Macrophage polarization in the tumor microenvironment: Emerging roles and therapeutic potentials.

The tumor microenvironment (TME) is a combination of tumor cells and indigenous host stroma, which consists of tumor-infiltrating immune cells, endothelial cells, fibroblasts, pericytes, and non-cellular elements. Tumor-associated macrophages (TAMs) represent the major tumor-infiltrating immune cell type and are generally polarized into two functionally contradictory subtypes, namely classical activated M1 macrophages and alternatively activated M2 macrophages. Macrophage polarization refers to how macrophages are activated at a given time and space. The interplay between the TME and macrophage polarization can influence tumor initiation and progression, making TAM a potential target for cancer therapy. Here, we review the latest investigations on factors orchestrating macrophage polarization in the TME, how macrophage polarization affects tumor progression, and the perspectives in modulating macrophage polarization for cancer immunotherapy.

A method to study human synaptic protein-protein interactions by using flow cytometry coupled to proximity ligation assay (Syn-FlowPLA).

BACKGROUND: Synapses are highly specialized sites characterized by intricate networks of protein-protein interactions (PPIs) important to maintain healthy synapses. Therefore, mapping these networks could address unsolved questions about human cognition, synaptic plasticity, learning, and memory in physiological and pathological conditions. The limitation of analyzing synaptic interactions in living humans has led to the development of methods to isolate synaptic terminals (synaptosomes) from cryopreserved human brains. NEW METHOD: Here, we established a method to detect synaptic PPIs by applying flow cytometric proximity ligation assay (FlowPLA) to synaptosomes isolated from frozen human frontal cortex (FC) and hippocampus (HP) (Syn-FlowPLA). RESULTS: Applying this method in synaptosomes, we were able to detect the known post-synaptic interactions between distinct subtypes of N-methyl-D-aspartate glutamate receptors (NMDARs) and their anchoring postsynaptic density 95 protein (PSD95). Moreover, we detected the known pre-synaptic interactions between the SNARE complex proteins synaptosomal-associated protein of 25 kDa (SNAP25), synaptobrevin (VAMP2), and syntaxin 1a (STX1A). As a negative control, we analyzed the interaction between mitochondrial superoxide dismutase 2 (SOD2) and PSD95, which are not expected to be physically associated. COMPARISON WITH EXISTING METHODS: PPIs have been studied in vitro primarily by co-immunoprecipitation, affinity chromatography, protein-fragment complementation assays (PCAs), and flow cytometry. All these are valid approaches; however, they require more steps or combination with other techniques. PLA technology identifies PPIs with high specificity and sensitivity. CONCLUSIONS: The Syn-FlowPLA described here allows rapid analyses of PPIs, specifically within the synaptic compartment isolated from frozen autopsy specimens, achieving greater target sensitivity. Syn-FlowPLA, as presented here, is therefore a useful method to study human synaptic PPI in physiological and pathological conditions.

High-throughput screening of SARS-CoV-2 main and papain-like protease inhibitors.

The global COVID-19 coronavirus pandemic has infected over 109 million people, leading to over 2 million deaths up to date and still lacking of effective drugs for patient treatment. Here, we screened about 1.8 million small molecules against the main protease (Mpro) and papain like protease (PLpro), two major proteases in severe acute respiratory syndrome-coronavirus 2 genome, and identified 1851Mpro inhibitors and 205 PLpro inhibitors with low nmol/l activity of the best hits. Among these inhibitors, eight small molecules showed dual inhibition effects on both Mpro and PLpro, exhibiting potential as better candidates for COVID-19 treatment. The best inhibitors of each protease were tested in antiviral assay, with over 40% of Mpro inhibitors and over 20% of PLpro inhibitors showing high potency in viral inhibition with low cytotoxicity. The X-ray crystal structure of SARS-CoV-2 Mpro in complex with its potent inhibitor 4a was determined at 1.8 Å resolution. Together with docking assays, our results provide a comprehensive resource for future research on anti-SARS-CoV-2 drug development.

Research progress on chemical components and pharmacological action of Solanum lyratum Thunb.

