Although the treatment of brain tumors by targeting kinase-regulated macroautophagy/autophagy, is under investigation, the precise mechanism underlying autophagy initiation and its significance in glioblastoma (GBM) remains to be defined. Here, we report that PAK1 (p21 [RAC1] activated kinase 1) is significantly upregulated and promotes GBM development. The Cancer Genome Atlas analysis suggests that the oncogenic role of PAK1 in GBM is mainly associated with autophagy. Subsequent experiments demonstrate that PAK1 indeed serves as a positive modulator for hypoxia-induced autophagy in GBM. Mechanistically, hypoxia induces ELP3-mediated PAK1 acetylation at K420, which suppresses the dimerization of PAK1 and enhances its activity, thereby leading to subsequent PAK1-mediated ATG5 (autophagy related 5) phosphorylation at the T101 residue. This event not only protects ATG5 from ubiquitination-dependent degradation but also increases the affinity between the ATG12-ATG5 complex and ATG16L1 (autophagy related 16 like 1). Consequently, ELP3-dependent PAK1 (K420) acetylation and PAK1-mediated ATG5 (T101) phosphorylation are required for hypoxia-induced autophagy and brain tumorigenesis by promoting autophagosome formation. Silencing PAK1 with shRNA or small molecule inhibitor FRAX597 potentially blocks autophagy and GBM growth. Furthermore, SIRT1-mediated PAK1-deacetylation at K420 hinders autophagy and GBM growth. Clinically, the levels of PAK1 (K420) acetylation significantly correlate with the expression of ATG5 (T101) phosphorylation in GBM patients. Together, this report uncovers that the acetylation modification and kinase activity of PAK1 plays an instrumental role in hypoxia-induced autophagy initiation and maintaining GBM growth. Therefore, PAK1 and its regulator in the autophagy pathway might represent potential therapeutic targets for GBM treatment.Abbreviations: 3-MA: 3-methyladenine; Ac-CoA: acetyl coenzyme A; ATG5: autophagy related 5; ATG16L1, autophagy related 16 like 1; BafA1: bafilomycin A1; CDC42: cell division cycle 42; CGGA: Chinese Glioma Genome Atlas; CHX, cycloheximide; ELP3: elongator acetyltransferase complex subunit 3; GBM, glioblastoma; HBSS: Hanks balanced salts solution; MAP1LC3B/LC3: microtubule associated protein 1 light chain 3 beta; MAP2K1: mitogen-activated protein kinase kinase 1; MAPK14, mitogen-activated protein kinase 14; PAK1: p21 (RAC1) activated kinase 1; PDK1: pyruvate dehydrogenase kinase 1; PGK1, phosphoglycerate kinase 1; PTMs: post-translational modifications; RAC1: Rac family small GTPase 1; SQSTM1: sequestosome 1; TCGA, The Cancer Genome Atlas.

Beyond classical roles in thrombosis and haemostasis, it becomes increasingly clear that platelets contribute as key players to inflammatory processes. The involvement of platelets in these processes is often mediated through a variety of platelet-derived chemokines which are released upon activation and act as paracrine and autocrine factors. In this study, we investigate CXCL14, a newly described platelet chemokine and its role in thrombus formation as well as monocyte and platelet migration. In addition, we examine the chemokine receptor CXCR4 as a possible receptor for CXCL14 on platelets. Furthermore, with the use of artificially generated platelets derived from induced pluripotent stem cells (iPSC), we investigate the importance of CXCR4 for CXCL14-mediated platelet functions.

Nilotinib is used for treating patients with imatinib-sensitive or -resistant chronic myeloid leukemia (CML); however, nilotinib-resistant cases have been observed in recent years. In addition, a considerable number of patients receiving nilotinib developed diabetes. Metformin is a front-line drug for the treatment of type 2 diabetes, and several studies have shown that diabetes patients treated with metformin have reduced incidence of cancer. This study aimed to define the effect of metformin on CML cells to determine whether metformin overcomes nilotinib resistance, and to identify novel targets for the treatment of nilotinib resistance.

