New insights on the involvement of Nerve Growth Factor in allergic inflammation and fibrosis

Nerve Growth Factor (NGF), that was originally discovered for its properties of stimulating growth and differentiation of neurons, is now also considered responsible for several activities in the immune system and beyond. Mast cells and eosinophils, key cells of allergic inflammation, are a source of NGF and are influenced by it. These observations have prompted studies on NGF in allergy and tissue repair. Recent evidences link NGF and these two processes. While NGF is clearly a new tool in the management of untreatable ulcers, its role in allergic inflammation, although appearing to be pro-inflammatory, is still not clearly defined.     

The variance of icosahedral virus models is a key indicator in the structure determination: a model-free reconstruction of viruses,suitable for refractory particles

A model-free method to determine the three-dimensional structure of icosahedral viruses is described. The novel strategy is based upon the approximate principle that correct virus structures have high variance as do all other well-detailed structures, even wrong ones. The original projections of individual particles are reduced to a radius of 25 pixels and are used to compute single particle reconstruction models by assigning them 1800 different Euler triads. The variance of the models obtained from all projections is stored in maps and a decimation process is carried out. In a first stage, thresholds are adopted for the variance values, and in a second stage, carried out by correspondence analysis and classification, 30 clusters of models are sorted out. The clusters are refined to yield models contained in boxes of 64(3) voxels. The refined models with highest variance and closest similarity represent the correct solution. Once enlarged, these models can be used to align all available projections in their original scale in a customary projection-matching process. The method has proved successful in determining the structures of poliovirus, of the empty and filled capsids of L-A virus, and of a modified capsid of hepatitis B virus.     

Metabolic derangement and cardiac injury early after reperfusion following intermittent cross-clamp fibrillation in patients undergoing coronary artery bypass graft surgery using conventional or miniaturized cardiopulmonary bypass

Reactive oxygen species (ROS)-induced oxidative stress increases in skeletal muscle with aging and decreases the viability of implanted cells. Type 1 insulin-like growth factor (IGF-1) promotes the survival of skeletal muscle cells under oxidative stress. It is unknown whether IGF-1 protects muscle-derived stem cells (MDSCs) from oxidative stress. In this study, we genetically engineered rat MDSCs to overexpress IGF-1 and determined cell viability, apoptosis, and VEGF secretion under oxidative stress. Overexpression of IGF-1 prevented MDSCs from H₂O₂-induced caspase-dependent apoptotic cell death by upregulating the PI3K/AKT pathway, accompanied with an increase of NF-κB, p-NF-κB, Bcl-2, and VEGF, as well as a decrease of Bax. In contrast, pre-administration of picropodophyllinb, wortmannin, 1L-6-hydroxymethyl-chiro-inositol-2-((R)-2-O-methyl-3-O-octadecylcarbonate), or pyrrolidine-dithiocarbamate, specific inhibitors of IGF-1R, PI3K, AKT, and NF-κB, respectively, followed by treatment with H₂O₂, resulted in cell death of MDSCs. Our data indicated that IGF-1 suppresses apoptosis and enhances the paracrine function of MDSCs under oxidative stress via enhancing IGF-1R/PI3K/AKT signaling. Thus, IGF-1 gene-modified MDSCs present a potential application in the treatment of muscle wasting, such as urethra intrinsic sphincter deficiency.     

ELF-MF attenuates quercetin-induced apoptosis in K562 cells through modulating the expression of Bcl-2 family proteins

This study investigated the effects of sinusoidal ELF-MF (1 mT; 50 Hz) on the apoptosis induced by four different compounds, namely vinblastine, etoposide, quercetin, and resveratrol, in human K562 chronic myeloid leukemia cells. The exposure to ELF-MF did not affect growth and viability of untreated K562 cells and did not influence the anti-proliferative effects of resveratrol, vinblastine, and etoposide. On the contrary, in quercetin-treated cells, exposure to ELF-MF significantly reduced the percentage of apoptotic cells and the caspase-3 activity and modified the cell cycle profile especially after 48 h of exposure. In addition, the simultaneous treatments for 24 h with quercetin plus ELF-MF increased Bcl-2 protein expression and prevented quercetin-induced downregulation of Mcl-1 and Bcl-xL. Finally, an increase of HSP70 expression was also observed after prolonged ELF-MF treatment. The ELF-MF-dependent modulation of the expression of anti-apoptotic Bcl-2 family and Hsp70 proteins could act as a pro-survival mechanism in K562 cells.     