OBJECTIVES: Solanum lyratum Thunb (SLT) is a perennial plant of the Solanaceae family, and is extensively used in the clinical practice of traditional Chinese medicine. Malaria, oedema, gonorrhoea, cancer, wind and fever, jaundiced hepatitis, cholecystitis and rheumatoid arthritis are among the diseases that it is used to treat. To offer a foundation for further development and usage of SLT, the pieces of literature about the chemical composition and pharmacological action of SLT were reviewed and analysed. KEY FINDINGS: The chemical constituents of SLT mainly included steroids, alkaloids, flavonoids, terpenoids, anthraquinones, phenylpropanoids and others. Pharmacological action mainly contains anti-tumour, antibacterial, anti-inflammatory, anti-oxidation and other pharmacological actions, among them, the anti-tumour effect is particularly outstanding. SUMMARY: At present, studies on the pharmacological effects of SLT mainly focus on alkaloids and steroidal saponins. In the follow-up studies, studies on the pharmacological activities of other chemical components in SLT, such as flavonoids and terpenoids, should be strengthened. It has the potential to pave the way for more research and development of novel SLT medicines.

Ligand recognition and activation of neuromedin U receptor 2.

Neuromedin U receptor 2 (NMU2), an emerging attractive target for treating obesity, has shown the capability in reducing food intake and regulating energy metabolism when activated. However, drug development of NMU2 was deferred partially due to the lack of structural information. Here, we present the cryo-electron microscopy (cryo-EM) structure of NMU2 bound to the endogenous agonist NmU-25 and Gi1 at 3.3 Å resolution. Combined with functional and computational data, the structure reveals the key factors that govern the recognition and selectivity of peptide agonist as well as non-peptide antagonist, providing the structural basis for design of novel and highly selective drugs targeting NMU2. In addition, a 25-degree rotation of Gi protein in reference to NMU2 is also observed compared in other structures of class A GPCR-Gi complexes, suggesting heterogeneity in the processes of G protein-coupled receptors (GPCRs) activation and G protein coupling.

The Genus Broussonetia: An Updated Review of Phytochemistry, Pharmacology and Applications.

The Broussonetia genus (Moraceae), recognized for its value in many Chinese traditional herbs, mainly includes Broussonetia papyrifera (L.) L'Hér. ex Vent. (BP), Broussonetia kazinoki Siebold (BK), and Broussonetia luzonica (Blanco) Bureau (BL). Hitherto, researchers have found 338 compounds isolated from BP, BK, and BL, which included flavonoids, polyphenols, phenylpropanoids, alkaloids, terpenoids, steroids, and others. Moreover, its active compounds and extracts have exhibited a variety of pharmacological effects such as antitumor, antioxidant, anti-inflammatory, antidiabetic, anti-obesity, antibacterial, and antiviral properties, and its use against skin wrinkles. In this review, the phytochemistry and pharmacology of Broussonetia are updated systematically, after its applications are first summarized. In addition, this review also discusses the limitations of investigations and the potential direction of Broussonetia. This review can help to further understand the phytochemistry, pharmacology, and other applications of Broussonetia, which paves the way for future research.

Tapinanthus species: A review of botany and biology, secondary metabolites, ethnomedical uses, current pharmacology and toxicology.

