Increasing evidence suggests the important role of metabolic reprogramming in the regulation of the innate inflammatory response, but the underlying mechanism remains unclear. Here, we provide evidence to support a novel role for the pyruvate kinase M2 (PKM2)-mediated Warburg effect, namely aerobic glycolysis, in the regulation of high mobility group box 1 (HMGB1) release. PKM2 interacts with hypoxia-inducible factor 1α (HIF1α) and activates the HIF-1α-dependent transcription of enzymes necessary for aerobic glycolysis in macrophages. Knockdown of PKM2, HIF1α, and glycolysis-related genes uniformly decreases lactate production and HMGB1 release. Similarly, a potential PKM2 inhibitor, shikonin, reduces serum lactate and HMGB1 levels and protects mice from lethal endotoxemia and sepsis. Collectively, these findings shed light on a novel mechanism for metabolic control of inflammation by regulating HMGB1 release and highlight the importance of targeting aerobic glycolysis in the treatment of sepsis and other inflammatory diseases.
System restoration from cascading failures is an integral part of the overall defense against catastrophic breakdown in networked critical infrastructures. From the outbreak of cascading failures to the system complete breakdown, actions can be taken to prevent failure propagation through the entire network. While most analysis efforts have been carried out before or after cascading failures, restoration during cascading failures has been rarely studied. In this paper, we present a modeling framework to investigate the effects of in-process restoration, which depends strongly on the timing and strength of the restoration actions. Furthermore, in the model we also consider additional disturbances to the system due to restoration actions themselves. We demonstrate that the effect of restoration is also influenced by the combination of system loading level and restoration disturbance. Our modeling framework will help to provide insights on practical restoration from cascading failures and guide improvements of reliability and resilience of actual network systems.
Cell death and inflammation are key pathologic responses of acute pancreatitis (AP), the leading cause of hospital admissions for gastrointestinal disorders. It is becoming increasingly clear that damage-associated molecular pattern molecules (DAMPs) play an important role in the pathogenesis of AP by linking local tissue damage to systemic inflammation syndrome. Endogenous DAMPs released from dead, dying or injured cells initiate and extend sterile inflammation via specific pattern recognition receptors. Inhibition of the release and activity of DAMPs (for example, high mobility group box 1, DNA, histones and adenosine triphosphate) provides significant protection against experimental AP. Moreover, increased serum levels of DAMPs in patients with AP correlate with disease severity. These findings provide novel insight into the mechanism, diagnosis and management of AP. DAMPs might be an attractive therapeutic target in AP.
High mobility group box 1 (HMGB1) is an evolutionarily ancient protein that is present in one form or another in all eukaryotes. It fundamentally resides in the nucleus but translocates to the cytosol with stress and is subsequently released into the extracellular space. HMGB1 global knockout mice exhibit lethal hypoglycemia, whereas tissues and cells from conditional knockout or knock-in mice are born alive without apparent significant functional deficit. An aberrant response to targeted stress in the liver, pancreas, heart or myeloid cells is consistent with a protective role for HMGB1 in sustaining nuclear homeostasis and enabling other stress responses, including autophagy. Under some conditions, HMGB1 is not required for liver and heart function. Many challenges remain with respect to understanding the multiple roles of HMGB1 in health and disease.
Sepsis is caused by an overwhelming immune response to bacterial infection. The discovery of high mobility group box 1 (HMGB1) as a late mediator of lethal sepsis has prompted investigation into the development of new therapeutics which specifically target this protein. Here, we show that chloroquine, an anti-malarial drug, prevents lethality in mice with established endotoxemia or sepsis. This effect is still observed even if administration of chloroquine is delayed. The protective effects of chloroquine were mediated through inhibition of HMGB1 release in macrophages, monocytes, and endothelial cells, thereby preventing its cytokine-like activities. As an inhibitor of autophagy, chloroquine specifically inhibited HMGB1-induced Iκ-B degradation and NF-κB activation. These findings define a novel mechanism for the anti-inflammatory effects of chloroquine and also suggest a new potential clinical use for this drug in the setting of sepsis.
