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The breakthrough potentials of research papers can be explained by their boundary-spanning qualities. Here, for the first time, we apply the structural variation analysis (SVA) model and its affiliated metrics to investigate the extent to which such qualities characterize a group of Nobel Prize winning papers. We find that these papers share remarkable boundary-spanning traits, marked by exceptional abilities to connect disparate and topically-diverse clusters of research papers. Further, their publications exert structural variations on the scale that significantly alters the betweenness centrality distributions of existing intellectual space. Overall, SVA not only provides a set of leading indicators for describing future Nobel Prize winning papers, but also broadens our understanding of the similar prize-winning properties that may have been overlooked among other regular publications., 27 pages, 8 figures, 9 tables. Submitted to Frontiers in Research Metrics and Analytics

Many people are studying how autophagy intersects with cell death. While most of those studies relate to autophagy acting as a protective mechanism (e.g., to block apoptosis), many papers conclude that autophagy is a death mechanism, and there is a widespread belief that autophagy (in most, but not all cases, we are talking about macroautophagy) can both kill and protect cells depending on the circumstances. Not surprisingly therefore, many of the papers submitted to Autophagy study the relationship between autophagy and cell death.

Anti-cancer chemo-drugs can cause a rapid elevation of intracellular reactive oxygen species (ROS) levels. An imbalance in ROS production and elimination systems leads to cancer cell resistance to chemotherapy. This study aimed to evaluate the mechanism and effect of ROS on multidrug resistance in various human chemoresistant cancer cells by detecting the changes in the amount of ROS, the expression of ROS-related and glycolysis-related genes, and cell death. We found that ROS was decreased while oxidative phosphorylation was increased in chemoresistant cells. We verified that the chemoresistance of cancer cells was achieved in two ways. First, chemoresistant cells preferred oxidative phosphorylation instead of anaerobic glycolysis for energy generation, which increased ATPase activity and produced much more ATP to provide energy. Second, ROS-scavenging systems were enhanced in chemoresistant cancer cells, which in turn decreased ROS amount and thus inhibited chemo-induced cell death. Our in vitro and in vivo photodynamic therapy further demonstrated that elevated ROS production efficiently inhibited chemo-drug resistance and promoted chemoresistant cell death. Taken together, targeting ROS systems has a great potential to treat cancer patients with chemoresistance.

Eco‐friendly synthesis of the silver nanoparticles (AgNPs) has a number of advantages like simplicity, biocompatibility, low toxicity in nature over their physical and chemical methods. In the present study, the authors report biosynthesized AgNPs using the root extract of the perennial plant ‘Spiny gourd’ (Momordica dioica) and investigated their anti‐bacterial application with mechanistic approaches. Different biophysical techniques such as UV‐Vis spectroscopy, FTIR, XRD, TEM, SAED, and DLS were employed for AgNPs characterization. The synthesized AgNPs were polydispersed, crystalline in nature, with anionic surface (−22.3 mV), spherical in shape with an average size of 13.2 nm. In addition, the AgNPs were stable in room temperature and in different biological buffers. The anti‐bacterial activities of AgNPs were studied with respect to the pathogens such as Bacillus subtilis, Staphylococcus aureus (Gram‐positive), Pseudomonas aeruginosa, Escherichia coli, Klebsiella planticola (Gram‐negative), and Candida albicans. Also, mechanistic studies of AgNPs such as protein leakage assay, nucleic acid leakage assay, ATP leakage assay, ROS accumulation, determination of biofilm degrading activity, measurement of potassium, showing that the synthesized AgNPs are capable of containing a potential application in the antimicrobial therapeutic agents and the pharmaceutical industry.

Osteoarthritis (OA) is a cartilage degenerative disease commonly observed in the elderly population and significantly impacts the normal life of OA patients. It has been reported that the development of pathological cell senescence in chondrocytes is involved in the pathogenesis of OA. Celecoxib is a common non-steroidal anti-inflammatory drug, and it has been recently reported to exert therapeutic effects on OA. However, its underlying mechanism is still unclear. The present study intends to explore its mechanism and provide fundamental evidence for the application of Celecoxib in the treatment of clinical OA. Tumor necrosis factor-α (TNF-α) was utilized to establish an in vitro model of chondrocytes senescence. The elevated reactive oxygen species (ROS) generation, increased cell cycle arrest in G0/G1 phase, reduced telomerase activity, and upregulated senescence-associatedβ-galactosidase (SA-β-Gal) staining were all observed in TNF-α-treated chondrocytes, which were then dramatically reversed by 10 and 20 μM Celecoxib. In addition, the upregulated DNA damage biomarkers, p-ATM, and p-CHK2, observed in TNF-α-treated chondrocytes were significantly downregulated by 10 and 20 μM Celecoxib. Lastly, the expression level of p21 and p53 was greatly elevated in chondrocytes by stimulation with TNF-α which was then pronouncedly repressed by treatment with Celecoxib. Taken together, our data reveal that Celecoxib ameliorated TNF-α-induced cellular senescence in human chondrocytes.