ETHNOPHARMACOLOGICAL RELEVANCE: Tapinanthus species are hemiparasites that grow on diverse hosts in African regions. Tapinanthus species are locally known as "all purpose herbs" as they are traditionally used to treat various diseases such as diabetes, hypertension, cancer, inflammation, malaria, anemia, anxiety, itching, and so on. AIM OF THE STUDY: A comprehensive review on research outcomes and future perspectives of Tapinanthus species are presented to provide a reference for relevant researchers. MATERIALS AND METHODS: The references regarding Tapinanthus species were retrieved from Google Scholar, Web of Science, Sci-finder, PubMed, Elsevier, Wiley, China National Knowledge Infrastructure, Open Access Library, and SpringerLink between 1963 and 2022. Scientific plant names were provided by "The Plant List" (www.theplantlist.org) and "The world Flora Online" (www.worldfloraonline.org). RESULTS: Even though Tapinanthus species are regarded as notorious pests that can undermine various hosts, they are, as omnipotent herbs in folklore, meaningful for the development of potential phytomedicine sources. Phytochemistry screening has revealed the presence of glycosides, triterpenoids, flavonoids, alkaloids, tannins, steroids, anthraquinones. Among them, the chemical structures of 40 compounds have been elucidated by phytochemical methods without alkaloids and anthraquinones. These secondary metabolites might be responsible for ethnomedical uses and bioactivities of Tapinanthus species. Current research has provided scientific evidence for traditional uses of Tapinanthus species, especially unraveling hypoglycemic, hepatoprotective, antioxidant, antibacterial, anti-anxiety, anti-depression, anti-inflammatory, and other pharmacological properties. Given the fact that ethnomedical uses served as a valuable reference for pharmacology, however, some records to treat arthritis, fever, itching, dysentery, stomach pain, and anemia, have not been confirmed in current research. Furthermore, the toxic effects of Tapinanthus species were susceptible to the dosages, with relative safety across a wide range. CONCLUSIONS: To reasonably yield Tapinanthus species, artificial culture might be a promising method to develop in the future. The discrepancies between phytochemistry screening and structure elucidation, as well as between ethnomedical uses and current pharmacology, need to be further clarified. The identification of bioactive compounds in crude extracts and fractions, the illustration of the underlying mechanisms of pharmacology, along with the addition of cytotoxicity, genotoxicity, and clinical trials of toxic tests, should be carried out in depth. This review highlights that Tapinanthus species can be considered promising phytomedicine sources as long as we adhere to digging more deeply into their potential role.

Structural insights into ligand recognition and selectivity of somatostatin receptors.

Somatostatin receptors (SSTRs) play versatile roles in inhibiting the secretion of multiple hormones such as growth hormone and thyroid-stimulating hormone, and thus are considered as targets for treating multiple tumors. Despite great progress made in therapeutic development against this diverse receptor family, drugs that target SSTRs still show limited efficacy with preferential binding affinity and conspicuous side-effects. Here, we report five structures of SSTR2 and SSTR4 in different states, including two crystal structures of SSTR2 in complex with a selective peptide antagonist and a non-peptide agonist, respectively, a cryo-electron microscopy (cryo-EM) structure of Gi1-bound SSTR2 in the presence of the endogenous ligand SST-14, as well as two cryo-EM structures of Gi1-bound SSTR4 in complex with SST-14 and a small-molecule agonist J-2156, respectively. By comparison of the SSTR structures in different states, molecular mechanisms of agonism and antagonism were illustrated. Together with computational and functional analyses, the key determinants responsible for ligand recognition and selectivity of different SSTR subtypes and multiform binding modes of peptide and non-peptide ligands were identified. Insights gained in this study will help uncover ligand selectivity of various SSTRs and accelerate the development of new molecules with better efficacy by targeting SSTRs.

Tyrosine Kinase Inhibitors Reduce NMDA NR1 Subunit Expression, Nuclear Translocation, and Behavioral Pain Measures in Experimental Arthritis.

In the lumbar spinal cord dorsal horn, release of afferent nerve glutamate activates the neurons that relay information about injury pain. Here, we examined the effects of protein tyrosine kinase (PTK) inhibition on NMDA receptor NR1 subunit protein expression and subcellular localization in an acute experimental arthritis model. PTK inhibitors genistein and lavendustin A reduced cellular histological translocation of NMDA NR1 in the spinal cord occurring after the inflammatory insult and the nociceptive behavioral responses to heat. The PTK inhibitors were administered into lumbar spinal cord by microdialysis, and secondary heat hyperalgesia was determined using the Hargreaves test. NMDA NR1 cellular protein expression and nuclear translocation were determined by immunocytochemical localization with light and electron microscopy, as well as with Western blot analysis utilizing both C- and N-terminal antibodies. Genistein and lavendustin A (but not inactive lavendustin B or diadzein) effectively reduced (i) pain related behavior, (ii) NMDA NR1 subunit expression increases in spinal cord, and (iii) the shift of NR1 from a cell membrane to a nuclear localization. Genistein pre-treatment reduced these events that occur in vivo within 4 h after inflammatory insult to the knee joint with kaolin and carrageenan (k/c). Cycloheximide reduced glutamate activated upregulation of NR1 content confirming synthesis of new protein in response to the inflammatory insult. In addition to this in vivo data, genistein or staurosporin inhibited upregulation of NMDA NR1 protein and nuclear translocation in vitro after treatment of human neuroblastoma clonal cell cultures (SH-SY5Y) with glutamate or NMDA (4 h). These studies provide evidence that inflammatory activation of peripheral nerves initiates increase in NMDA NR1 in the spinal cord coincident with development of pain related behaviors through glutamate non-receptor, PTK dependent cascades.