Spreading depolarization (SD) and seizures are pathophysiological events associated with cerebral ischemia. Here, we investigated their role for injury progression in the cerebral cortex. Cerebral ischemia was induced in anesthetized male Wistar rats using the photothrombosis (PT) stroke model. SD and spontaneous neuronal activity were recorded in the presence of either urethane or ketamine/xylazine anesthesia. Blood-brain barrier (BBB) permeability, cerebral perfusion, and cellular damage were assessed through a cranial window and repeated intravenous injection of fluorescein sodium salt and propidium iodide until 4 h after PT. Neuronal injury and early lesion volume were quantified by stereological cell counting and manual and automated assessment of ex vivo T2-weighted magnetic resonance imaging. Onset SDs originated at the thrombotic core and invaded neighboring cortex, whereas delayed SDs often showed opposite propagation patterns. Seizure induction by 4-aminopyridine caused no increase in lesion volume or neuronal injury in urethane-anesthetized animals. Ketamine/xylazine anesthesia was associated with a lower number of onset SDs, reduced lesion volume, and neuronal injury despite a longer duration of seizures. BBB permeability increase inversely correlated with the number of SDs at 3 and 4 h after PT. Our results provide further evidence that ketamine may counteract the early progression of ischemic injury.

Next-generation sequencing (NGS) is used to investigate the presence of somatic mutations. The utility of incorporating routine sequencing to guide diagnosis and therapeutic decisions remains unclear. We report the findings of an observational, multicenter study that aimed to assess the impact of somatic mutation testing by NGS in a reallife setting of chronic myeloid malignancies. A total of 177 patients were enrolled, partitioned into two overlapping groups. In group A (n=94), the indication was to search for clonal hematopoiesis, in a context of suspected myelodysplastic syndrome or myeloproliferative neoplasia. In group B (n=95), the theranostic impact of somatic mutations was studied. A panel of 34 genes was used on DNA extracted from blood or bone marrow samples. Within group A, the detection of clonal hematopoiesis supported the diagnosis of chronic myeloid malignancies for 31 patients while the absence of clonal hematopoiesis ruled out the suspected diagnosis in 47 patients. Within group B, NGS identified prognostically relevant somatic mutations in 32 patients, which had a therapeutic impact in 18 cases. By determining the presence or absence of somatic mutations, the application of NGS in daily practice was found to be useful for an integrated final diagnosis in 83% of the patients. Moreover, the search for somatic mutations had a prognostic impact that led to treatment modification in 19% of the cases. This study outlines the fact that adequate implementation of new investigations may have a significant positive medico-economic impact by enabling appropriate management of patients.

Effectively representing Medical Subject Headings (MeSH) headings (terms) such as disease and drug as discriminative vectors could greatly improve the performance of downstream computational prediction models. However, these terms are often abstract and difficult to quantify. In this paper, we converted the MeSH tree structure into a relationship network and applied several graph embedding algorithms on it to represent these terms. Specifically, the relationship network consisting of nodes (MeSH headings) and edges (relationships), which can be constructed by the tree num. Then, five graph embedding algorithms including DeepWalk, LINE, SDNE, LAP and HOPE were implemented on the relationship network to represent MeSH headings as vectors. In order to evaluate the performance of the proposed methods, we carried out the node classification and relationship prediction tasks. The results show that the MeSH headings characterized by graph embedding algorithms can not only be treated as an independent carrier for representation, but also can be utilized as additional information to enhance the representation ability of vectors. Thus, it can serve as an input and continue to play a significant role in any computational models related to disease, drug, microbe, etc. Besides, our method holds great hope to inspire relevant researchers to study the representation of terms in this network perspective.

Pregnane X receptor (PXR) is activated by chemicals to transcriptionally regulate drug disposition and possibly decrease drug efficacy and increase resistance, suggesting therapeutic value for PXR antagonists. We previously reported the antagonist SPA70 and its analog SJB7, which unexpectedly is an agonist. Here, we describe another unexpected observation: mutating a single residue (W299A) within the PXR ligand-binding domain converts SPA70 to an agonist. After characterizing wild-type and W299A PXR activity profiles, we used molecular dynamics simulations to reveal that in wild-type PXR, agonists stabilize the activation function 2 (AF-2) helix in an "inward" position, but SPA70 displaces the AF-2. In W299A, however, SPA70 stabilizes the AF-2 "inward", like agonists. We validated our model by predicting the antagonist SJC2 to be a W299A agonist, which was confirmed experimentally. Our work correlates previously unobserved ligand-induced conformational changes to PXR cellular activity and, for the first time, reveals how PXR antagonists work.

