GRT-X Stimulates Dorsal Root Ganglia Axonal Growth in Culture via TSPO and Kv7.2/3 Potassium Channel Activation.

GRT-X, which targets both the mitochondrial translocator protein (TSPO) and the Kv7.2/3 (KCNQ2/3) potassium channels, has been shown to efficiently promote recovery from cervical spine injury. In the present work, we investigate the role of GRT-X and its two targets in the axonal growth of dorsal root ganglion (DRG) neurons. Neurite outgrowth was quantified in DRG explant cultures prepared from wild-type C57BL6/J and TSPO-KO mice. TSPO was pharmacologically targeted with the agonist XBD173 and the Kv7 channels with the activator ICA-27243 and the inhibitor XE991. GRT-X efficiently stimulated DRG axonal growth at 4 and 8 days after its single administration. XBD173 also promoted axonal elongation, but only after 8 days and its repeated administration. In contrast, both ICA27243 and XE991 tended to decrease axonal elongation. In dissociated DRG neuron/Schwann cell co-cultures, GRT-X upregulated the expression of genes associated with axonal growth and myelination. In the TSPO-KO DRG cultures, the stimulatory effect of GRT-X on axonal growth was completely lost. However, GRT-X and XBD173 activated neuronal and Schwann cell gene expression after TSPO knockout, indicating the presence of additional targets warranting further investigation. These findings uncover a key role of the dual mode of action of GRT-X in the axonal elongation of DRG neurons.

The Comprehensive Steroidome in Complete TSPO/PBR Knockout Mice under Basal Conditions.

The 18 kDa translocator protein (TSPO/PBR) is a multifunctional evolutionary highly conserved outer mitochondrial membrane protein. Decades of research has reported an obligatory role of TSPO/PBR in both mitochondrial cholesterol transport and, thus, steroid production. However, the strict dependency of steroidogenesis on TSPO/PBR has remained controversial. The aim of this study was to provide insight into the steroid profile in complete C57BL/6-Tspotm1GuWu(GuwiyangWurra)-knockout male mice (TSPO-KO) under basal conditions. The steroidome in the brain, adrenal glands, testes and plasma was measured by gas chromatography coupled to tandem mass spectrometry (GC-MS/MS). We found that steroids present in wild-type (WT) mice were also detected in TSPO-KO mice, including pregnenolone (PREG), progestogens, mineralo-glucocorticosteroids and androgens. The concentrations of PREG and most metabolites were similar between genotypes, except a significant decrease in the levels of the 5α-reduced metabolites of progesterone (PROG) in adrenal glands and plasma and of the 5α-reduced metabolites of corticosterone (B) in plasma in TSPO-KO compared to WT animals, suggesting other regulatory functions for the TSPO/PBR. The expression levels of the voltage-dependent anion-selective channel (VDAC-1), CYP11A1 and 5α-reductase were not significantly different between both groups. Thus, the complete deletion of the tspo gene in male mice does not impair de novo steroidogenesis in vivo.

C-type lectin-like receptor 2 and zonulin are associated with mild cognitive impairment and Alzheimer's disease.

OBJECTIVE: Increased permeability and changes in gut microbiota contributed to the pathogenesis of Alzheimer's disease (AD). Zonulin is a key modulator that regulates intestinal barrier function. Peripheral platelet alterations have been involved in AD pathology. C-type lectin-like receptor 2 (CLEC-2) is a receptor on the platelet surface for activation. The purpose of this study was to determine zonulin and CLEC-2 levels in mild cognitive impairment (MCI) and AD, and investigate the relationship between zonulin and CLEC-2. METHODS: In this study, CLEC-2 and zonulin levels were measured using ELISA assay in 110 AD patients, 110 MCI patients, and 110 non-demented control subjects. RESULTS: Increased CLEC-2 and zonulin levels were observed in MCI and AD patients. Furthermore, AD patients had higher CLEC-2 and zonulin levels compared with MCI patients. In addition, CLEC-2 levels were positively correlated with zonulin levels, after adjusting confounding factors (r = .592, P < .001). Multivariate analysis revealed that increased CLEC-2 and zonulin levels were significantly associated with reduced Mini-Mental State Examination (MMSE) score. CONCLUSIONS: C-type lectin-like receptor 2 is correlated with zonulin after adjusting confounding covariates. Moreover, increased CLEC-2 and zonulin are the significant factors for reduced MMSE score in MCI and AD. Further studies are needed.