Hippocampal stem cells promotes synaptic resistance to the dysfunctional impact of amyloid beta oligomers via secreted exosomes.

BACKGROUND: Adult hippocampal neurogenesis plays an important role in synaptic plasticity and cogntive function. We reported that higher numbers of neural stem cells (NSC) in the hippocampus of cognitively-intact individuals with high Alzheimer's disease (AD) pathology (plaques and tangles) is associated with decreased synaptic amyloid beta oligomers (Aßο), an event linked to onset of dementia in AD. While these findings suggest a link between NSC and synaptic resistance to Aßο, the involved mechanism remains to be determined. With this goal in mind, here we investigated the ability of exosomes secreted from hippocampal NSC to promote synaptic resilience to Aßo. METHODS: Exosomes isolated from media of hippocampus NSC (NSC-exo) or mature hippocampal neuronal (MN-exo) cultures were delivered intracerebroventricularly (ICV) to mice before assessment of Aßο-induced suppression of hippocampal long-term potentiation (LTP) and memory deficits. Aßο binding to synapses was assessed in cultured hippocampal neurons and on synaptosomes isolated from hippocampal slices from wild type mice and from an inducible mouse model of NSC ablation (Nestin-δ-HSV-TK mice) treated with exosomes. Expression of CaMKII and of AMPA and NMDA glutamate receptor subunits in synaptosomes was measured by western blot. Small RNA Deep sequencing was performed to identify microRNAs enriched in NSC-exo as compared to MN-exo. Mimics of select miRNAs were injected ICV. RESULTS: NSC-exo, but not MN-exo, abolished Aßo-induced suppression of LTP and subsequent memory deficits. Furthermore, in hippocampal slices and cultured neurons, NSC-exo significantly decreased Aßo binding to the synapse. Similarly, transgenic ablation of endogenous NSC increased synaptic Aßo binding, which was reversed by exogenous NSC-exo. Phosphorylation of synaptic CaMKII was increased by NSC-exo, while AMPA and NMDA receptors were not affected. Lastly, we identified a set of miRNAs enriched in NSC-exo that, when injected ICV, protected the synapses from Aßo-binding and Aßo-induced LTP inhibition. CONCLUSIONS: These results identify a novel mechanism linking NSC-exo and synaptic susceptibility to Aßo that may underscore cognitive resilience of certain individuals with increased neurogenesis in spite of AD neuropathology and unmask a novel target for the development of a new treatment concept for AD centered on promoting synaptic resilience to toxic amyloid proteins.

Two disparate ligand-binding sites in the human P2Y1 receptor.

In response to adenosine 5'-diphosphate, the P2Y1 receptor (P2Y1R) facilitates platelet aggregation, and thus serves as an important antithrombotic drug target. Here we report the crystal structures of the human P2Y1R in complex with a nucleotide antagonist MRS2500 at 2.7 Å resolution, and with a non-nucleotide antagonist BPTU at 2.2 Å resolution. The structures reveal two distinct ligand-binding sites, providing atomic details of P2Y1R's unique ligand-binding modes. MRS2500 recognizes a binding site within the seven transmembrane bundle of P2Y1R, which is different in shape and location from the nucleotide binding site in the previously determined structure of P2Y12R, representative of another P2YR subfamily. BPTU binds to an allosteric pocket on the external receptor interface with the lipid bilayer, making it the first structurally characterized selective G-protein-coupled receptor (GPCR) ligand located entirely outside of the helical bundle. These high-resolution insights into P2Y1R should enable discovery of new orthosteric and allosteric antithrombotic drugs with reduced adverse effects.