Rotigotine is a non-ergoline dopamine agonist that has been licensed for the treatment of Parkinson’s disease. Cardiovascular diseases are the world’s leading cause of death. Ox-LDL- induced endothelial damages are involved in the initiation and progression of cardiovascular diseases. In this study, we assessed the beneficial properties of Rotigotine on ox-LDL-induced insults to HUVECs to highlight its potential use in the treatment of cardiovascular diseases. Our findings show that Rotigotine suppresses the expressions of low-density lipoprotein receptor (LDL-R), proprotein convertase subtilisin/kexin type 9 (PCSK-9), and sterol regulatory element-binding protein (SREBP-2). It also inhibits ox-LDL-induced cholesterol accumulation in endothelial cells (ECs), improves U937 monocytes adhesion, and decreases the representation of NADPH oxidase (NOX-4) and generation of reactive oxygen species (ROS) in endothelial cells (ECs). Furthermore, Rotigotine inhibited the expressions of both vascular cellular adhesion molecule-1 (VCAM-1) and intercellular adhesion molecule-1 (ICAM-1) in HUVECs and had anti-inflammatory efficacy in ox-LDL-induced cells by inhibiting the expressions of pro-inflammatory cytokines. Notably, Rotigotine inhibits the activation of nuclear factor-kappaB (NF-κB) by preventing nuclear translocation of NF-κB p65 and reducing the luciferase activity of NF-κB reporter. We, therefore, conclude that these effects of Rotigotine on HUVECs suggest that it may play a therapeutic role in cardiovascular diseases.

Stroke is a leading cause of morbidity and mortality worldwide. As the most common type of stroke cases, treatment effectiveness is still limited despite intensive research. Recently, traditional Chinese medicine has attracted attention because of potential benefits for stroke treatment. Among these, luteolin, a natural plant flavonoid compound, offers neuroprotection following against ischemic stroke, although the specific mechanisms are unknown. Here we used network pharmacology, molecular docking, and experimental verification to explore the mechanisms whereby luteolin can benefit stroke recovery. The pharmacological and molecular properties of luteolin were obtained from Traditional Chinese Medicine Systems Pharmacology Database and Analysis Platform. The potential targets of luteolin and ischemic stroke were collected from interrogating public databases. Gene Ontology and Kyoto Encyclopedia of Genes and Genomes pathway analyses were performed by Funrich and Database for Annotation, Visualization and Integrated Discovery respectively, a luteolin-target-pathway network constructed using Cytoscape, Autodock vina was used for molecular docking simulation with Discovery Studio was used to visualize and analyze the docked conformations. Lastly, we employed an in vitro model of stroke injury to evaluate the effects of luteolin on cell survival and expression of the putative targets. From 95 candidate luteolin target genes, our analysis identified six core targets . KEGG analysis of the candidate targets identified that luteolin provides therapeutic effects on stroke through TNF signaling and other pathways. Our experimental analyses confirmed the conclusions analyzed above. In summary, the molecular and pharmacological mechanisms of luteolin against stroke are indicated in our study from a systematic perspective.

Docetaxel-associated liver injury has become a serious public health problem, resulting in therapy discontinuation, liver failure, and death. Zafirlukast is a typical leukotriene receptor antagonist used for prophylaxis and chronic treatment of asthma. In this study, we investigate whether treatment with Zafirlukast could alleviate Docetaxel-induced cytotoxicity in hepatocytes. Our results indicate that Zafirlukast mitigated Docetaxel-induced toxicity in LO-2 hepatocytes. Firstly, Zafirlukast reduced the production of 8-hydroxy-2p-deoxyguanosine (8-OHdG) and increased the levels of reduced glutathione (GSH) against Docetaxel. Secondly, Zafirlukast elevated the levels of mitochondrial membrane potential (ΔΨm) and adenosine triphosphate (ATP). Thirdly, Zafirlukast prevented Docetaxel-induced release of lactate dehydrogenase (LDH) and increased cell viability of LO-2 hepatocytes against Docetaxel. We also found that Zafirlukast ameliorated Docetaxel-induced apoptosis by reducing Caspase-3 and Caspase-9 activity. Mechanistically, our results demonstrate that Zafirlukast inhibited the activation of NOD-like receptor protein 3 (NLRP3), mediated by SIRT1. Based on these findings, we conclude that the administration of Zafirlukast might have a protective effect against Docetaxel-induced cytotoxicity in hepatocytes.

Applying toxic chemical to the synthesis of stable gold nanoparticles is one of the limitations of gold nanoparticles for therapeutic applications such as photothermal therapy. Plant compounds such as apigenin (API) with therapeutic potential can be applied in the synthesis of gold nanoparticles. API‐coated gold nanoparticles (Api@AuNPs) with an average size of 19.1 nm and a surface charge of −4.3 mV have been synthesized by a simple and efficient technique. The stability of Api@AuNPs in the biological environment was verified through UV‐Vis spectroscopy. Based on Raman and FTIR spectroscopy analysis, chemical binding of API on the surface of Api@AuNPs through hydroxyl and carbonyl functional groups was found to be the main reason for the stability of the Api@AuNPs in comparison with citrate‐coated gold nanoparticles (Cit@AuNPs). The synthesized Api@AuNPs do not cause major toxic effects up to 128 ppm. Api@AuNP‐mediated photothermal therapy leads to the indiscriminate eradication of almost half of both mouse fibroblastic (L929) and colorectal cancer (CT26) cells. Flow‐cytometry analysis revealed that the cell death mechanism is mainly apoptosis. In the apoptosis triggered cell death in photothermal treatment, Api@AuNPs are preferred over commonly used gold nanoparticles in photothermal treatments which mostly trigger the necrosis cell death pathway.