HMGB1; chloroquine; sepsis; autophagy; NF-κB; Beclin 1
Forty years ago, high mobility group box 1 (HMGB1) was discovered in calf thymus and named according to its electrophoretic mobility in polyacrylamide gels. Now, we know that HMGB1 performs dual functions. Inside the cell, HMGB1 is a highly conserved chromosomal protein acting as a DNA chaperone. Outside of the cell, HMGB1 is a prototypical damage-associated molecular pattern, acting with cytokine, chemokine, and growth factor. During tumor development and in cancer therapy, HMGB1 has been reported to play paradoxical roles in promoting both cell survival and death by regulating multiple signaling pathways, including inflammation, immunity, genome stability, proliferation, metastasis, metabolism, apoptosis, and autophagy. Here, we review the current knowledge of both HMGB1’s oncogenic and tumor suppressive roles and the potential strategies that target HMGB1 for the prevention and treatment of cancer.
Pancreatic ductal adenocarcinoma (PDA) has an aggressive natural history and is resistant to therapy. The receptor for advanced glycation end-products (RAGE) is a pattern recognition receptor for many damage associated molecular pattern (DAMP) molecules. RAGE is overexpressed in both human and murine models of PDA as well as most advanced epithelial neoplasms. The immunosuppressive nature of the PDA micro-environment is facilitated, in part, by the accumulation of regulatory immune cell infiltrates such as myeloid-derived suppressor cells (MDSCs). To study the role of RAGE expression in the setting of mutant Ras-promoted pancreatic carcinogenesis (KC), a triple transgenic model of spontaneous murine PDA in a RAGE-null background (KCR) was generated. KCR mice had markedly delayed pancreatic carcinogenesis and a significant diminution of MDSCs compared to KC mice at comparable time points post weaning. While RAGE was not required for the development or suppressor activity of MDSCs, its absence was associated with temporally limited pancreatic neoplasia and altered phenotype and function of the myeloid cells. In lieu of MDSCs, KCR animals at comparable time points exhibited mature CD11b+Gr1−F4/80+ cells which were not immunosuppressive in vitro. KCR mice also maintained a significantly less suppressive milieu evidenced by marked decreases in CCL22 in relation to CXCL10 and diminished serum levels of IL-6.
Rodent; Monocytes/Macrophages; Inflammation; Tolerance/Suppression/Anergy; Tumor Immunity; Transgenic/Knockout Mice
Autophagy is a double-edged sword in tumorigenesis and plays an important role in the resistance of cancer cells to chemotherapy. S100A8 is a member of the S100 calcium-binding protein family and plays an important role in the drug resistance of leukemia cells, with the mechanisms largely unknown. Here we report that S100A8 contributes to drug resistance in leukemia by promoting autophagy. S100A8 level was elevated in drug resistance leukemia cell lines relative to the nondrug resistant cell lines. Adriamycin and vincristine increased S100A8 in human leukemia cells, accompanied with upregulation of autophagy. RNA interference-mediated knockdown of S100A8 restored the chemosensitivity of leukemia cells, while overexpression of S100A8 enhanced drug resistance and increased autophagy. S100A8 physically interacted with the autophagy regulator BECN1 and was required for the formation of the BECN1-PI3KC3 complex. In addition, interaction between S100A8 and BECN1 relied upon the autophagic complex ULK1-mAtg13. Furthermore, we discovered that exogenous S100A8 induced autophagy, and RAGE was involved in exogenous S100A8-regulated autophagy. Our data demonstrated that S100A8 is involved in the development of chemoresistance in leukemia cells by regulating autophagy, and suggest that S100A8 may be a novel target for improving leukemia therapy.
Autophagy is a lysosome-mediated catabolic process involving the degradation of intracellular contents (e.g., proteins and organelles) as well as invading microbes (e.g., parasites, bacteria and viruses). Multiple forms of cellular stress can stimulate this pathway, including nutritional imbalances, oxygen deprivation, immunological response, genetic defects, chromosomal anomalies and cytotoxic stress. Damage-associated molecular pattern molecules (DAMPs) are released by stressed cells undergoing autophagy or injury, and act as endogenous danger signals to regulate the subsequent inflammatory and immune response. A complex relationship exists between DAMPs and autophagy in cellular adaption to injury and unscheduled cell death. Since both autophagy and DAMPs are important for pathogenesis of human disease, it is crucial to understand how they interplay to sustain homeostasis in stressful or dangerous environments.
autophagy; DAMP; stress; HMGB1; ATP; IL1B; injury
Colorectal cancer (CRC) remains a major worldwide cause of cancer-related morbidity and mortality largely due to the insidious onset of the disease. The current clinical procedures utilized for disease diagnosis are invasive, unpleasant, and inconvenient. Hence, the need for simple blood tests that could be used for the early detection is crucial for its ultimate control and prevention.