Peripheral adipose tissue insulin resistance alters lipid composition and function of hippocampal synapses.

Compelling evidence indicates that type 2 diabetes mellitus, insulin resistance (IR), and metabolic syndrome are often accompanied by cognitive impairment. However, the mechanistic link between these metabolic abnormalities and CNS dysfunction requires further investigations. Here, we evaluated whether adipose tissue IR and related metabolic alterations resulted in CNS changes by studying synapse lipid composition and function in the adipocyte-specific ecto-nucleotide pyrophosphate phosphodiesterase over-expressing transgenic (AtENPP1-Tg) mouse, a model characterized by white adipocyte IR, systemic IR, and ectopic fat deposition. When fed a high-fat diet, AtENPP1-Tg mice recapitulate essential features of the human metabolic syndrome, making them an ideal model to characterize peripherally induced CNS deficits. Using a combination of gas chromatography and western blot analysis, we found evidence of altered lipid composition, including decreased phospholipids and increased triglycerides (TG) and free fatty acid in hippocampal synaptosomes isolated from high-fat diet-fed AtENPP1-Tg mice. These changes were associated with impaired basal synaptic transmission at the Schaffer collaterals to hippocampal cornu ammonis 1 (CA1) synapses, decreased phosphorylation of the GluN1 glutamate receptor subunit, down-regulation of insulin receptor expression, and up-regulation of the free fatty acid receptor 1.

Agonist-bound structure of the human P2Y12 receptor.

The P2Y12 receptor (P2Y12R), one of eight members of the P2YR family expressed in humans, is one of the most prominent clinical drug targets for inhibition of platelet aggregation. Although mutagenesis and modelling studies of the P2Y12R provided useful insights into ligand binding, the agonist and antagonist recognition and function at the P2Y12R remain poorly understood at the molecular level. Here we report the structures of the human P2Y12R in complex with the full agonist 2-methylthio-adenosine-5'-diphosphate (2MeSADP, a close analogue of endogenous agonist ADP) at 2.5 Šresolution, and the corresponding ATP derivative 2-methylthio-adenosine-5'-triphosphate (2MeSATP) at 3.1 Šresolution. These structures, together with the structure of the P2Y12R with antagonist ethyl 6-(4-((benzylsulfonyl)carbamoyl)piperidin-1-yl)-5-cyano-2-methylnicotinate (AZD1283), reveal striking conformational changes between nucleotide and non-nucleotide ligand complexes in the extracellular regions. Further analysis of these changes provides insight into a distinct ligand binding landscape in the δ-group of class A G-protein-coupled receptors (GPCRs). Agonist and non-nucleotide antagonist adopt different orientations in the P2Y12R, with only partially overlapped binding pockets. The agonist-bound P2Y12R structure answers long-standing questions surrounding P2Y12R-agonist recognition, and reveals interactions with several residues that had not been reported to be involved in agonist binding. As a first example, to our knowledge, of a GPCR in which agonist access to the binding pocket requires large-scale rearrangements in the highly malleable extracellular region, the structural and docking studies will therefore provide invaluable insight into the pharmacology and mechanisms of action of agonists and different classes of antagonists for the P2Y12R and potentially for other closely related P2YRs.

[Study of the effect of fluor protector on resistance to demineralization of milk beverages on enamel of primary teeth].

PURPOSE: To study the effect of fluor protector against demineralization of deciduous teeth enamel surface in milk beverages, to certify the mechanism of anti-demineralization of fluor protector, and provide laboratory basis for clinical practice of fluor protector and the prevention of infants and young children. METHODS: The enamel surfaces of 30 prepared deciduous teeth without caries were randomly divided into 3 groups. Group A was control group. Milk beverages were used in group B, and 0.1% fluor protector was used in group C. Each group included 10 specimens. All specimens were soaked in water tank at 37 degrees centigrade for 4 days, but specimens in groups B and C were soaked at intervals in 10mL yogurt drinks in water tank at 37 degrees centigrade, 9 minutes for one time and 3 times for one day. Scanning electron microscope (SEM) and spectrum analyzer were used to detect the changes in surface morphology of deciduous teeth enamel surface. The concentration values and the ratio of calcium/phosphorus were analyzed with SPSS11.5 software package. RESULTS: Enamel soaked in milk beverages had decreased Ca(2+) and P(3+) content significantly (P<0.01). Ca(2+) and P(3+) content in enamel in group C was significantly higher than in group B (P<0.01). Scanning electron microscopy showed enamel surface in group C had mild demineralization. CONCLUSIONS: Milk beverages can cause a strong acid demineralization in deciduous teeth enamel surface. The application of fluor protector in enamel surface can inhibit enamel demineralization in the milk beverages.