DNA physical properties outperform sequence compositional information in classifying nucleosome-enriched and -depleted regions.

The nucleosome is the fundamental structural unit of eukaryotic chromatin and plays an essential role in the epigenetic regulation of cellular processes, such as DNA replication, recombination, and transcription. Hence, it is important to identify nucleosome positions in the genome. Our previous model based on DNA deformation energy, in which a set of DNA physical descriptors was used, performed well in predicting nucleosome dyad positions and occupancy. In this study, we established a machine-learning model for predicting nucleosome occupancy in order to further verify the physical descriptors. Results showed that (1) our model outperformed several other sequence compositional information-based models, indicating a stronger dependence of nucleosome positioning on DNA physical properties; (2) nucleosome-enriched and -depleted regions have distinct features in terms of DNA physical descriptors like sequence-dependent flexibility and equilibrium structure parameters; (3) gene transcription start sites and termination sites can be well characterized with the distribution patterns of the physical descriptors, indicating the regulatory role of DNA physical properties in gene transcription. In addition, we developed a web server for the model, which is freely accessible at http://lin-group.cn/server/iNuc-force/.

Calcium-axonemal microtubuli interactions underlie mechanism(s) of primary cilia morphological changes.

We have used cell culture of astrocytes aligned within microchannels to investigate calcium effects on primary cilia morphology. In the absence of calcium and in the presence of flow of media (10 µL.s-1) the majority (90%) of primary cilia showed reversible bending with an average curvature of 2.1 ± 0.9 × 10-4 nm-1. When 1.0 mM calcium was present, 90% of cilia underwent bending. Forty percent of these cilia demonstrated strong irreversible bending, resulting in a final average curvature of 3.9 ± 1 × 10-4 nm-1, while 50% of cilia underwent bending similar to that observed during calcium-free flow. The average length of cilia was shifted toward shorter values (3.67 ± 0.34 µm) when exposed to excess calcium (1.0 mM), compared to media devoid of calcium (3.96 ± 0.26 µm). The number of primary cilia that became curved after calcium application was reduced when the cell culture was pre-incubated with 15 µM of the microtubule stabilizer, taxol, for 60 min prior to calcium application. Calcium caused single microtubules to curve at a concentration ≈1.0 mM in vitro, but at higher concentration (≈1.5 mM) multiple microtubule curving occurred. Additionally, calcium causes microtubule-associated protein-2 conformational changes and its dislocation from the microtubule wall at the location of microtubule curvature. A very small amount of calcium, that is 1.45 × 1011 times lower than the maximal capacity of TRPPs calcium channels, may cause gross morphological changes (curving) of primary cilia, while global cytosol calcium levels are expected to remain unchanged. These findings reflect the non-linear manner in which primary cilia may respond to calcium signaling, which in turn may influence the course of development of ciliopathies and cancer.

Cellular Sources and Regional Variations in the Expression of the Neuroinflammatory Marker Translocator Protein (TSPO) in the Normal Brain.

The inducible expression of the mitochondrial translocator protein 18 kDa (TSPO) by activated microglia is a prominent, regular feature of acute and chronic-progressive brain pathology. This expression is also the rationale for the continual development of new TSPO binding molecules for the diagnosis of "neuroinflammation" by molecular imaging. However, there is in the normal brain an ill-defined, low-level constitutive expression of TSPO. Taking advantage of healthy TSPO knockout mouse brain tissue to validate TSPO antibody specificity, this study uses immunohistochemistry to determine the regional distribution and cellular sources of TSPO in the normal mouse brain. Fluorescence microscopy revealed punctate TSPO immunostaining in vascular endothelial cells throughout the brain. In the olfactory nerve layers and glomeruli of the olfactory bulb, choroid plexus and ependymal layers, we confirm constitutive TSPO expression levels similar to peripheral organs, while some low TSPO expression is present in regions of known neurogenesis, as well as cerebellar Purkinje cells. The distributed-sparse expression of TSPO in endothelial mitochondria throughout the normal brain can be expected to give rise to a low baseline signal in TSPO molecular imaging studies. Finally, our study emphasises the need for valid and methodologically robust verification of the selectivity of TSPO ligands through the use of TSPO knockout tissues.