The present work is a case–control study focused on proteomic analysis of serum of healthy volunteers and CRC patients by the ClinProt profiling technology based on mass spectrometry. This approach allowed to identifying a pattern of proteins/peptides able to differentiate the studied populations. Moreover, some of peptides differentially expressed in the serum of patients as compared to healthy volunteers were identified by LTQ Orbitrap XL.
A Quick Classifier Algorithm was used to construct the peptidome patterns (m/z 1208, 1467, 1505, 1618, 1656 and 4215) for the identification of CRC from healthy volunteers with accuracy close to 100% (>CEA, P < 0.05). Peaks at m/z 1505 and 1618 were identified as alpha-2-HS-glycoprotein precursor and tubulin beta chain, respectively.
Alpha-2-HS-glycoprotein precursor and tubulin beta chain could be involved in the pathogenesis of CRC and perform as potential serology diagnosis biomarker.
The virtual slide(s) for this article can be found here: http://www.diagnosticpathology.diagnomx.eu/vs/4796578761089186.
Colorectal neoplasms; Diagnosis; Biological markers; Proteomics; Alpha-2-HS-glycoprotein; Tubulin beta chain
Autophagy is an evolutionarily conserved pathway responsible for delivery of cytoplasmic material into the lysosomal degradation pathway to enable vesicular exocytosis. Interleukin (IL)-2 is produced by T-cells and its activity is important for immunoregulation. Fibroblasts are an immune competent cell type, playing a critical role in wound healing, chronic inflammation, and tumor development. Although autophagy plays an important role in each of these processes, whether it regulates IL-2 activity in fibroblasts is unknown. Here, we show that autophagy is required for IL-2-induced cell growth in fibroblasts. IL-2 significantly induced autophagy in mouse embryonic fibroblasts (MEFs) and primary lung fibroblasts. Autophagy inhibitors (e.g., 3-methylamphetamine and bafilomycin A1) or knockdown of ATG5 and beclin 1 blocked clinical grade IL-2-induced autophagy. Moreover, IL-2 induced HMGB1 cytoplasmic translocation in MEFs and promoted interaction between HMGB1 and beclin1, which is required for autophagy induction. Pharmacological and genetic inhibition of autophagy inhibited IL-2-induced cell proliferation and enhanced IL-2-induced apoptosis. These findings suggest that autophagy is an important pro-survival regulator for IL-2-induced cell growth in fibroblasts.
IL-2; Autophagy; Apoptosis; Immunotherapy; HMGB1
Damage-associated molecular pattern (DAMP) molecules are essential for the initiation of innate inflammatory responses to infection and injury. The prototypic DAMP molecule, high-mobility group box 1 (HMGB1), is an abundant architectural chromosomal protein that has location-specific biological functions: within the nucleus as a DNA chaperone, within the cytosol to sustain autophagy and outside the cell as a DAMP molecule. Recent research indicates that aberrant activation of HMGB1 signaling can promote the onset of inflammatory and autoimmune diseases, raising interest in the development of therapeutic strategies to control their function. The importance of HMGB1 activation in various forms of liver disease in relation to liver damage, steatosis, inflammation, fibrosis, tumorigenesis and regeneration is discussed in this review.