New strategies to eradicate cancer stem cells in chronic myeloid leukemia (CML) include a combination of imatinib with peroxisome proliferator‐activated receptor gamma (PPARγ) ligands. Recently, we identified the partial PPARγ agonist telmisartan as effective sensitizer of resistant K562 CML cells to imatinib treatment. Here, the importance of the heterocyclic core on the cell death‐modulating effects of the telmisartan‐derived lead 4′‐((2‐propyl‐1H‐benzo[d]imidazol‐1‐yl)methyl)‐[1,1′‐biphenyl]‐2‐carboxylic acid (3 b) was investigated. Inspired by the pharmacodynamics of HYL‐6d and the selective PPARγ ligand VSP‐51, the benzimidazole was replaced by a carbazole or an indole core. The results indicate no correlation between PPARγ activation and sensitization of resistant CML cells to imatinib. The 2‐COOH derivatives of the carbazoles or indoles achieved low activity at PPARγ, while the benzimidazoles showed 60‐100 % activation. Among the 2‐CO2CH3 derivatives, only the ester of the lead (2 b) slightly activated PPARγ. Sensitizing effects were further observed for this non‐cytotoxic 2 b (80 % cell death), and to a lesser extent for the lead 3 b or the 5‐Br‐substituted ester of the benzimidazoles (5 b)., Heterocyclic compounds with carbazole, benzimidazole, or indole core were synthesized as sensitizers to imatinib treatment in resistant chronic myelogenous leukemia cells. The benzimidazole derivatives were clearly more potent as cell death modulators than the respective carbazoles or indoles. These results are important for the further development of effective sensitizers to circumvent tyrosine kinase inhibitor resistance.

Hepatitis B virus (HBV) infection is a worldwide health problem with high morbidity and mortality rates. The therapeutic vaccine is a promising method of treatment, and HBV polymerase plays a vital role in viral replication. Therefore, a therapeutic vaccine that binds to HBV DNA polymerase may control HBV infection. We predicted and selected epitopes of polymerase using online databases and analysis software. We then performed molecular docking and peptide binding assays to evaluate the binding energies and affinities between polymerase epitopes and the HLA-A0201 molecule. Finally, we induced T cells from the peripheral blood mononuclear cells (PBMCs) of healthy donors using each epitope and quantified the functions of epitope-specific T cells by IFN-γELISPOT assay, T2 cell cytotoxicity assay, HepG2.2.15 cell cytotoxicity assay and HBV gene expression assays. Four epitopes (RVTGGVFLV, GLLGFAAPF, LLDDEAGPL and YMDDVVLGA) had low binding energy and two epitopes (RVTGGVFLV and GLLGFAAPF) had a high binding affinity. The T cells stimulated by two epitopes (GLLGFAAPF and HLYSHPIIL) had a greater ability to induce immune response and suppress HBV. The HBV DNA polymerase epitopes identified in this study are promising targets for designing an epitope-based therapeutic vaccine against HBV.

Nonalcoholic fatty liver disease (NAFLD) is the leading cause of chronic liver disease globally. NAFLD is a consequence of fat accumulation in the liver leading to lipotoxicity. Increasing evidence has demonstrated the critical role of autophagy in NAFLD. This study uncovers the unexpected role of immune surveillance protein DDX58/Rig-1 (DExD/H box helicase 58) in activating macroautophagy/autophagy and protecting from lipotoxicity associated with NAFLD. Here we show for the first time that DDX58 protein is significantly reduced in nonalcoholic steatohepatitis (NASH) mouse model, an aggressive form of NAFLD characterized by inflammation and fibrosis of the liver. In addition to decreased expression of DDX58, we found that DDX58 activity can be attenuated by treatments with palmitic acid (PA), a saturated fatty acid. To investigate whether PA inhibition of DDX58 is harmful to the cell, we characterized DDX58 function in hepatocytes when exposed to high doses of PA in the presence and/or absence of DDX58. We show that siRNA knockdown of DDX58 promotes apoptosis. Importantly, we show that stable overexpression of DDX58 is protective against toxic levels of PA and stimulates autophagy. This study begins to demonstrate the regulation of the autophagy receptor protein SQSTM1/p62 through DDX58. DDX58 expression directly influences SQSTM1 mRNA and protein levels. This work proposes a model in which activating DDX58 increases an autophagic response and this aids in clearing toxic lipid inclusion bodies, which leads to inflammation and apoptosis. Activating a DDX58-induced autophagy response may be a strategy for treating NAFLD. Abbreviations:5ʹpppdsRNA: 5ʹ triphosphate double-stranded RNA; CDAHFD: choline-deficient, L-amino acid defined high-fat diet; CEBPB: CCAAT/enhancer binding protein (C/EBP), beta; CQ: chloroquine; DDX58/retinoic acid inducible gene 1/Rig-1: DExD/H box helicase 58; h: hours; IFIH1/MDA5: interferon induced with helicase C domain 1; IFNB/IFN-β: interferon beta 1, fibroblast; KO: knockout; MAVS: mitochondrial antiviral signaling protein; NAFLD: nonalcoholic fatty liver disease; NASH: nonalcoholic steatohepatitis; NFKB/NF-κB: nuclear factor of kappa light polypeptide gene enhancer in B cells; PA: palmitic acid; poly:IC: polyinosinic:polycytidylic acid; PRR: pattern recognition receptors; PSR: picrosirus red; RAP: rapamycin; RLR: RIG-I-like receptor; SQSTM1/p62: sequestosome 1; STING1: stimulator of interferon response cGAMP interactor 1; TBK1: TANK-binding kinase 1.