Synthesis characterization and cytotoxicity studies of platinum(II) complexes with reduced amino pyridine schiff base and its derivatives as ligands.

A series of reduced amino pyridine Schiff base platinum(II) complexes were prepared as potential anticancer drugs, and characterized by NMR, IR spectroscopy, elemental analysis, and molar conductivity. UV and CD results showed the binding mode between these compounds and salmon sperm DNA may be intercalation. The cytotoxicity of these complexes was validated against A549, Hela, and MCF-7 cell lines by MTT assay. Some complexes exhibited better cytotoxic activity than cisplatin against Hela and MCF-7 cell lines.

Checkpoints to the Brain: Directing Myeloid Cell Migration to the Central Nervous System.

Myeloid cells are a unique subset of leukocytes with a diverse array of functions within the central nervous system during health and disease. Advances in understanding of the unique properties of these cells have inspired interest in their use as delivery vehicles for therapeutic genes, proteins, and drugs, or as "assistants" in the clean-up of aggregated proteins and other molecules when existing drainage systems are no longer adequate. The trafficking of myeloid cells from the periphery to the central nervous system is subject to complex cellular and molecular controls with several 'checkpoints' from the blood to their destination in the brain parenchyma. As important components of the neurovascular unit, the functional state changes associated with lineage heterogeneity of myeloid cells are increasingly recognized as important for disease progression. In this review, we discuss some of the cellular elements associated with formation and function of the neurovascular unit, and present an update on the impact of myeloid cells on central nervous system (CNS) diseases in the laboratory and the clinic. We then discuss emerging strategies for harnessing the potential of site-directed myeloid cell homing to the CNS, and identify promising avenues for future research, with particular emphasis on the importance of untangling the functional heterogeneity within existing myeloid subsets.

Investigating the interactions of the 18kDa translocator protein and its ligand PK11195 in planar lipid bilayers.

The functional effects of a drug ligand may be due not only to an interaction with its membrane protein target, but also with the surrounding lipid membrane. We have investigated the interaction of a drug ligand, PK11195, with its primary protein target, the integral membrane 18kDa translocator protein (TSPO), and model membranes using Langmuir monolayers, quartz crystal microbalance with dissipation monitoring (QCM-D) and neutron reflectometry (NR). We found that PK11195 is incorporated into lipid monolayers and lipid bilayers, causing a decrease in lipid area/molecule and an increase in lipid bilayer rigidity. NR revealed that PK11195 is incorporated into the lipid chain region at a volume fraction of ~10%. We reconstituted isolated mouse TSPO into a lipid bilayer and studied its interaction with PK11195 using QCM-D, which revealed a larger than expected frequency response and indicated a possible conformational change of the protein. NR measurements revealed a TSPO surface coverage of 23% when immobilised to a modified surface via its polyhistidine tag, and a thickness of 51Å for the TSPO layer. These techniques allowed us to probe both the interaction of TSPO with PK11195, and PK11195 with model membranes. It is possible that previously reported TSPO-independent effects of PK11195 are due to incorporation into the lipid bilayer and alteration of its physical properties. There are also implications for the variable binding profiles observed for TSPO ligands, as drug-membrane interactions may contribute to the apparent affinity of TSPO ligands.

Guwiyang Wurra--'Fire Mouse': a global gene knockout model for TSPO/PBR drug development, loss-of-function and mechanisms of compensation studies.

The highly conserved 18-kDa translocator protein (TSPO) or peripheral benzodiazepine receptor (PBR), is being investigated as a diagnostic and therapeutic target for disease conditions ranging from inflammation to neurodegeneration and behavioural illnesses. Many functions have been attributed to TSPO/PBR including a role in the mitochondrial permeability transition pore (MPTP), steroidogenesis and energy metabolism. In this review, we detail the recent developments in determining the physiological role of TSPO/PBR, specifically based on data obtained from the recently generated Tspo knockout mouse models. In addition to defining the role of TSPO/PBR, we also describe the value of Tspo knockout mice in determining the selectivity, specificity and presence of any off-target effects of TSPO/PBR ligands.