Structure of the human P2Y12 receptor in complex with an antithrombotic drug.

P2Y receptors (P2YRs), a family of purinergic G-protein-coupled receptors (GPCRs), are activated by extracellular nucleotides. There are a total of eight distinct functional P2YRs expressed in human, which are subdivided into P2Y1-like receptors and P2Y12-like receptors. Their ligands are generally charged molecules with relatively low bioavailability and stability in vivo, which limits our understanding of this receptor family. P2Y12R regulates platelet activation and thrombus formation, and several antithrombotic drugs targeting P2Y12R--including the prodrugs clopidogrel (Plavix) and prasugrel (Effient) that are metabolized and bind covalently, and the nucleoside analogue ticagrelor (Brilinta) that acts directly on the receptor--have been approved for the prevention of stroke and myocardial infarction. However, limitations of these drugs (for example, a very long half-life of clopidogrel action and a characteristic adverse effect profile of ticagrelor) suggest that there is an unfulfilled medical need for developing a new generation of P2Y12R inhibitors. Here we report the 2.6 Å resolution crystal structure of human P2Y12R in complex with a non-nucleotide reversible antagonist, AZD1283. The structure reveals a distinct straight conformation of helix V, which sets P2Y12R apart from all other known class A GPCR structures. With AZD1283 bound, the highly conserved disulphide bridge in GPCRs between helix III and extracellular loop 2 is not observed and appears to be dynamic. Along with the details of the AZD1283-binding site, analysis of the extracellular interface reveals an adjacent ligand-binding region and suggests that both pockets could be required for dinucleotide binding. The structure provides essential insights for the development of improved P2Y12R ligands and allosteric modulators as drug candidates.

Structure of the CCR5 chemokine receptor-HIV entry inhibitor maraviroc complex.

The CCR5 chemokine receptor acts as a co-receptor for HIV-1 viral entry. Here we report the 2.7 angstrom-resolution crystal structure of human CCR5 bound to the marketed HIV drug maraviroc. The structure reveals a ligand-binding site that is distinct from the proposed major recognition sites for chemokines and the viral glycoprotein gp120, providing insights into the mechanism of allosteric inhibition of chemokine signaling and viral entry. A comparison between CCR5 and CXCR4 crystal structures, along with models of co-receptor-gp120-V3 complexes, suggests that different charge distributions and steric hindrances caused by residue substitutions may be major determinants of HIV-1 co-receptor selectivity. These high-resolution insights into CCR5 can enable structure-based drug discovery for the treatment of HIV-1 infection.

Oxidation-sensing regulator AbfR regulates oxidative stress responses, bacterial aggregation, and biofilm formation in Staphylococcus epidermidis.

Staphylococcus epidermidis is a notorious human pathogen that is the major cause of infections related to implanted medical devices. Although redox regulation involving reactive oxygen species is now recognized as a critical component of bacterial signaling and regulation, the mechanism by which S. epidermidis senses and responds to oxidative stress remains largely unknown. Here, we report a new oxidation-sensing regulator, AbfR (aggregation and biofilm formation regulator) in S. epidermidis. An environment of oxidative stress mediated by H(2)O(2) or cumene hydroperoxide markedly up-regulates the expression of abfR gene. Similar to Pseudomonas aeruginosa OspR, AbfR is negatively autoregulated and dissociates from promoter DNA in the presence of oxidants. In vivo and in vitro analyses indicate that Cys-13 and Cys-116 are the key functional residues to form an intersubunit disulfide bond upon oxidation in AbfR. We further show that deletion of abfR leads to a significant induction in H(2)O(2) or cumene hydroperoxide resistance, enhanced bacterial aggregation, and reduced biofilm formation. These effects are mediated by derepression of SERP2195 and gpxA-2 that lie immediately downstream of the abfR gene in the same operon. Thus, oxidative stress likely acts as a signal to modulate S. epidermidis key virulence properties through AbfR.