Pulmonary fibrosis is a common pulmonary interstitial disease of pathogenesis without effective drugs for treatment. Therefore, discovering new and effective drugs is urgently needed. In the present study, we prepared a novel compound named acetyl oxygen benzoate engeletin ester (AOBEE), investigated its effect on experimental pulmonary fibrosis, and proposed a long non-coding RNA (lncRNA)-mediated mechanism of its action. Bleomycin-induced pulmonary fibrosis in mice exhibited that AOBEE improved forced vital capacity (FVC) and alveolar structure and inhibited α-SMA, vimentin, and collagen expression. TGFβ1-stimulated fibroblast L929 cells showed that AOBEE reduced these fibrotic proteins expression and inhibited the activated-fibroblast proliferation and migration. Whole transcriptome sequencing was performed to screen out lncRNA-lnc865 and lnc556 with high expression under bleomycin treatment, but AOBEE caused a considerable decrease in lnc865 and lnc556. Mechanistic study elucidated that AOBEE alleviated pulmonary fibrosis through lnc865- and lnc556-mediated mechanism, in which both lnc865 and lnc556 sponged miR-29b-2-5p to target signal transducer and activator of transcription 3 (STAT3). Further signal pathway inhibitors and the Cignal Finder 45-pathway reporter array illustrated that the up- and downstream pathways were TGFβ1-smad2/3 and p38MAPK, and Krüppel-like factor 4 (KLF4), respectively. In conclusion, AOBEE promoted KLF4 degradation leading to the attenuation of pulmonary fibrosis by inhibiting TGFβ1-smad/p38MAPK-lnc865/lnc556-miR-29b-2-5p-STAT3 signal pathway. We hope this work will provide valuable information to design new drugs and therapeutic targets of lncRNAs for pulmonary fibrosis treatment.

While treatments for colorectal cancer continue to improve, some 50% of patients succumb within 5 years, pointing to the need for additional therapeutic options. We have developed a modified non-replicating adenoviral vector for gene transfer, called AdRGD-PG, which offers improved levels of transduction and transgene expression. Here, we employ the p53-responsive PG promoter to drive expression of p53 or human interferon-β (hIFNβ) in human colorectal cancer cell lines HCT116(wt) (wtp53), HCT116(−/-) (p53 deficient) and HT29 (mutant p53). The HCT116 cell lines were both easily killed with p53 gene transfer, while combined p53 and hIFNβ cooperated for the induction of HT29 cell death and emission of immunogenic cell death (ICD) markers. Elevated annexinV staining and caspase 3/7 activity point to cell death by a mechanism consistent with apoptosis. P53 gene transfer alone or in combination with hIFNβ sensitized all cell lines to chemotherapy, permitting the application of low drug doses while still achieving significant loss of viability. While endogenous p53 status was not sufficient to predict response to treatment, combined p53 and hIFNβ provided an additive effect in HT29 cells. We propose that this approach may prove effective for the treatment of colorectal cancer, permitting the use of limited drug doses.

Rationale: Accumulated evidence indicates that environmental plasticizers are a threat to human and animal fertility. Di (2-ethylhexyl) phthalate (DEHP), a plasticizer to which humans are exposed daily, can trigger reproductive toxicity by acting as an endocrine-disrupting chemical. In mammals, the female primordial follicle pool forms the lifetime available ovarian reserve, which does not undergo regeneration once it is established during the fetal and neonatal period. It is therefore critical to examine the toxicity of DEHP regarding the establishment of the ovarian reserve as it has not been well investigated. Methods: The ovarian cells of postnatal pups, following maternal DEHP exposure, were prepared for single cell-RNA sequencing, and the effects of DEHP on primordial follicle formation were revealed using gene differential expression analysis and single-cell developmental trajectory. In addition, further biochemical experiments, including immunohistochemical staining, apoptosis detection, and Western blotting, were performed to verify the dataset results. Results: Using single-cell RNA sequencing, we revealed the gene expression dynamics of female germ cells and granulosa cells following exposure to DEHP in mice. Regarding germ cells: DEHP impeded the progression of follicle assembly and interfered with their developmental status, while key genes such as Lhx8, Figla, and others, strongly evidenced the reduction. As for granulosa cells: DEHP likely inhibited their proliferative activity, and activated the regulation of cell death. Furthermore, the interaction between ovarian cells mediated by transforming growth factor-beta signaling, was disrupted by DEHP exposure, since the expression of GDF9, BMPR1A, and SMAD3 was affected. In addition, DNA damage and apoptosis were elevated in germ cells and/or somatic cells. Conclusion: These findings offer substantial novel insights into the reproductive toxicity of DEHP exposure during murine germ cell cyst breakdown and primordial follicle formation. These results may enhance the understanding of DEHP exposure on reproductive health.