Structure and Potential Cellular Targets of HAMLET-like Anti-Cancer Compounds made from Milk Components.

The HAMLET family of compounds (Human Alpha-lactalbumin Made Lethal to Tumours) was discovered during studies on the properties of human milk, and is a class of protein-lipid complexes having broad spectrum anti-cancer, and some specific anti-bacterial properties. The structure of HAMLET-like compounds consists of an aggregation of partially unfolded protein making up the majority of the compound's mass, with fatty acid molecules bound in the hydrophobic core. This is a novel protein-lipid structure and has only recently been derived by small-angle X-ray scattering analysis. The structure is the basis of a novel cytotoxicity mechanism responsible for anti-cancer activity to all of the around 50 different cancer cell types for which the HAMLET family has been trialled. Multiple cytotoxic mechanisms have been hypothesised for the HAMLET-like compounds, but it is not yet clear which of those are the initiating cytotoxic mechanism(s) and which are subsequent activities triggered by the initiating mechanism(s). In addition to the studies into the structure of these compounds, this review presents the state of knowledge of the anti-cancer aspects of HAMLET-like compounds, the HAMLET-induced cytotoxic activities to cancer and non-cancer cells, and the several prospective cell membrane and intracellular targets of the HAMLET family. The emerging picture is that HAMLET-like compounds initiate their cytotoxic effects on what may be a cancer-specific target in the cell membrane that has yet to be identified. This article is open to POST-PUBLICATION REVIEW. Registered readers (see "For Readers") may comment by clicking on ABSTRACT on the issue's contents page.

[Optical Properties Investigation of p-Type ZnO Film Based on Doping by Diffusion].

The main purpose of this paper is to investigate the optical properties of p-type ZnO film based on P doping. ZnO film was grown by Atomic layer deposition (ALD) on InP subsrate in this experiment, and phosphorus diffused into ZnO lattice by annealing treatment at different temperature (500, 700 °C). The optical properties of samples were investigated by photoluminescence (PL) spectroscopy, which indicated that the annealing temperature is the important factor influencing the phosphorus diffusion doping. The low-temperature PL spectra of the sample which annealed at 700 °C for 1 h exhibited acceptor related emission peaks located at 3.351, 3.311, 3.246 and 3.177 eV, which were attributed to A °X, FA, DAP and DA-1LO, respectively. The acceptor binding energy is estimated to be about 122 meV, which is agreed with the theoretic values in phosphorus-doped ZnO films. In this paper, through thermal diffusion method to realize the p-type doped ZnO thin films, it solved the main problems which limited the development of ZnO based optoelectronic devices, and has an important significance for the development of the ZnO semiconductor materials and ZnO based photoelectric device.

Cytotoxicity of phosphoramidate, bis-amidate and cycloSal prodrug metabolites against tumour and normal cells.

Phosphonate and phosphate prodrugs are integral to enhancing drug permeability, but the potential toxicity of their metabolites requires careful consideration. This study evaluates the impact of widely used phosphoramidate, bis-amidate, and cycloSal phosph(on)ate prodrug metabolites on BxPC3 pancreatic cancer cells, GL261-Luc glioblastoma cells, and primary cultured mouse astrocytes. 1-Naphthol and 2-naphthol demonstrated the greatest toxicity. Notably, 2-naphthol exhibited an ED50 of 21 µM on BxPC3 cells, surpassing 1-naphthol with an ED50 of 82 µM. Real-time xCELLigence experiments revealed notable activity for both metabolites at a low concentration of 16 µM. On primary cultured mouse astrocyte cells, all prodrugs exhibited reduced viability at 128 to 256 µM after only 4 hours of exposure. A cell-type-dependent sensitivity to phosph(on)ate prodrug metabolites was evident, with normal cells showing greater susceptibility than corresponding tumour cells. The results suggest it is essential to consider the potential cytotoxicity of phosph(on)ate prodrugs in the drug design and evaluation process.

Diagnostic Value of a Combined Serum α-Hydroxybutyrate Dehydrogenase, Carcinoembryonic Antigen and Glycoantigen 125 Test for Early-Stage Breast Cancer.

