Mapping transmembrane residues of proteinase activated recpetor 2 (PAR2) that influence ligand-modulated calcium signaling

Proteinase-activated receptor 2 (PAR(2)) is a G protein -coupled receptor involved in metabolism, inflammation, and cancers. It is activated by proteolysis, which exposes a nascent N -terminal sequence that becomes a tethered agonist. Short synthetic peptides corresponding to this sequence also activate PAR(2), while small organic molecules show promising PAR(2) antagonism. Developing PAR(2) ligands into pharmaceuticals is hindered by a lack of knowledge of how synthetic ligands interact with and differentially modulate PAR(2). Guided by PAR(2) homology modeling and ligand docking based on bovine rhodopsin, followed by cross-checking with newer PAR(2) models based on ORL-1 and PART, site-directed mutagenesis of PAR(2) was used to investigate the pharmacology of three agonists (two synthetic agonists and trypsin-exposed tethered ligand) and one antagonist for modulation of PAR(2) signaling. Effects of 28 PAR2 mutations were examined for PAR(2)-mediated calcium mobilization and key mutants were selected for measuring ligand binding. Nineteen of twenty-eight PAR(2) mutations reduced the potency of at least one ligand by>10-fold. Key residues mapped predominantly to a cluster in the transmembrane (TM) domains of PAR(2), differentially influence intracellular Ca2+ induced by synthetic agonists versus a native agonist, and highlight subtly different TM residues involved in receptor activation. This is the first evidence highlighting the importance of the PAR(2) TM regions for receptor activation by synthetic PAR(2) agonists and antagonists. The trypsin-cleaved N-terminus that activates PAR(2) was unaffected by residues that affected synthetic peptides, challenging the widespread practice of substituting peptides for proteases to characterize PAR(2) physiology. (C) 2017 Elsevier Ltd. All rights reserved

Modulating mitophagy in mitochondrial disease

Mitochondrial diseases may result from mutations in the maternally-inherited mitochondrial DNA (mtDNA) or from mutations in nuclear genes encoding mitochondrial proteins. Their bi-genomic nature makes mitochondrial diseases a very heterogeneous group of disorders that can present at any age and can affect any type of tissue. The autophagic-lysosomal degradation pathway plays an important role in clearing dysfunctional and redundant mitochondria through a specific quality control mechanism termed mitophagy. Mitochondria could be targeted for autophagic degradation for a variety of reasons including basal turnover for recycling, starvation induced degradation, and degradation due to damage. While the core autophagic machinery is highly conserved and common to most pathways, the signaling pathways leading to the selective degradation of damaged mitochondria are still not completely understood. Type 1 mitophagy due to nutrient starvation is dependent on PI3K (phosphoinositide 3-kinase) for autophagosome formation but independent of mitophagy proteins, PINK1 (PTEN-induced putative kinase 1) and Parkin. Whereas type 2 mitophagy that occurs due to damage is dependent on PINK1 and Parkin but does not require PI3K. Autophagy and mitophagy play an important role in human disease and hence could serve as therapeutic targets for the treatment of mitochondrial as well as neurodegenerative disorders. Therefore, we reviewed drugs that are known modulators of autophagy (AICAR and metformin) and may effect this by activating the AMP-activated protein kinase signaling pathways. Furthermore, we reviewed data available on supplements, such as Coenzyme Q and the quinone idebenone, that we assert rescue increased mitophagy in mitochondrial disease by benefiting mitochondrial function

Preparation of porous PMMA/Na+-montmorillonite cation-exchange membranes for cationic dye adsorption