MOAP1 (modulator of apoptosis 1) is a BAX-binding protein tightly regulated by the ubiquitin-proteasome system. Apoptotic stimuli stabilize MOAP1 protein and facilitate its interaction with BAX to promote apoptosis. Here we show that in contrast to being resistant to apoptotic stimuli, MOAP1-deficient cells are hypersensitive to cell death mediated by starvation rendered by EBSS treatment. MOAP1-deficient cells exhibited impairment in macroautophagy/autophagy signaling induced by EBSS. Mechanistic analysis revealed that MOAP1-deficient cells had no notable defect in the recruitment of the pre-autophagosomal phosphatidylinositol-3-phosphate (PtdIns3P)-binding proteins, ZFYVE1/DFCP1 and WIPI2, nor in the LC3 lipidation mechanism regulated by the ATG12–ATG5-ATG16L1 complex upon EBSS treatment. Interestingly, MOAP1 is required for facilitating efficient closure of phagophore in the EBSS-treated cells. Analysis of LC3-positive membrane structures using Halo-tagged LC3 autophagosome completion assay showed that predominantly unclosed phagophore rather than closed autophagosome was present in the EBSS-treated MOAP1-deficient cells. The autophagy substrate SQSTM1/p62, which is normally contained within the enclosed autophagosome under EBSS condition, was also highly sensitive to degradation by proteinase K in the absence of MOAP1. MOAP1 binds LC3 and the binding is critically dependent on a LC3-interacting region (LIR) motif detected at its N-terminal region. Re-expression of MOAP1, but not its LC3-binding defective mutant, MOAP1-LIR, in the MOAP1-deficient cells, restored EBSS-induced autophagy. Together, these observations suggest that MOAP1 serves a distinct role in facilitating autophagy through interacting with LC3 to promote efficient phagophore closure during starvation. Abbreviations: CQ: Chloroquine; EBSS: Earle’s Balanced Salt Solution; GABARAP: Gamma-Amino Butyric Acid Receptor Associated Protein; IF: Immunofluorescence; IP: Immunoprecipitation; LAMP1: Lysosomal-Associated Membrane Protein 1; LIR: LC3-Interacting Region; MAP1LC3/LC3: Microtubule Associated Protein 1 Light Chain 3; MEF: Mouse Embryonic Fibroblast; MOAP1: Modulator of Apoptosis 1; PE: Phosphatidylethanolamine; PtdIns3K: class III PtdIns3K complex I; PtdIns3P: Phosphatidylinositol-3-phosphate; STX17: Syntaxin 17; ULK1: unc-51 like autophagy activating kinase 1.

Oxygen glucose deprivation (OGD)/re-oxygenation (OGDR) induces profound oxidative injury and neuronal cell death. It mimics ischemia-reperfusion neuronal injury. CPI-1189 is a novel tumor necrosis factor alpha-inhibiting compound with potential neuroprotective function. Here in SH-SY5Y neuronal cells and primary murine cortical neurons, CPI-1189 pretreatment potently inhibited OGDR-induced viability reduction and cell death. In OGDR-stimulated neuronal cells, p38 phosphorylation was blocked by CPI-1189. In addition, CPI-1189 alleviated OGDR-induced reactive oxygen species production, lipid peroxidation, and glutathione consumption. OGDR-induced neuronal cell apoptosis was also inhibited by CPI-1189 pretreatment. Furthermore, in SH-SY5Y cells and cortical neurons, CPI-1189 alleviated OGDR-induced programmed necrosis by inhibiting mitochondrial p53-cyclophilin D-adenine nucleotide translocase 1 association, mitochondrial depolarization, and lactate dehydrogenase release to the medium. In summary, CPI-1189 potently inhibited OGDR-induced oxidative injury and neuronal cell death.