Objective: To investigate the diagnostic value of the combined detection of α-hydroxybutyrate dehydrogenase (α-HBDH), carcinoembryonic antigen (CEA) and cancer antigen 125 (CA125) in early-stage breast cancer (ESBC). Methods: This was a retrospective analysis of 169 patients with ESBC, 138 patients with benign breast disease (BBD) and 200 normal healthy controls (NHCs). The levels of serum α-HBDH, CEA and CA125 in the two groups were detected. The receiver operating characteristic (ROC) curve and area under the curve (AUC) were used to analyse the diagnostic value of the above indicators alone and in combination for ESBC. Results: The levels of α-HBDH, CEA and CA125 in the ESBC group were significantly higher than those in the BBD and NHC groups ([118.18 ± 11.19 vs 91.24 ± 9.17 vs 89.38 ± 9.01, F = 6.189, p = 0.004], [2.39 ± 1.12 vs 1.48 ± 0.76 vs 1.58 ± 0.58, F = 5.362, p = 0.017] and [14.44 ± 6.78 vs 11.19 ± 3.17 vs 7.18 ± 4.71, F = 8.912, p = 0.001], respectively). In the ESBC group, the positive rate of combined detection was higher than that of single detection (96.12% vs 72.64% vs 53.67% vs 42.41%, X2 = 27.174, p < 0.05). ROC curve analysis showed that serum α-HBDH, CEA, CA125 alone and combined detection in the diagnosis of ESBC. The sensitivity was 48.1%, 63.6%, 44.2% and 54.5%, the specificity was 75.4%, 75.4%, 86.0% and 91.2% and the AUC was 0.654, 0.715, 0.636 and 0.772, respectively. The diagnostic value of combined detection was the highest. Conclusion: The levels of serum α-HBDH, CEA and CA125 in ESBC are high, and the combined detection of the three has a high diagnostic value for ESBC.

The cancer-testis antigen a-kinase anchor protein 3 facilitates breast cancer progression via activation of the PTEN/PI3K/AKT/mTOR signaling.

The cancer-testis antigen A-kinase anchor protein 3 (AKAP3) has been shown to have a strong association with breast cancer (BC). However, its role in BC progression received scant attention. We aimed to explore the prognostic implication of aberrant AKAP3 expression for a better knowledge of BC progression and improved treatment. AKAP3 expression was quantitated using tissue microarrays and immunohistochemistry (IHC). Cell viability, invasion, migration, apoptosis, and expressions of PTEN/PI3K/AKT/mTOR signaling components were assessed in AKAP3-overexpressed or si-AKAP3-transfected BC cells. Finally, elevated AKAP3 expression was observed in BC versus paracancerous tissues. BC patients with high AKAP3 expression showed a worse prognosis than low expression patients (P < 0.0001). AKAP3 overexpressions fueled cell growth, proliferation, migration, and invasion in HCC1937 and MDA-MB-468 BC cell lines, alongside increased expressions of PI3K/AKT/mTOR signaling components and PTEN suppression. These effects were pronouncedly reversed, together with elevated apoptosis, in cells transfected with si-AKAP3. Therefore, AKAP3 is upregulated in BC and promotes BC cell growth, invasion, and migration via PTEN/PI3K/AKT/mTOR signaling activation. It may serve as a prognosis indicator for BC survival.

Long-term diazepam treatment enhances microglial spine engulfment and impairs cognitive performance via the mitochondrial 18 kDa translocator protein (TSPO).

Benzodiazepines are widely administered drugs to treat anxiety and insomnia. In addition to tolerance development and abuse liability, their chronic use may cause cognitive impairment and increase the risk for dementia. However, the mechanism by which benzodiazepines might contribute to persistent cognitive decline remains unknown. Here we report that diazepam, a widely prescribed benzodiazepine, impairs the structural plasticity of dendritic spines, causing cognitive impairment in mice. Diazepam induces these deficits via the mitochondrial 18 kDa translocator protein (TSPO), rather than classical γ-aminobutyric acid type A receptors, which alters microglial morphology, and phagocytosis of synaptic material. Collectively, our findings demonstrate a mechanism by which TSPO ligands alter synaptic plasticity and, as a consequence, cause cognitive impairment.

