

Solute carrier family 7 member 11 (SLC7A11), glutathione peroxidase 4 (GPX4), and apoptosis inducing factor mitochondria associated 2 (AIFM2) are the key regulators in ferroptosis. However, the expression patterns and prognostic roles of these genes in pan-cancer are still largely unclear. The expression patterns and prognostic roles of SLC7A11, GPX4, and AIFM2 and the relationships between the expression levels of these genes and immune infiltration levels in pan-cancer were analyzed by using TIMER, gene expression profiling interactive analysis (GEPIA), Oncomine, and Kaplan–Meier databases. Our results showed that both SLC7A11 and GPX4 were overexpressed in colorectal cancer, and SLC7A11 was overexpressed in lung cancer. High levels of SLC7A11 and AIFM2 were significantly linked with the shortened disease-free survival and overall survival (OS) in adrenocortical carcinoma (ACC), respectively. And high expression of SLC7A11, GPX4, and AIFM2 were significantly correlated with the shortened OS of acute myeloid leukemia patients. In esophageal carcinoma (ESCA), GPX4 expression was significantly associated with the infiltration of macrophage and myeloid dendritic cell, and AIFM2 expression was significantly associated with the infiltration of CD4+ T cell. Importantly, GPX4 expression was positively correlated with the expression levels of monocyte markers (CD14 and CD115) and M2 macrophage markers (VSIG4 and MS4A4A) both in ESCA and in head and neck squamous cell carcinoma (HNSC). In summary, SLC7A11, GPX4, and AIFM2 are dysregulated in many types of cancers, and are candidate prognostic biomarkers for many types of cancers, and can be used to evaluate the infiltration of immune cells in tumor tissues.

Bisphenol a Induces Autophagy Defects and AIF-Dependent Apoptosis via HO-1 and AMPK to Degenerate N2a Neurons

Bisphenol A (BPA) is an environmental contaminant widely suspected to be a neurological toxicant. Epidemiological studies have demonstrated close links between BPA exposure, pathogenetic brain degeneration, and altered neurobehaviors, considering BPA a risk factor for cognitive dysfunction. However, the mechanisms of BPA resulting in neurodegeneration remain unclear. Herein, cultured N2a neurons were subjected to BPA treatment, and neurotoxicity was assessed using neuronal viability and differentiation assays. Signaling cascades related to cellular self-degradation were also evaluated. BPA decreased cell viability and axon outgrowth (e.g., by down-regulating MAP2 and GAP43), thus confirming its role as a neurotoxicant. BPA induced neurotoxicity by down-regulating Bcl-2 and initiating apoptosis and autophagy flux inhibition (featured by nuclear translocation of apoptosis-inducing factor (AIF), light chain 3B (LC3B) aggregation, and p62 accumulation). Both heme oxygenase (HO)-1 and AMP-activated protein kinase (AMPK) up-regulated/activated by BPA mediated the molecular signalings involved in apoptosis and autophagy. HO-1 inhibition or AIF silencing effectively reduced BPA-induced neuronal death. Although BPA elicited intracellular oxygen free radical production, ROS scavenger NAC exerted no effect against BPA insults. These results suggest that BPA induces N2a neurotoxicity characterized by AIF-dependent apoptosis and p62-related autophagy defects via HO-1 up-regulation and AMPK activation, thereby resulting in neuronal degeneration.

Connexin Expression Is Altered in Liver Development of Yotari (dab1 -/-) Mice

Disabled-1 (Dab1) protein is an intracellular adaptor of reelin signaling required for prenatal neuronal migration, as well as postnatal neurotransmission, memory formation and synaptic plasticity. Yotari, an autosomal recessive mutant of the mouse Dab1 gene is recognizable by its premature death, unstable gait and tremor. Previous findings are mostly based on neuronal abnormalities caused by Dab1 deficiency, but the role of the reelin signaling pathway in nonneuronal tissues and organs has not been studied until recently. Hepatocytes, the most abundant cells in the liver, communicate via gap junctions (GJ) are composed of connexins. Cell communication disruption in yotari mice was examined by analyzing the expression of connexins (Cxs): Cx26, Cx32, Cx37, Cx40, Cx43 and Cx45 during liver development at 13.5 and 15.5 gestation days (E13.5 and E15.5). Analyses were performed using immunohistochemistry and fluorescent microscopy, followed by quantification of area percentage covered by positive signal. Data are expressed as a mean±SD and analyzed by one-way ANOVA. All Cxs examined displayed a significant decrease in yotari compared to wild type (wt) individuals at E13.5. Looking at E15.5 we have similar results with exception of Cx37 showing negligible expression in wt. Channels formation triggered by pathological stimuli, as well as propensity to apoptosis, was studied by measuring the expression of Pannexin1 (Panx1) and Apoptosis-inducing factor (AIF) through developmental stages mentioned above. An increase in Panx1 expression of E15.5 yotari mice, as well as a strong jump of AIF in both phases suggesting that yotari mice are more prone to apoptosis. Our results emphasize the importance of gap junction intercellular communication (GJIC) during liver development and their possible involvement in liver pathology and diagnostics where they can serve as potential biomarkers and drug targets.

Staurosporine-induced cleavage of apoptosis-inducing factor in human fibrosarcoma cells is independent of matrix metalloproteinase-2

Apoptosis-inducing factor (AIF) is a mitochondrial flavoprotein which mediates staurosporine (STS)-induced cell death. AIF cleavage and translocation to the cytosol is thought to be calpain-1-dependent as calpain inhibitors reduced AIF proteolysis. However, many calpain inhibitors also inhibit matrix metalloproteinase-2 (MMP-2) activity, an intracellular and extracellular protease implicated in apoptosis. Here we investigated whether MMP-2 activity is affected in response to STS and if contributes to AIF cleavage. Human fibrosarcoma HT1080 cells were treated with STS (0.1 µM, 0.25-24 hr). A significant increase in cellular MMP-2 activity was seen by gelatin zymography after 6 hr STS treatment, prior to induction of cell necrosis. Western blot showed the time-dependent appearance of two forms of AIF (~60 and 45 kDa) in the cytosol which were significantly increased at 6 hr. Surprisingly, knocking down MMP-2 or inhibiting its activity with MMP-2 preferring inhibitors ARP-100 or ONO-4817, or inhibiting calpain activity with ALLM or PD150606, did not prevent the STS-induced increase in cytosolic AIF. These results show that although STS rapidly increases MMP-2 activity, the cytosolic release of AIF may be independent of the proteolytic activities of MMP-2 or calpain.

Molecular characterization of a complex of apoptosis-inducing factor 1 with cytochrome c oxidase of the mitochondrial respiratory chain

Combining mass spectrometry–based chemical cross-linking and complexome profiling, we analyzed the interactome of heart mitochondria. We focused on complexes of oxidative phosphorylation and found that dimeric apoptosis-inducing factor 1 (AIFM1) forms a defined complex with ∼10% of monomeric cytochrome c oxidase (COX) but hardly interacts with respiratory chain supercomplexes. Multiple AIFM1 intercross-links engaging six different COX subunits provided structural restraints to build a detailed atomic model of the COX-AIFM12 complex (PDBDEV_00000092). An application of two complementary proteomic approaches thus provided unexpected insight into the macromolecular organization of the mitochondrial complexome. Our structural model excludes direct electron transfer between AIFM1 and COX. Notably, however, the binding site of cytochrome c remains accessible, allowing formation of a ternary complex. The discovery of the previously overlooked COX-AIFM12 complex and clues provided by the structural model hint at potential roles of AIFM1 in oxidative phosphorylation biogenesis and in programmed cell death.