α-Synuclein oligomers oppose long-term potentiation and impair memory through a calcineurin-dependent mechanism: relevance to human synucleopathic diseases.

Intracellular deposition of fibrillar aggregates of α-synuclein (αSyn) characterizes neurodegenerative diseases such as Parkinson's disease (PD) and dementia with Lewy bodies. However, recent evidence indicates that small αSyn oligomeric aggregates that precede fibril formation may be the most neurotoxic species and can be found extracellularly. This new evidence has changed the view of pathological αSyn aggregation from a self-contained cellular phenomenon to an extracellular event and prompted investigation of the putative effects of extracellular αSyn oligomers. In this study, we report that extracellular application of αSyn oligomers detrimentally impacts neuronal welfare and memory function. We found that oligomeric αSyn increased intracellular Ca(2+) levels, induced calcineurin (CaN) activity, decreased cAMP response element-binding protein (CREB) transcriptional activity and resulted in calcineurin-dependent death of human neuroblastoma cells. Similarly, CaN induction and CREB inhibition were observed when αSyn oligomers were applied to organotypic brain slices, which opposed hippocampal long-term potentiation. Furthermore, αSyn oligomers induced CaN, inhibited CREB and evoked memory impairments in mice that received acute intracerebroventricular injections. Notably, all these events were reversed by pharmacological inhibition of CaN. Moreover, we found decreased active CaN and reduced levels of phosphorylated CREB in autopsy brain tissue from patients affected by dementia with Lewy bodies, which is characterized by deposition of αSyn aggregates and progressive cognitive decline. These results indicate that exogenously applied αSyn oligomers impact neuronal function and produce memory deficits through mechanisms that involve CaN activation.

Percutaneous endoscopic gastrostomy/jejunostomy combined with percutaneous transhepatic biliary drainage in treating malignant biliary obstruction.

OBJECTIVE: To investigate the safety and efficacy of percutaneous endoscopic gastrostomy/jejunostomy (PEG/PEJ) combined with percutaneous transhepatic biliary drainage (PTCD) in treating malignant biliary obstruction. SUBJECTS AND METHODS: Nine patients (6 males and 3 females, average age 71.3 ± 5.5 years) with complete obstruction of the biliary tract were treated with PEG/PEJ after PTCD. The PEG/PEJ and PTCD tubes were linked outside of the abdominal wall to direct the externally drained bile back to the jejunum through the PEG/PEJ intestinal tube. Clinical symptoms and liver function were assessed following the treatment. RESULTS: The operations were successfully completed in the 9 patients within 40 min (average 35 ± 2.9 min). Clinical symptoms such as jaundice, abdominal distension, stomachache and diarrhea appeared but improved within 7 days of the operation. Serum levels of bilirubin, aspartate aminotransferase and alanine aminotransferase were reduced (p < 0.01) 4 weeks following the treatment. There were no procedural complications. CONCLUSIONS: Combined PEG/PEJ and PTCD appeared to be safe and effective in the management of malignant biliary obstruction. Further, larger-scale studies will be needed to verify findings of this report.

Structural insight into the oxidation-sensing mechanism of the antibiotic resistance of regulator MexR.

MexR functions as the primary regulator of the mexAB-oprM multidrug efflux expression in Pseudomonas aeruginosa. It has been shown that MexR senses oxidative stress by interprotomer disulphide bond formation between redox-active cysteines. This oxidation induces MexR to dissociate from the promoter DNA, thus activating the transcriptional expression of efflux pump genes. In this study, we present the crystal structure of MexR in its oxidized form at a resolution of 2.1 A. This crystal structure reveals the mechanism by which oxidative signal allosterically derepresses the MexR-controlled transcription activation.