Toll-like receptor (TLR) signaling pathways need to be tightly controlled to avoid excessive inflammation and unwanted damage to the host. Myeloid differentiation primary response gene 88 (MyD88) is a critical adaptor of TLR signaling. Here, we identified the speckle-type POZ protein (SPOP) as a MyD88-associated protein. SPOP was recruited to MyD88 following TLR4 activation. TLR4 activation also caused the translocation of SPOP from the nucleus to the cytoplasm. SPOP depletion promoted the aggregation of MyD88 and recruitment of the downstream signaling kinases IRAK4, IRAK1 and IRAK2. Consistently, overexpression of SPOP inhibited the TLR4-mediated activation of NF-κB and production of inflammatory cytokines, whereas SPOP depletion had the opposite effects. Furthermore, knockdown of SPOP increased MyD88 aggregation and inflammatory cytokine production upon TLR2, TLR7 and TLR9 activation. Our findings reveal a mechanism by which MyD88 is regulated and highlight a role for SPOP in limiting inflammatory responses.

Induction of broadly neutralizing monoclonal antibodies (bNAbs) that bind to the viral envelope glycoproteins is a major goal of hepatitis C virus (HCV) vaccine research. The study of bNAbs arising in natural infection is essential in this endeavor. We generated a human antibody, 8D6, recognizing the E2 protein of HCV isolated from a chronic hepatitis C patient. This antibody shows broadly neutralizing activity, which covers a pan-genotypic panel of cell culture-derived HCV virions (HCVcc). Functional and epitope analyses demonstrated that 8D6 can block the interaction between E2 and CD81 by targeting a highly conserved epitope on E2. We describe how the 8D6 lineage evolved via somatic hypermutation to achieve broad neutralization. We found that the V(D)J recombination-generated junctional and somatic hypermutation-induced disulfide bridge (C-C) motif in the CDRH3 is critical for the broad neutralization and binding activity of 8D6. This motif is conserved among a series of broadly neutralizing HCV antibodies, indicating a common binding model. Next, the 8D6 inferred germline (iGL) was reconstructed and tested for its binding affinity and neutralization activity. Interestingly, 8D6 iGL-mediated relatively strong inhibition of the 1b genotype PR79L9 strain, suggesting that PR79L9 may serve as a potential natural viral strain that provides E2 sequences that induce bNAbs. Overall, our detailed epitope mapping and genetic studies of the HCV E2-specific mAb 8D6 have allowed for further refinement of antigenic sites on E2 and reveal a new mechanism to generate a functional CDRH3, while its iGL can serve as a probe to identify potential HCV vaccine strains.

The autoimmune encephalitis (AE) syndromes have been characterised by the detection of autoantibodies in serum and/or cerebrospinal fluid which target the extracellular domains of specific neuroglial antigens. The clinical syndromes have phenotypes which are often highly characteristic of their associated antigen-specific autoantibody. For example, the constellation of psychiatric features and the multi-faceted movement disorder observed in patients with NMDAR antibodies are highly distinctive, as are the faciobrachial dystonic seizures observed in close association with LGI1 antibodies. These typically tight correlations may be conferred by the presence of autoantibodies which can directly access and modulate their antigens in vivo. AE remains an under-recognised clinical syndrome but one where early and accurate detection is critical as prompt initiation of immunotherapy is closely associated with improved outcomes. In this review of a rapidly emerging field, we outline molecular observations with translational value. We focus on contemporary methodologies of autoantibody detection, the evolution and distinctive nature of the clinical phenotypes, generalisable therapeutic paradigms, and finally discuss the likely mechanisms of autoimmunity in these patients which may inform future precision therapies.