Pathogen-associated molecular pattern molecules (PAMPs) are derived from microorganisms and recognized by pattern recognition receptor (PRR)-bearing cells of the innate immune system as well as many epithelial cells. In contrast, damage-associated molecular pattern molecules (DAMPs) are cell-derived and initiate and perpetuate immunity in response to trauma, ischemia, and tissue damage, either in the absence or presence of pathogenic infection. Most PAMPs and DAMPs serve as so-called ‘Signal 0s’ that bind specific receptors [Toll-like receptors, NOD-like receptors, RIG-I-like receptors, AIM2-like receptors, and the receptor for advanced glycation end products (RAGE)] to promote autophagy. Autophagy, a conserved lysosomal degradation pathway, is a cell survival mechanism invoked in response to environmental and cellular stress. Autophagy is inferred to have been present in the last common eukaryotic ancestor and only to have been lost by some obligatory intracellular parasites. As such, autophagy represents a unifying biology, subserving survival and the earliest host defense strategies, predating apoptosis, within eukaryotes. Here, we review recent advances in our understanding of autophagic molecular mechanisms and functions in emergent immunity.
PAMPs; DAMPs; autophagy; apoptosis; danger signals; inflammation; programmed cell death
Pancreatic ductal adenocarcinoma (PDA), the fourth leading cause of cancer death in the United States, is a complex disease that arises in the setting of genetic alterations (KRAS, BRCA1, SMAD4, CDKN2A/p16INK4a and TP53), epigenetic perturbations (MIR155, acetylation and methylation) and epicellular events (diabetes and inflammation). We have demonstrated that the advanced glycation end product-specific receptor (AGER, also called RAGE) contributes to pancreatic tumorigenesis. Targeted ablation of AGER diminishes the amount of autophagic flux and attenuates the development of early pancreatic intraepithelial neoplasia (PanIN) lesions in a murine model of KRAS-drivien carcinogenesis. Autophagy (programmed cell survival), a metabolic process of lysosome-mediated self-digestion, promotes pancreatic cancer growth. In pancreatic tumor cell lines, AGER-mediated autophagy promotes interleukin-6 (IL6)-induced phosphorylation of signal transducer and activator of transcription 3 (pSTAT3) and mitochondrial localization of pSTAT3. Enhanced mitochondrial pSTAT3 increases the pool of available ATP and increases cellular proliferation. Moreover, we observed a positive feedback loop between activation of autophagy and the IL6-pSTAT3 pathway, perhaps different from the role of cytosolic nonphosphorylated STAT3, which has been reported to inhibit autophagy. These AGER-dependent changes were found during the earliest stages of pancreatic cancer development. These observations of inflammation and altered metabolism in PDA provide a pathological link to early precursor lesion development. Thus, AGER is an important inflammatory mediator that modulates crosstalk between prosurvival pathways, IL6-pSTAT3 and autophagy, in PDA tumor cells, and contributes to early PanIN formation.
RAGE; autophagy; oncogene; KRAS; IL6; STAT3
Double-stranded RNA–dependent protein kinase (PKR) is implicated in inflammation and immune dysfunction through its regulation of mitogen-activated protein kinases, interferon regulatory factor 3, nuclear factorκB, apoptosis, and autophagy pathways. A study shows that PKR is also required for the activation of inflammasomes and the subsequent release of high-mobility group box 1 (HMGB1) protein, a proinflammatory cytokine. Thus, the cell stress kinase PKR has multifaceted roles in the regulation of inflammatory immune responses, and PKR and HMGB1 are attractive targets for inflammasome-associated diseases.
Tumorigenesis and the efficacy of cancer therapeutics are both defined by the balance between autophagy and apoptosis. High-mobility group box 1 (HMGB1) is a DNA chaperone and extracellular damage-associated molecular pattern molecule (DAMP) with pro-autophagic activity. TP53/p53 plays a transcription-dependent and -independent role in the regulation of apoptosis, autophagy, metabolism, cell cycle progression, and many other processes. Both HMGB1 and TP53 are tightly linked with the development of cancer, associated with many of the hallmarks defining the altered biology of cancer. We have demonstrated that TP53-HMGB1 complexes regulate the balance between apoptosis and autophagy through regulation of the cytosolic localization of the reciprocal binding partner, whereby increased cytosolic HMGB1 enhances autophagy and increased cytosolic TP53 enhances apoptosis in colon cancer cells. We found that HMGB1-mediated autophagy promotes cell survival in TP53-dependent processes, and that TP53 inhibits autophagy through negative regulation of HMGB1-BECN1 complexes. Nuclear localization of TP53 and HMGB1 in tumors from patients with colon adenocarcinoma had a positive trend with survival time from diagnosis. Thus, HMGB1 and TP53 are critical in the crossregulation of apoptosis and autophagy and central to colon cancer biology.