Porous PMMA/Na+-montmorillonite (MMT) cation-exchange membranes were successfully prepared by entrapment method in this study. One approach (simple mixing) was to mix commercial PMMA polymer with Na+-MMT clays in solvent for membrane preparation (Membrane A). The other approach (emulsion polymerization) was to synthesize the PMMA/Na+-MMT polymer composite via emulsion polymerization first, followed by membrane casting (Membrane B for Kunipia F clays and Membrane C for PK-802 clays). Membrane morphology and properties were characterized. The thermogravimetric analysis (TGA) verified the near complete incorporation of feed Na+-MMT clays in the PMMA/Na+-MMT composite membranes, while X-ray diffractograms (WXRD) exhibited the slightly enlarged interlayer spacing of Na+-MMT. The range of cation-exchange capacity (CEC) was 9-32 mu equiv./47 mm disc. For batch cationic dye adsorption, the best performance was achieved by Membrane B with feed Na+-MMT/MMA (M/P) ratio (w/w) = 0.5 and Membrane C with feed M/P = 0.6, where about 95% Methyl violet adsorption was attained in 2 h. The optimal desorption solution was 1 M KSCN in 80% methanol and its related dye desorption efficiency was 92%. In the flow process using one piece of 47 mm disc of Membrane B (M/P = 0.5), dye solution was recirculated for 6 h and >= 85% dye could be removed. Higher than 94% of dye was desorbed at 1 or 4 mL/min, and the membrane regenerability was proved by successfully performing three consecutive cycles. (C) 2008 Elsevier B.V. All rights reserved

Preparation of polysulfone-based cation-exchange membranes and their application in protein separation with a plate-and-frame module

In this study, polysulfone-based cation-exchange membranes were successfully prepared through a sulfonation procedure and a membrane preparation process. By comparing the results from different conditions, the most favorable sulfonation condition is to react at 75 degreesC for 4 h, whereas the optimal preparation process is casting a 70 degreesC solution of 2 g sulfonated polysulfone in a 8 ml mixture of polyethylene glycol: 1-methyl-2-pyrrolidinone (v:v) = 5:3, exposing the film to an air of 50% relative humidity at room temperature for 3 min, and then immersing it into a 27 degreesC water bath. The prepared cation-exchange membranes (90 pin thick) had a water content of 0.68 and an ion-exchange capacity of 2.9 mueq/cm(2). In the batch adsorption at pH 7.4, the saturation capacity of cationic lysozyme was 0.0098 mumol/cm(2) and anionic BSA was not adsorbed. In the flow adsorption process with a 10-piece membrane stack in the plate-and-frame module, the results show that lysozyme and BSA were effectively separated and the lysozyme recovery was higher than 93%. The effects of inlet flow rate and operation mode were insignificant on the separation performance. In the last part of this paper, the isolation of lysozyme from hen egg white was conducted in a flow process with only one piece of the prepared cation-exchange membrane. A purification effectiveness of 20.7 and a lysozyme recovery of 51.1% were achieved at a flow rate of 10 ml/min. (C) 2004 Elsevier B.V. All rights reserved

Neurological Manifestations of Coronavirus Disease 2019: A Comprehensive Review and Meta-Analysis of the First 6 Months of Pandemic Reporting

Background: There is growing evidence that SARS-Cov-2 infection is associated with severe neurological complications. Understanding the nature and prevalence of these neurologic manifestations is essential for identifying higher-risk patients and projecting demand for ongoing resource utilisation. This review and meta-analysis report the neurologic manifestations identified in hospitalised COVID-19 patients and provide a preliminary estimate of disease prevalence. Methods: MEDLINE, Embase and Scopus were searched for studies reporting the occurrence of neurological complications in hospitalised COVID-19 patients. Results: A total of 2,207 unique entries were identified and screened, among which 14 cohort studies and 53 case reports were included, reporting on a total of 8,577 patients. Central nervous system manifestations included ischemic stroke (n = 226), delirium (n = 79), intracranial haemorrhage (ICH, n = 57), meningoencephalitis (n = 13), seizures (n = 3), and acute demyelinating encephalitis (n = 2). Peripheral nervous system manifestations included Guillain-Barrè Syndrome (n = 21) and other peripheral neuropathies (n = 3). The pooled period prevalence of ischemic stroke from identified studies was 1.3% [95%CI: 0.9–1.8%, 102/7,715] in all hospitalised COVID-19 patients, and 2.8% [95%CI: 1.0–4.6%, 9/318] among COVID-19 patients admitted to ICU. The pooled prevalence of ICH was estimated at 0.4% [95%CI: 0–0.8%, 6/1,006]. Conclusions: The COVID-19 pandemic exerts a substantial neurologic burden which may have residual effects on patients and healthcare systems for years. Low quality evidence impedes the ability to accurately predict the magnitude of this burden. Robust studies with standardised screening and case definitions are required to improve understanding of this disease and optimise treatment of individuals at higher risk for neurologic sequelae.SCOPUS: re.jinfo:eu-repo/semantics/publishe