Autistic individuals experience significantly elevated rates of childhood trauma, self-harm and suicidal behaviour and ideation (SSBI). Is this purely the result of negative environmental experiences, or does this interact with genetic predisposition? In this study we investigated if a genetic predisposition for autism is associated with childhood trauma using polygenic scores (PGS) and genetic correlations in the UK Biobank (105,222 < N < 105,638), and tested potential mediators and moderators of the association between autism, childhood trauma and SSBI. Autism PGS were significantly associated with childhood trauma (max R2 = 0.096%, P < 2 × 10-16), self-harm ideation (max R2 = 0.108%, P < 2 × 10-16), and self-harm (max R2 = 0.13%, P < 2 × 10-16). Supporting this, we identified significant genetic correlations between autism and childhood trauma (rg = 0.36 ± 0.05, P = 8.13 × 10-11), self-harm ideation (rg = 0.49 ± 0.05, P = 4.17 × 10-21) and self-harm (rg = 0.48 ± 0.05, P = 4.58 × 10-21), and an over-transmission of PGS for the two SSBI phenotypes from parents to autistic probands. Male sex negatively moderated the effect of autism PGS on childhood trauma (β = -0.023 ± 0.005, P = 6.74 × 10-5). Further, childhood trauma positively moderated the effect of autism PGS on self-harm score (β = 8.37 × 10-3 ± 2.76 × 10-3, P = 2.42 × 10-3) and self-harm ideation (β = 7.47 × 10-3 ± 2.76 × 10-3, P = 6.71 × 10-3). Finally, depressive symptoms, quality and frequency of social interactions, and educational attainment were significant mediators of the effect of autism PGS on SSBI, with the proportion of effect mediated ranging from 0.23 (95% CI: 0.09-0.32) for depression to 0.008 (95% CI: 0.004-0.01) for educational attainment. Our findings identify that a genetic predisposition for autism is associated with adverse life-time outcomes, which represent complex gene-environment interactions, and prioritizes potential mediators and moderators of this shared biology. It is important to identify sources of trauma for autistic individuals in order to reduce their occurrence and impact.

Bipolar Disorder is costly and debilitating, and many treatments have side effects. Transcranial Direct Current Stimulation (tDCS) is a well-tolerated neuromodulation technique that may be a useful treatment for Bipolar Disorder if targeted to neural regions implicated in the disorder. One potential region is the left ventrolateral prefrontal cortex (vlPFC), which shows abnormally elevated activity during reward expectancy in individuals with Bipolar Disorder. We used a counterbalanced repeated measures design to assess the impact of cathodal (inhibitory) tDCS over the left vlPFC on reward circuitry activity, functional connectivity, and affect in adults with Bipolar Disorder, as a step toward developing novel interventions for individuals with the disorder. -1mA cathodal tDCS was administered over the left vlPFC versus a control region, left somatosensory cortex, concurrently with neuroimaging. Affect was assessed pre and post scan in remitted Bipolar Disorder (n = 27) and age/gender-matched healthy (n = 31) adults. Relative to cathodal tDCS over the left somatosensory cortex, cathodal tDCS over the left vlPFC lowered reward expectancy-related left ventral striatal activity (F(1,51) = 9.61, p = 0.003), and was associated with lower negative affect post scan, controlling for pre-scan negative affect, (F(1,49) = 5.57, p = 0.02) in all participants. Acute cathodal tDCS over the left vlPFC relative to the left somatosensory cortex reduces reward expectancy-related activity and negative affect post tDCS. Build on these findings, future studies can determine whether chronic cathodal tDCS over the left vlPFC has sustained effects on mood in individuals with Bipolar Disorder, to guide new treatment developments for the disorder.

Attention-deficit hyperactivity disorder (ADHD) is associated with pervasive impairments in attention and cognitive control. Although brain circuits underlying these impairments have been extensively investigated with resting-state fMRI, little is known about task-evoked functional brain circuits and their relation to cognitive control deficits and inattention symptoms in children with ADHD. Children with ADHD and age, gender and head motion matched typically developing (TD) children completed a Go/NoGo fMRI task. We used multivariate and dimensional analyses to investigate impairments in two core cognitive control systems: (i) cingulo-opercular "salience" network (SN) anchored in the right anterior insula, dorsal anterior cingulate cortex (rdACC), and ventrolateral prefrontal cortex (rVLPFC) and (ii) dorsal frontoparietal "central executive" (FPN) network anchored in right dorsolateral prefrontal cortex (rDLPFC) and posterior parietal cortex (rPPC). We found that multivariate patterns of task-evoked effective connectivity between brain regions in SN and FPN distinguished the ADHD and TD groups, with rDLPFC-rPPC connectivity emerging as the most distinguishing link. Task-evoked rdACC-rVLPFC connectivity was positively correlated with NoGo accuracy, and negatively correlated with severity of inattention symptoms. Brain-behavior relationships were robust against potential age, gender, and head motion confounds. Our findings highlight aberrancies in task-evoked modulation of SN and FPN connectivity in children with ADHD. Crucially, cingulo-frontal connectivity was a common locus of deficits in cognitive control and clinical measures of inattention symptoms. Our study provides insights into a parsimonious systems neuroscience model of cognitive control deficits in ADHD, and suggests specific circuit biomarkers for predicting treatment outcomes in childhood ADHD.