Apoptosis; autophagy; colorectal cancer; HMGB1; TP53
The balance between apoptosis (“programmed cell death”) and autophagy (“programmed cell survival”) is important in tumor development and response to therapy. Here we show that HMGB1 and p53 form a complex which regulates the balance between tumor cell death and survival. We demonstrate that knockout of p53 inHCT116 cells increases expression of cytosolic HMGB1 and induces autophagy. Conversely, knockout of HMGB1 in mouse embryonic fibroblasts increases p53 cytosolic localization and decreases autophagy. p53 is thus a negative regulator of the HMGB1/Beclin 1 complex, and HMGB1 promotes autophagy in the setting of diminished p53. HMGB1-mediated autophagy promotes tumor cell survival in the setting of p53-dependent processes. The HMGB1/p53 complex affects the cytoplasmic localization of the reciprocal binding partner thereby regulating subsequent levels of autophagy and apoptosis. These insights provide a novel link between HMGB1 and p53 in the crossregulation of apoptosis and autophagy in the setting of cell stress, providing insights into their reciprocal roles in carcinogenesis.
HMGB1; p53; Autophagy; Apoptosis; Colorectal cancer
IFN1@ (interferon, type 1, cluster, also called IFNα) has been extensively studied as a treatment for patients with chronic myeloid leukemia (CML). The mechanism of anticancer activity of IFN1@ is complex and not well understood. Here, we demonstrate that autophagy, a mechanism of cellular homeostasis for the removal of dysfunctional organelles and proteins, regulates IFN1@-mediated cell death. IFN1@ activated the cellular autophagic machinery in immortalized or primary CML cells. Activation of JAK1-STAT1 and RELA signaling were required for IFN1@-induced expression of BECN1, a key regulator of autophagy. Moreover, pharmacological and genetic inhibition of autophagy enhanced IFN1@-induced apoptosis by activation of the CASP8-BID pathway. Taken together, these findings provide evidence for an important mechanism that links autophagy to immunotherapy in leukemia.
IFN1@; autophagy; apoptosis; immunotherapy; chronic myeloid leukemia
The title co-crystal, [Eu(NCS)3(C18H15OP)3][Eu(NCS)2(NO3)(C18H15OP)3], contains two distinct neutral complexes. Each complex has threefold symmetry about its central Eu3+ ion. As a result, the nitrate-containing molecule contains disorder of its bidentate nitrate and two N-bound thiocyanate anions, while the [Eu(NCS)3(OPPh3)3] complex is fully ordered. There is a weak π–π stacking interaction between the phenyl rings of the two molecules [centroid–centroid distance = 4.138 (4) Å].
The title compound, [Tb(NCS)3(C18H15OP)3], contains a six-coordinate TbII cation surrounded by three O-bound triphenylphosphine oxide ligands and three N-bound thiocyanate ligands, each in a fac arrangement. There are two crystallographically unique TbIII atoms in the asymmetric unit. One TbIII atom resides on a threefold rotation axis, while the other has no imposed crystallographic symmetry. The thiocyanate ligands are bound through N atoms, illustrating the hard–hard bonding principles of metal complex chemistry.
In the title compound, C23H38O5, the oxabicyclo[2.2.1]heptane-2,3-dicarboxylic anhydride unit has a normal geometry and the tetradecoxymethyl side chain is fully extended. In the crystal, molecules are linked head-to-head by C—H⋯O hydrogen bonds, forming two-dimensional networks propagating along the a and c-axis directions.