Large aperture vertical cavity surface emitting laser with distributed-ring contact.

To overcome the serious current crowding effect in top-emitting vertical cavity surface emitting lasers (VCSELs) with large aperture, a distributed-ring-contact (DRC) VCSEL is proposed and demonstrated. A maximal cw light output power of more than 0.3 W and a wall-plug efficiency of 17.4% are achieved for a 300 µm-diameter VCSEL. The DRC VCSEL exhibits a more homogeneous emission profile, and the laser emits at 803.3 nm with a narrow spectrum (less than 0.2 nm FWHM).

[Correlation between distributions of pathogenic bacteria on hands and the position of funeral staffs].

OBJECTIVE: This study aims to investigate the bacteria contamination on hands of funeral staffs in different positions. METHODS: Bacterial samples were collected from the hands of 105 funeral staffs in different positions (including 90 frontline staffs and 15 administrative workers) from 13 funeral parlors nationwide, and were subsequently tested by bacterium inspection. RESULTS: In total, 1783 strains of bacteria were isolated, including 1027 Gram-positive bacteria, most of which were Staphylococcus; and 756 Gram-negative bacteria, most of which were Pseudomonas. Out of the 1783 strains of bacteria, 570 pathogens and opportunistic pathogens were isolated, accounted to 31.96%. The isolated ratio of pathogens and conditional pathogens in embalmed/cosmetologist of cadavers was 35.67% (370/1037), which was higher than those in the funeral workers in other positions, such as cremators, pick-up and administrative workers, whose ratios were 24.42% (95/389), 22.41% (52/232) and 10.40% (12/125), respectively (χ(2) were 13.682, 10.967 and 32.263, respectively; P values were all < 0.05). And the isolated ratios of pathogens and conditional pathogens in cremators and pick-up workers were significantly higher than that in administrative workers (χ(2) were 11.206 and 7.873, respectively; P values were all < 0.05). CONCLUSION: Lots of bacteria were found in the samples from hands of funeral staffs. The isolated ratio of pathogens and conditional pathogens was different between the funeral staffs in different positions; while the highest was from embalmed/cosmetologist of cadavers and the lowest was from administrators.

Control of Neuroinflammation through Radiation-Induced Microglial Changes.

Microglia, the innate immune cells of the central nervous system, play a pivotal role in the modulation of neuroinflammation. Neuroinflammation has been implicated in many diseases of the CNS, including Alzheimer's disease and Parkinson's disease. It is well documented that microglial activation, initiated by a variety of stressors, can trigger a potentially destructive neuroinflammatory response via the release of pro-inflammatory molecules, and reactive oxygen and nitrogen species. However, the potential anti-inflammatory and neuroprotective effects that microglia are also thought to exhibit have been under-investigated. The application of ionising radiation at different doses and dose schedules may reveal novel methods for the control of microglial response to stressors, potentially highlighting avenues for treatment of neuroinflammation associated CNS disorders, such as Alzheimer's disease and Parkinson's disease. There remains a need to characterise the response of microglia to radiation, particularly low dose ionising radiation.

Steric Hindrance- and Work Function-Promoted High Performance for Electrochemical CO Methanation on Antisite Defects of MoS2 and WS2.

CO methanation from electrochemical CO reduction reaction (CORR) is significant for sustainable environment and energy, but electrocatalysts with excellent selectivity and activity are still lacking. Selectivity is sensitive to the structure of active sites, and activity can be tailored by work function. Moreover, intrinsic active sites usually possess relatively high concentration compared to artificial ones. Here, antisite defects MoS2 and WS2 , intrinsic atomic defects of MoS2 and WS2 with a transition metal atom substituting a S2 column, were investigated for CORR by density functional theory calculations. The steric hindrance from the special bowl structure of MoS2 and WS2 ensured good selectivity towards CO methanation. Coordination environment variation of the active sites, the under-coordinated Mo or W atoms, effectively lowered the work function, making MoS2 and WS2 highly active for CO methanation with the required potential of -0.47 and -0.49 V vs. reversible hydrogen electrode, respectively. Moreover, high concentration of active sites and minimal structural deformation during the catalytic process of MoS2 and WS2 enhanced their attraction for future commercial application.