Is low dose inhaled corticosteroid therapy as effective for inflammation and remodeling in asthma? A randomized,parallel group study

Background

While most of the clinical benefits of inhaled corticosteroid (ICS) therapy may occur at low doses, results of dose-ranging studies are inconsistent. Although symptom/lung function response to low and high dose ICS medication is comparable, it is uncertain whether low dose ICSs are as effective as high dose in the treatment of inflammation and remodeling.

Methods

22 mild or moderate asthmatic adult subjects (corticosteroid free for > 2 months) participated in a randomized, parallel group study to compare effects of fluticasone propionate (FP) 200 mcg/day and 1000 mcg/day. Alveolar macrophage (AM)-derived cytokines and basement membrane thickness (BMT) were measured at baseline and after 7 weeks treatment while symptoms, spirometry, exhaled nitric oxide (eNO) and airway hyperresponsiveness (AHR) to mannitol at baseline and 6 weeks.

Results

FP improved spirometry, eNO, symptoms and AHR with no difference between low and high dose FP. Both high and low dose FP reduced GM-CSF, TNF-alpha and IL-1ra, with no change in BMT and with no differences between low and high dose FP.

Conclusions

200 μg/day of FP was as effective as 1000 μg/day in improving asthma control, airway inflammation, lung function and AHR in adults in the short term. Future studies should examine potential differential effects between low and high dose combination therapy (ICS/long acting beta agonist) on inflammation and airway remodeling over longer treatment periods.     

Intercellular communication via the endo-lysosomal system: Translocation of granzymes through membrane barriers

Cytotoxic lymphocytes (CLs) are responsible for the clearance of virally infected or neoplastic cells. CLs possess specialised lysosome-related organelles called granules which contain the granzyme family of serine proteases and perforin. Granzymes may induce apoptosis in the target cell when delivered by the pore forming protein, perforin. Here we follow the perforin-granzyme pathway from synthesis and storage in the granule, to exocytosis and finally delivery into the target cell. This review focuses on the controversial subject of perforin-mediated translocation of granzymes into the target cell cytoplasm. It remains unclear whether this occurs at the cell surface with granzymes moving through a perforin pore in the plasma membrane, or if it involves internalisation of perforin and granzymes and subsequent release from an endocytic compartment. The latter mechanism would represent an example of cross talk between the endo-lysosomal pathways of individual cells. This article is part of a Special Issue entitled: Proteolysis 50 years after the discovery of lysosome.     

Nanostructured p‐n Junctions for Kinetic‐to‐Electrical Energy Conversion

Piezoelectric ZnO nanorods grown on a flexible substrate are combined with the p‐type semiconducting polymer PEDOT:PSS to produce a p‐n junction device that successfully demonstrates kinetic‐to‐electrical energy conversion. Both the voltage and current output of the devices are measured to be in the range of 10 mV and 10 μA cm^?2. Combining these figures for the best device gives a maximum possible power density of 0.4 mW cm^?3. Systematic testing of the devices is performed showing that the voltage output increases linearly with applied stress, and is reduced significantly by illumination with super‐band gap light. This provides strong evidence that the voltage output results from piezoelectric effects in the ZnO. The behavior of the devices is explained by considering the time‐dependent changes in band structure resulting from the straining of a piezoelectric material within a p‐n junction. It is shown that the rate of screening of the depolarisation field determines the power output of a piezoelectric energy harvesting device. This model is consistent with the behavior of a number of previous devices utilising the piezoelectric effect in ZnO.     

Choline-releasing glycerophosphodiesterase EDI3 links the tumor metabolome to signaling network activities

Recently, EDI3 was identified as a key factor for choline metabolism that controls tumor cell migration and is associated with metastasis in endometrial carcinomas. EDI3 cleaves glycerophosphocholine (GPC) to form choline and glycerol-3-phosphate (G3P). Choline is then further metabolized to phosphatidylcholine (PtdC), the major lipid in membranes and a key player in membrane-mediated cell signaling. The second product, G3P, is a precursor molecule for several lipids with central roles in signaling, for example lysophosphatidic acid (LPA), phosphatidic acid (PA) and diacylglycerol (DAG). LPA activates intracellular signaling pathways by binding to specific LPA receptors, including membrane-bound G protein-coupled receptors and the intracellular nuclear receptor, PPARγ. Conversely, PA and DAG mediate signaling by acting as lipid anchors that bind and activate several signaling proteins. For example, binding of GTPases and PKC to PA and DAG, respectively, increases the activation of signaling networks, mediating processes such as migration, adhesion, proliferation or anti-apoptosis—all relevant for tumor development. We present a concept by which EDI3 either directly generates signaling molecules or provides “membrane anchors” for downstream signaling factors. As a result, EDI3 links choline metabolism to signaling activities resulting in a more malignant phenotype.     