Reactive oxygen species, including hydrogen peroxide (H2O2), can cause toxicity and act as signaling molecules in various pathways regulating both cell survival and cell death. However, the sequence of events between the oxidative insult and cell damage remains unclear. In the current study, we investigated the effect of oxidative stress on activation of the Receptor for Advanced Glycation End-products (RAGE) and subsequent protection against H2O2-induced pancreatic tumor cell damage. We found that exposure of pancreatic tumor cells to H2O2 provoked a nuclear factor kappa B (NF-κB)-dependent increase in RAGE expression. Further, suppression of RAGE expression by RNA interference increased the sensitivity of pancreatic tumor cells to oxidative injury. Furthermore, targeted knockdown of RAGE led to increased cell death by apoptosis and diminished cell survival by autophagy during H2O2-induced oxidative injury. Moreover, we demonstrate that RAGE is a positive feedback regulator for NF-κB as knockdown of RAGE decreased H2O2-induced activity of NF-κB. Taken together, these results suggest that RAGE is an important regulator of oxidative injury. Antioxid. Redox Signal. 15, 2175–2184.
Autophagy, the process by which cells break down spent biochemical and damaged components, plays an important role in cell survival following stress. High mobility group box 1 (HMGB1) regulates autophagy in response to oxidative stress.
Exogenous hydrogen peroxide (H2O2) treatment or knockdown of the major superoxide scavenger enzyme, superoxide dismutase 1 (SOD1), by small interfering RNA (siRNA) increases autophagy in mouse and human cell lines. Addition of either SOD1 siRNA or H2O2 promotes cytosolic HMGB1 expression and extracellular release. Importantly, inhibition of HMGB1 release or loss of HMGB1 decreases the number of autophagolysosomes and autophagic flux under oxidative stress in vivo and in vitro.
HMGB1 release may be a common mediator of response to oxidative stress.
HMGB1 is important for oxidative stress-mediated autophagy and serves as a new target for the treatment of stress-associated disorders. Antioxid. Redox Signal. 15, 2185–2195.
Activation of the induced receptor for advanced glycation endproducts (RAGE) leads to initiation of NF-κB and MAP kinase signaling pathways resulting in propagation and perpetuation of inflammation. RAGE knock out animals are less susceptible to acute inflammation and carcinogen induced tumor development. We have reported that most forms of tumor cell death result in release of the RAGE ligand, HMGB1. We now report a novel role for RAGE in the tumor cell response to stress. Targeted knockdown of RAGE in the tumor cell, leads to increased apoptosis, diminished autophagy and decreased tumor cell survival . In contrast, overexpression of RAGE is associated with enhanced autophagy, diminished apoptosis and greater tumor cell viability. RAGE limits apoptosis through a p53 dependent mitochondrial pathway. Moreover, RAGE-sustained autophagy is associated with decreased phosphorylation of mTOR and increased Beclin-1/VPS34 autophagosome formation. These findings demonstrate that the inflammatory receptor RAGE plays a heretofore unrecognized role in the tumor cell response to stress. Furthermore, these studies establish a direct link between inflammatory mediators in the tumor microenvironment and resistance to programmed cell death. Our data suggest that targeted inhibition of RAGE or its ligands may serve as novel targets to enhance current cancer therapies.
Pancreatic cancer is the fourth most common cancer to cause death due to advanced stage at diagnosis and poor response to current treatment. Autophagy is the lysosome-mediated degradation pathway which plays a critical role in cellular defense, quality control, and energy metabolism. Targeting autophagy is now an exciting field for translational cancer research, as autophagy dysfunction is among the hallmarks of cancer. Pancreatic tumors have elevated autophagy under basal conditions when compared with other epithelial cancers. This review describes our current understanding of the interaction between autophagy and pancreatic cancer development, including risk factors (e.g., pancreatitis, smoking, and alcohol use), tumor microenvironment (e.g., hypoxia and stromal cells), and molecular biology (e.g., K-Ras and p53) of pancreatic cancer. The importance of the HMGB1-RAGE pathway in regulation of autophagy and pancreatic cancer is also presented. Finally, we describe current studies involving autophagy inhibition using either pharmacological inhibitors (e.g., chloroquine) or RNA interference of essential autophagy genes that regulate chemotherapy sensitivity in pancreatic cancer. Summarily, autophagy plays multiple roles in the regulation of pancreatic cancer pathogenesis and treatment, although the exact mechanisms remain unknown.
Autophagy; pancreatic cancer; oncogene; hypoxia; pancreatitis; HMGB1; RAGE; p53; HIF1α; AMPK