Development of modulators targeting specific interactions of ubiquitin‐based conjugates with their partners is a formidable task since it requires a suitable screening assay and homogeneous ubiquitin conjugates. We developed a novel high‐throughput strategy for screening ligands for Lys48‐linked tetraubiquitin chain in a relatively simple, fast, and affordable manner. This approach combined with a state‐of‐the‐art toolbox of chemical protein synthesis and a specially optimized Cys deprotection protocol enabled us to design highly potent, Lys48‐linked tetraubiquitin chain selective “next generation” dimeric peptide modulators. The dimeric peptide exhibited cancer cell permeability and induced cell death with higher efficiency compared to its monocyclic analogue. These features make our dimeric peptide a promising candidate for further studies using in vivo models. Our assay can be adopted for other various ubiquitin chains in their free or anchored forms as well as conjugates for Ub‐like modifiers., A fluorescence‐based competitive assay was developed, which enabled the high‐throughput screening of various cyclic peptides targeting the Lys48‐linked tetraubiquitin chain. This assay combined with chemical tools enabled the design of a potent dimeric cyclic peptide, which induced cell death with higher efficiency compared to its monocyclic analogue.

This study investigated the neuroprotective effects of triptolide (TPL) in a rat model of cardiopulmonary bypass with deep hypothermia circulatory arrest (DHCA). Rats were randomly divided into six groups: control, sham, DHCA, and DHCA + TPL (100, 200, 300 μg/kg). Neurobehavioral functions were measured using the elevated plus-maze, Y-maze, and Morris water maze tests. Levels of inflammatory cytokines, oxidative stress indices, and brain neurotrophins were measured by ELISA. Microglial activation and cell death was measured by immunofluorescence staining and TUNEL assay, respectively. Finally, activation of the Nrf2 pathway and NF-κB were detected by western blot. The elevated plus-maze, Y-maze, and Morris water maze tests all showed that TPL mitigated anxiety-like behavior, working memory, spatial learning, and memory in DHCA rats. TPL inhibited inflammatory responses and oxidative stress, as well as increased brain neurotrophin levels in DHCA rats. Moreover, TPL attenuated microglia activation and cell death in DHCA rats. Finally, TPL activated the Nrf2 pathway and inhibited NF-κB activity in DHCA rats. These results demonstrated that TPL improved neurobehavioral functions, neuroinflammation, and oxidative stress in DHCA rats, which may be associated with the Nrf2 and NF-κB pathways.

Rationale: Multiple myeloma (MM) is a multifocal malignancy of bone marrow plasma cells, characterized by vicious cycles of remission and relapse that eventually culminate in death. The disease remains mostly incurable largely due to the complex interactions between the bone microenvironment (BME) and MM cells (MMC). In the “vicious cycle” of bone disease, abnormal activation of osteoclasts (OCs) by MMC causes severe osteolysis, promotes immune evasion, and stimulates the growth of MMC. Disrupting these cancer-stroma interactions would enhance treatment response. Methods: To disrupt this cycle, we orthogonally targeted nanomicelles (NM) loaded with non-therapeutic doses of a photosensitizer, titanocene (TC), to VLA-4 (α4ß1, CD49d/CD29) expressing MMC (MM1.S) and αvß3 (CD51/CD61) expressing OC. Concurrently, a non-lethal dose of a radiopharmaceutical, 18F-fluorodeoxyglucose ([18F]FDG) administered systemically interacted with TC (radionuclide stimulated therapy, RaST) to generate cytotoxic reactive oxygen species (ROS). The in vitro and in vivo effects of RaST were characterized in MM1.S cell line, as well as in xenograft and isograft MM animal models. Results: Our data revealed that RaST induced non-enzymatic hydroperoxidation of cellular lipids culminating in mitochondrial dysfunction, DNA fragmentation, and caspase-dependent apoptosis of MMC using VLA-4 avid TC-NMs. RaST upregulated the expression of BAX, Bcl-2, and p53, highlighting the induction of apoptosis via the BAK-independent pathway. The enhancement of multicopper oxidase enzyme F5 expression, which inhibits lipid hydroperoxidation and Fenton reaction, was not sufficient to overcome RaST-induced increase in the accumulation of irreversible function-perturbing α,ß-aldehydes that exerted significant and long-lasting damage to both DNA and proteins. In vivo, either VLA-4-TC-NM or αvß3-TC-NMs RaST induced a significant therapeutic effect on immunocompromised but not immunocompetent MM-bearing mouse models. Combined treatment with both VLA-4-TC-NM and αvß3-TC-NMs synergistically inhibited osteolysis, reduced tumor burden, and prevented rapid relapse in both in vivo models of MM. Conclusions: By targeting MM and bone cells simultaneously, combination RaST suppressed MM disease progression through a multi-prong action on the vicious cycle of bone cancer. Instead of using the standard multidrug approach, our work reveals a unique photophysical treatment paradigm that uses nontoxic doses of a single light-sensitive drug directed orthogonally to cancer and bone cells, followed by radionuclide-stimulated generation of ROS to inhibit tumor progression and minimize osteolysis in both immunocompetent murine and immunocompromised human MM models.