Older patients with severe physical trauma are at high risk of developing neuropsychiatric syndromes with global impairment of cognition, attention, and consciousness. We employed a thoracic trauma (TxT) mouse model and thoroughly analyzed age-dependent spatial and temporal posttraumatic alterations in the central nervous system. Up to 5 days after trauma, we observed a transient 50% decrease in the number of excitatory synapses specifically in hippocampal pyramidal neurons accompanied by alterations in attention and motor activity and disruption of contextual memory consolidation. In parallel, hippocampal corticotropin-releasing hormone (CRH) expression was highly upregulated, and brain-derived neurotrophic factor (BDNF) levels were significantly reduced. In vitro experiments revealed that CRH application induced neuronal autophagy with rapid lysosomal degradation of BDNF via the NF-κB pathway. The subsequent synaptic loss was rescued by BDNF as well as by specific NF-κB and CRH receptor 1 (CRHR1) antagonists. In vivo, the chronic application of a CRHR1 antagonist after TxT resulted in reversal of the observed histological, molecular, and behavioral alterations. The data suggest that neuropsychiatric syndromes (i.e., delirium) after peripheral trauma might be at least in part due to the activation of the hippocampal CRH/NF-κB/BDNF pathway, which results in a dramatic loss of synaptic contacts. The successful rescue by stress hormone receptor antagonists should encourage clinical trials focusing on trauma-induced delirium and/or other posttraumatic syndromes.

Memory CD8 T cells can provide long-term protection against tumors, which depends on their enhanced proliferative capacity, self-renewal and unique metabolic rewiring to sustain cellular fitness. Specifically, memory CD8 T cells engage oxidative phosphorylation and fatty acid oxidation to fulfill their metabolic demands. In contrast, tumor-infiltrating lymphocytes (TILs) display severe metabolic defects, which may underlie their functional decline. Here, we show that overexpression of proliferator-activated receptor gamma coactivator 1-alpha (PGC-1α), the master regulator of mitochondrial biogenesis (MB), favors CD8 T cell central memory formation rather than resident memory generation. PGC-1α-overexpressing CD8 T cells persist and mediate more robust recall responses to bacterial infection or peptide vaccination. Importantly, CD8 T cells with enhanced PGC-1α expression provide stronger antitumor immunity in a mouse melanoma model. Moreover, TILs overexpressing PGC-1α maintain higher mitochondrial activity and improved expansion when rechallenged in a tumor-free host. Altogether, our findings indicate that enforcing mitochondrial biogenesis promotes CD8 T cell memory formation, metabolic fitness, and antitumor immunity in vivo.

Pancreatic beta-cells are central regulators of glucose homeostasis. By tightly coupling nutrient sensing and granule exocytosis, beta-cells adjust the secretion of insulin to the circulating blood glucose levels. Failure of beta-cells to augment insulin secretion in insulin-resistant individuals leads progressively to impaired glucose tolerance, Type 2 diabetes, and diabetes-related diseases. Mitochondria play a crucial role in β-cells during nutrient stimulation, linking the metabolism of glucose and other secretagogues to the generation of signals that promote insulin secretion. Mitochondria are double-membrane organelles containing numerous channels allowing the transport of ions across both membranes. These channels regulate mitochondrial energy production, signalling, and cell death. The mitochondria of β-cells express ion channels whose physio/pathological role is underappreciated. Here, we describe the mitochondrial ion channels identified in pancreatic β-cells, we further discuss the possibility of targeting specific β-cell mitochondrial channels for the treatment of Type 2 diabetes, and we finally highlight the evidence from clinical studies. LINKED ARTICLES: This article is part of a themed issue on Cellular metabolism and diseases. To view the other articles in this section visit http://onlinelibrary.wiley.com/doi/10.1111/bph.v178.10/issuetoc.