Synthesis of N-phenyl-N-(3-(piperidin-1-yl)propyl)benzofuran-2-carboxamides as new selective ligands for sigma receptors

Novel benzofuran-2-carboxamide ligands, which are selective for sigma receptors, have been synthesized via a microwave-assisted Perkin rearrangement reaction and a modified Finkelstein halogen-exchange used to facilitate N-alkylation. The ligands synthesized are the 3-methyl-N-phenyl-N-(3-(piperidin-1-yl)propyl)benzofuran-2-carboxamides (KSCM-1, KSCM-5 and KSCM-11). The benzofuran-2-carboxamide structure was N-arylated and N-alkylated to include both N-phenyl and N-(3-(piperidin-1-yl)propyl substituents, respectively. These new carboxamides exhibit high affinity at the sigma-1 receptor with K_i values ranging from 7.8 to 34 nM. Ligand KSCM-1 with two methoxy substituents at C-5 and C-6 of the benzofuran ring, and K_i = 27.5 nM at sigma-1 was found to be more selective for sigma-1 over sigma-2.     

EM-fold: de novo atomic-detail protein structure determination from medium-resolution density maps

Electron density maps of membrane proteins or large macromolecular complexes are frequently only determined at medium resolution between 4?? and 10??, either by cryo-electron microscopy or X-ray crystallography. In these density maps, the general arrangement of secondary structure elements (SSEs) is revealed, whereas their directionality and connectivity remain elusive. We demonstrate that the topology of proteins with up to 250 amino acids can be determined from such density maps when combined with a computational protein folding protocol. Furthermore, we accurately reconstruct atomic detail in loop regions and amino acid side chains not visible in the experimental data. The EM-Fold algorithm assembles the SSEs de novo before atomic detail is added using Rosetta. In a benchmark of 27 proteins, the protocol consistently and reproducibly achieves models with root mean square deviation values <3??.     

The Balance of Apoptotic and Necrotic Cell Death in Mycobacterium tuberculosis Infected Macrophages Is Not Dependent on Bacterial Virulence

Background

An important mechanism of Mycobacterium tuberculosis pathogenesis is the ability to control cell death pathways in infected macrophages: apoptotic cell death is bactericidal, whereas necrotic cell death may facilitate bacterial dissemination and transmission.

Methods

We examine M.tuberculosis control of spontaneous and chemically induced macrophage cell death using automated confocal fluorescence microscopy, image analysis, flow cytometry, plate-reader based vitality assays, and M.tuberculosis strains including H37Rv, and isogenic virulent and avirulent strains of the Beijing lineage isolate GC1237.

Results

We show that bacterial virulence influences the dynamics of caspase activation and the total level of cytotoxicity. We show that the powerful ability of M.tuberculosis to inhibit exogenously stimulated apoptosis is abrogated by loss of virulence. However, loss of virulence did not influence the balance of macrophage apoptosis and necrosis – both virulent and avirulent isogenic strains of GC1237 induced predominantly necrotic cell death compared to H37Rv which induced a higher relative level of apoptosis.

Conclusions

This reveals that macrophage necrosis and apoptosis are independently regulated during M. tuberculosis infection of macrophages. Virulence affects the level of host cell death and ability to inhibit apoptosis but other strain-specific characteristics influence the ultimate mode of host cell death and alter the balance of apoptosis and necrosis.     

First detection of Nosema ceranae, a microsporidian parasite of European honey bees (Apis mellifera), in Canada and central USA

Nosema ceranae is an emerging microsporidian parasite of European honey bees, Apis mellifera, but its distribution is not well known. Six Nosema-positive samples (determined from light microscopy of spores) of adult worker bees from Canada (two each from Nova Scotia, New Brunswick, and Prince Edward Island) and two from USA (Minnesota) were tested to determine Nosema species using previously-developed PCR primers of the 16S rRNA gene. We detected for the first time N. ceranae in Canada and central USA. One haplotype of N. ceranae was identified; its virulence may differ from that of other haplotypes.