Rationale: Cardiovascular diseases, such as myocardial infarction (MI), are the leading causes of death worldwide. Reperfusion therapy is the common standard treatment for MI. However, myocardial ischemia/reperfusion (I/R) causes cardiomyocyte injury, including apoptosis and fibrosis. We aimed to investigate the effects of conditioned medium from adipose-derived stem cells (ADSC-CM) on apoptosis and fibrosis in I/R-treated hearts and hypoxia/reoxygenation (H/R)-treated cardiomyocytes and the underlying mechanisms. Methods: ADSC-CM was collected from ADSCs. The effects of intramuscular injection of ADSC-CM on cardiac function, cardiac apoptosis, and fibrosis examined by echocardiography, Evans blue/TTC staining, TUNEL assay, and Masson's trichrome staining in I/R-treated mice. We also examined the effects of ADSC-CM on apoptosis and fibrosis in H/R-treated H9c2 cells by annexin V/PI flow cytometry, TUNEL assay, and immunocytochemistry. Results: ADSC-CM treatment significantly reduced heart damage and fibrosis of I/R-treated mice and H/R-treated cardiomyocytes. In addition, the expression of apoptosis-related proteins, such as p53 upregulated modulator of apoptosis (PUMA), p-p53 and B-cell lymphoma 2 (BCL2), as well as the fibrosis-related proteins ETS-1, fibronectin and collagen 3, were significantly reduced by ADSC-CM treatment. Moreover, we demonstrated that ADSC-CM contains a large amount of miR-221/222, which can target and regulate PUMA or ETS-1 protein levels. Furthermore, the knockdown of PUMA and ETS-1 decreased the induction of apoptosis and fibrosis, respectively. MiR-221/222 overexpression achieved similar results. We also observed that cardiac I/R markedly increased apoptosis and fibrosis in miR-221/222 knockout (KO) mice, while ADSC-CM decreased these effects. The increased phosphorylation of p38 and NF‐κB not only mediated myocardial apoptosis through the PUMA/p53/BCL2 pathway but also regulated fibrosis through the ETS-1/fibronectin/collagen 3 pathway. Conclusions: Overall, our results show that ADSC-CM attenuates cardiac apoptosis and fibrosis by reducing PUMA and ETS-1 expression, respectively. The protective effect is mediated via the miR-221/222/p38/NF-κB pathway.

Colorectal cancer (CRC) is one of the most common tumors, ranking second in the global cause of death from cancer. The prognosis of advanced patients is still very poor. In this study, hub modules with the highest association with tumor-infiltrating immune cells were identified by weighted gene co-expression network analysis based on CRC expression data from the Gene Expression Omnibus database. Next, three hub genes (ADAM8, IL-1A, VAV3) related to infiltrating immune cells were identified by co-expression network and prognostic analysis. After analysis and verification of the TIMER database, ADAM8 was selected as a prognostic biomarker. Finally, the result of functional test showed that ADAM8 gene expression down-regulation partially reversed the immune tolerance of CRC cells to TILs. By bioinformatics analysis methods and the experimental techniques, we identified ADAM8 as a prognostic biomarker and clinical therapeutic target related to tumor-infiltrating immune cells in CRC., Graphical abstract

Long non-coding RNAs (lncRNAs) have been proposed as potential targets in OSCC gene therapy. Thus, the study aims to analyze how they exert functions in OSCC. LINC00958, AIM2, Gasdermin D (GSDMD) and tumor protein p53 (TP53) expression levels are analyzed by Quantitative Real-time PCR (qPCR) or Western blotting (WB) in OSCC cells lines. The roles of LINC00958 in cell proliferation, cell death, and GSDMD expression respectively were analyzed by Cell Counting Kit-8 (CCK8) assay, flow cytometry and Immunofluorescence (IF) assay. In addition, expressions of pyroptosis- and autophagy-related proteins are evaluated by WB detection. The targeted binding of LINC00958 and miR-4306 or AIM2 mRNA is predicted by bioinformatics analysis and detected by biodual luciferase system. RIP and qPCR assays analyze whether LINC00958 interacts with SIRT1. We found that LINC00958 showed upregulation in OSCC cells compared to normal oral epithelial cells. LINC00958 silencing significantly suppressed OSCC cell proliferation, induced cell death and reduced autophagy. LINC00958 regulated the levels of miR-4306 which binds to the 3ʹUTR of AIM2, and interacts with and modulates SIRT1 protein expression. LINC00958 regulated GSDMD and AIM2 levels, as well as p53 and SIRT1 levels. SIRT1 overexpression markedly reversed aforementioned effects of LINC00958. LINC00958 not only downregulated miR-4306 levels to activate the pyroptosis pathway mediated by AIM2 and promoted cancer cell survival but also induced a decrease in SIRT protein expression to further reduce p53 levels., Graphical abstract

Rationale: Iron-saturated Lf (Holo-Lactoferrin, Holo-Lf) exhibits a superior anticancer property than low iron-saturated Lf (Apo-Lf). Ferroptosis is an iron-dependent cell death characterized by the accumulation of lipid peroxidation products and lethal reactive oxygen species (ROS). Radiotherapy also exerts its therapeutic effect through ROS. Methods: The effect of different iron-saturated Lf on ferroptosis and radiotherapy were tested on triple-negative breast cancer (TNBC) cell line MDA-MB-231 and non-TNBC cell line MCF-7. Results: Holo-Lf significantly increased the total iron content, promoted ROS generation, increased lipid peroxidation end product, malondialdehyde (MDA), and enhanced ferroptosis of MDA-MB-231 cells. By contrast, Apo-Lf upregulated SLC7a11 expression, increased GSH generation and inhibited ferroptosis of MDA-MB-231 cells. However, non-TNBC MCF-7 cells were resistant to Holo-Lf-induced ferroptosis because MCF-7 cells have a higher redox balance capacity than MDA-MB-231 cells. More importantly, Holo-Lf downregulated HIF-1α expression, ameliorated the hypoxia microenvironment in subcutaneous MDA-MB-231 tumors, and promoted radiation-induced DNA damage to hypoxic MDA-MB-231 cells. Finally, the efficacy of radiotherapy to MDA-MB-231 tumors was enhanced by Holo-Lf. Conclusion: Holo-Lf could induce ferroptosis in MDA-MB-231 cells and sensitize MDA-MB-231 tumors to radiotherapy.