Hydrazinocurcumin (HZC), a synthetic derivative of curcumin (CUR), has been documented to show anticancer potential in impeding tumor growth in several cancers, including hepatocellular carcinoma (HCC). However, the underlying molecular mechanisms remain unclear. This study aimed to explore the function and underlying mechanisms of HZC on HCC cells, which may involve the p38 mitogen activated protein kinase (MAPK) pathway. HZC was first purified and identified. HepG2 cells were then subjected to treatment with HZC or CUR of different concentrations and p38 MAPK signaling inhibitor (SB203580) to verify their effects on HCC cell apoptosis and proliferation. Furthermore, the functional relevance between HZC and the p38 MAPK pathway in HCC was examined. It was observed that 40 μM HZC exhibited the best pro-apoptosis effect in HCC cells. HZC was found to inhibit HCC cell proliferation and promote apoptosis, the effect of which was stronger than 5-fluorouracil (5-FU). More importantly, the anti-oncogenic effect of HZC and 5-FU was implicated with activation of the p38 MAPK pathway. In vivo experimental results showed that HZC inhibited tumor growth more effectively than 5-FU through the p38 MAPK pathway. These results provide evidence that HZC exerted anti-oncogenic and pro-apoptosis effects in HCC cells through activation of the p38 MAPK pathway.

Efficacy of the live-attenuated herpes zoster (HZ) vaccine (ZVL) wanes substantially over time. We evaluated immunogenicity and safety of the adjuvanted recombinant zoster vaccine (RZV) in previous ZVL recipients.

Cell microencapsulation is a rapidly expanding field with broad potential for stem cell therapies and tissue engineering research. Traditional alginate microspheres suffer from poor biocompatibility, and microencapsulation of more advanced hydrogels is challenging due to their slower gelation rates. We have developed a novel, noncytotoxic, nonemulsion-based method to produce hydrogel microspheres compatible with a wide variety of materials, called core-shell spherification (CSS). Fabrication of microspheres by CSS derived from two slow-hardening hydrogels, hyaluronic acid (HA) and polyethylene glycol diacrylate (PEGDA), was characterized. HA microspheres were manufactured with two different crosslinking methods: thiolation and methacrylation. Microspheres of methacrylated HA (MeHA) had the greatest swelling ratio, the largest average diameter, and the lowest diffusion barrier. In contrast, PEGDA microspheres had the smallest diameters, the lowest swelling ratio, and the highest diffusion barrier, while microspheres of thiolated HA had characteristics that were in between the other two groups. To test the ability of the hydrogels to protect cells, while promoting function, diabetic NOD mice received intraperitoneal injections of PEGDA or MeHA microencapsulated canine islets. PEGDA microspheres reversed diabetes for the length of the study (up to 16 weeks). In contrast, islets encapsulated in MeHA microspheres at the same dose restored normoglycemia, but only transiently (3-4 weeks). Nonencapsulated canine islet transplanted at the same dose did not restore normoglycemia for any length of time. In conclusion, CSS provides a nontoxic microencapsulation procedure compatible with various hydrogel types.

Lipid A is one of the three components of bacterial lipopolysaccharides constituting the outer membrane of Gram-negative bacteria, and is recognized to have an important biological role in the inflammatory response of mammalians. Its biological activity is modulated by the number of acyl-chains that are present in the lipid and by the dielectric medium, i.e., the type of counter-ions, through electrostatic interactions. In this paper, we report on a coarse-grained model of chemical variants of Lipid A based on the hybrid particle-field/molecular dynamics approach (hPF-MD). In particular, we investigate the stability of Lipid A bilayers for two different hexa- and tetra-acylated structures. Comparing particle density profiles along bilayer cross-sections, we find good agreement between the hPF-MD model and reference all-atom simulation for both chemical variants of Lipid A. hPF-MD models of constituted bilayers composed by hexa-acylated Lipid A in water are stable within the simulation time. We further validate our model by verifying that the phase behavior of Lipid A/counterion/water mixtures is correctly reproduced. In particular, hPF-MD simulations predict the correct self-assembly of different lamellar and micellar phases from an initially random distribution of Lipid A molecules with counterions in water. Finally, it is possible to observe the spontaneous formation and stability of Lipid A vesicles by fusion of micellar aggregates.