Background: Cancer is a leading cause of death worldwide. Extensive research over decades has led to the development of therapies that inhibit oncogenic signaling pathways. The mammalian target of rapamycin (mTOR) signaling pathway plays an important role in the development of many cancers. Several mTOR inhibitors are approved for the treatment of cancers. However, the anticancer efficacies of mTOR inhibitor monotherapy are still limited. Methods: Western blot was used to detect the expression of indicated molecules. Thioredoxin reductase (TrxR) activity in cells was determined by the endpoint insulin reduction assay. Immunofluorescence staining was used to analyze precise location and expression of target proteins. Nude mice were used for xenograft tumor models. Results: We identified a synergistic lethal interaction of mTOR and TrxR inhibitors and elucidated the underlying molecular mechanisms of this synergism. We demonstrated that mTOR and TrxR inhibitors cooperated to induce cell death by triggering oxidative stress, which led to activation of autophagy, endoplasmic reticulum (ER) stress and c-Jun N-terminal Kinase (JNK) signaling pathway in cancer cells. Remarkably, we found that auranofin (AF) combined with everolimus significantly suppressed tumor growth in HCT116 and SGC-7901 xenograft models with no significant signs of toxicity. Conclusion: Our findings identify a promising therapeutic combination for cancer and has important implications for developing mTOR inhibitor-based combination treatments.

Protein kinases are the family of attractive enzyme targets for drug design with relevance to cancer biology. Serine arginine protein kinase 1 (SRPK1) is responsible for the phosphorylation of serine/arginine (SR)-rich proteins. Alternative Splicing Factor/Splicing Factor 2 (ASF/SF2) involved in mRNA editing. ASF/SF2 is over expressed in many cancers and plays crucial roles in the cell survival. Phosphorylation of ASF/SF2 is decisive for its functions in cancer. In search of potential anticancer therapeutic agents for attenuating phosphorylation of ASF/SF2, we have explored specific and potential inhibitors of SRPK1 from natural and drug like compounds databases using in-silico methods. Compound ZINC02154892 (C02) was found to be the most potent inhibitor for SRPK1. In-vitro molecular and cell biology studies have shown C02 as a potent and specific inhibitor of phosphorylation of ASF/SF2 and cell survival in leukemic cell line. Structural analysis of SRPK1 with compound C02 revealed a unique pattern of binding targeting ATP binding site along with inhibiting recruitment of ASF/SF2 by SRPK1. The possibilities of compound C02 to be used as a lead compound paving way for the development of potent and specific inhibitors of SRPK1 for designing of novel potential anticancer inhibitor is inferred from the current studies.

Highlights • TRAF6 binds to both WT and the mutant form A30 P asyn in SH-SY5Y cell model. • The activation of NF-κB leads to changes in cytokines levels induced by TRAF6 - WT asyn interaction decreasing cell viability. • The interaction between TRAF6 and A30P asyn does not induce NF-κB activation and cytokine regulation in SH-SY5Y cells. • The present work demonstrates a novel role of TRAF6 in the pathophysiology of Parkinson's disease., Background Parkinson's disease (PD) is a neurodegenerative disease characterized by intracellular inclusions named Lewy bodies (LB), and alpha-synuclein (asyn) is the major component of these protein aggregates. The precise physiological and pathological roles of asyn are not fully understood. Nevertheless, asyn present in LB is ubiquitinated but fails to reach the 26S proteasome. The mutation A30 P is related to an aggressive and early-onset form of PD. Tumor necrosis factor receptor-associated factor 6 (TRAF6) is an E3 ubiquitin ligase, and it interacts and ubiquitinates the asyn in atypical chains (lysine K6, K27, K29, and K33). Methods: Here, we investigated the role of TRAF6 interaction with asyn and the involvement of nuclear factor κB (NF-κB), a key transcription factor in pro-inflammatory signaling pathway activation. Results and Conclusion We demonstrated that TRAF6 binds to both WT and the mutant form A30 P asyn in an SH-SY5Y cell model. Additionally, the interaction between TRAF6 and WT asyn induced an increase in the activation of NF-κB, leading to changes in TNF, IL-1β and IL-10 levels and culminating in reduced cell viability. Interestingly, the activation of NF-κB and gene regulation were not found in A30 P asyn. These data point to a novel role of TRAF6 in the pathophysiology of PD.