Glucose oxidase from Penicillium amagasakiense: Characterization of the transition state of its denaturation from molecular dynamics simulations

Glucose oxidase (GOx) is a flavoenzyme having applications in food and medical industries. However, GOx, as many other enzymes when extracted from the cells, has relatively short operational lifetimes. Several recent studies (both experimental and theoretical), carried out on small proteins (or small fractions of large proteins), show that a detailed knowledge of how the breakdown process starts and proceeds on molecular level could be of significant help to artificially improve the stability of fragile proteins. We have performed extended molecular dynamics (MD) simulations to study the denaturation of GOx (a protein dimer containing nearly 1200 amino acids) to identify weak points in its structure and in this way gather information to later make it more stable, for example, by mutations. A denaturation of a protein can be simulated by increasing the temperature far above physiological temperature. We have performed a series of MD simulations at different temperatures (300, 400, 500, and 600 K). The exit from the protein's native state has been successfully identified with the clustering method and supported by other methods used to analyze the simulation data. A common set of amino acids is regularly found to initiate the denaturation, suggesting a moiety where the enzyme could be strengthened by a suitable amino acid based modification. Proteins 2014; 82:2353–2363. © 2014 Wiley Periodicals, Inc.     

Acylated and unacylated ghrelin protect MC3T3-E1 cells against tert-butyl hydroperoxide-induced oxidative injury: pharmacological characterization of ghrelin receptor and possible epigenetic involvement

Increasing evidence suggests a role for oxidative stress in age-related decrease in osteoblast number and function leading to the development of osteoporosis. This study was undertaken to investigate whether ghrelin, previously reported to stimulate osteoblast proliferation, counteracts tert-butyl hydroperoxide (t-BHP)-induced oxidative damage in MC3T3-E1 osteoblastic cells as well as to characterize the ghrelin receptor (GHS-R) involved in such activity. Pretreatment with ghrelin (10^?7–10^?11 M) significantly increased viability and reduced apoptosis of MC3T3-E1 cells cultured with t-BHP (250 μM) for three hours at the low concentration of 10^?9 M as shown by MTT assay and Hoechst-33258 staining. Furthermore, ghrelin prevented t-BHP-induced osteoblastic dysfunction and changes in the cytoskeleton organization evidenced by the staining of the actin fibers with Phalloidin-FITC by reducing reactive oxygen species generation. The GHS-R type 1a agonist, EP1572 (10^?7–10^?11 M), had no effect against t-BHP-induced cytotoxicity and pretreatment with the selective GHS-R1a antagonist, d-Lys³-GHRP-6 (10^?7 M), failed to remove ghrelin (10^?9 M)-protective effects against oxidative injury, indicating that GHS-R1a is not involved in such ghrelin activity. Accordingly, unacylated ghrelin (DAG), not binding GHS-R1a, displays the same protective actions of ghrelin against t-BHP-induced cytotoxicity. Preliminary observations indicate that ghrelin increased the trimethylation of lys4 on histones H3, a known epigenetic mark activator, which may regulate the expression of some genes limiting oxidative damage. In conclusion, our data demonstrate that ghrelin and DAG promote survival of MC3T3-E1 cell exposed to t-BHP-induced oxidative damage. Such effect is independent of GHS-R1a and is likely mediated by a common ghrelin/DAG binding site.     

Direct chromatographic enantioresolution of fully constrained β-amino acids: exploring the use of high-molecular weight chiral selectors

To the best of our knowledge enantioselective chromatographic protocols on β-amino acids with polysaccharide-based chiral stationary phases (CSPs) have not yet appeared in the literature. Therefore, the primary objective of this work was the development of chromatographic methods based on the use of an amylose derivative CSP (Lux Amylose-2), enabling the direct normal-phase (NP) enantioresolution of four fully constrained β-amino acids. Also, the results obtained with the glycopeptide-type Chirobiotic T column employed in the usual polar-ionic (PI) mode of elution are compared with those achieved with the polysaccharide-based phase. The Lux Amylose-2 column, in combination with alkyl sulfonic acid containing NP eluent systems, prevailed over the Chirobiotic T one, when used under the PI mode of elution, and hence can be considered as the elective choice for the enantioseparation of this class of rigid β-amino acids. Moreover, the extraordinarily high α (up to 4.60) and R _S (up to 10.60) values provided by the polysaccharidic polymer, especially when used with camphor sulfonic acid containing eluent systems, make it also suitable for preparative-scale enantioisolations.     

Safety and efficacy of lactoferrin versus ferrous sulphate in curing iron deficiency and iron deficiency anaemia in hereditary thrombophilia pregnant women: an interventional study

Objective Evaluate the safety and efficacy of bovine lactoferrin (bLf) versus the ferrous sulphate standard intervention in curing iron deficiency (ID) and ID anaemia (IDA) in pregnant women affected by hereditary thrombophilia (HT). Design Interventional study. Setting Secondary-level hospital for complicated pregnancies in Rome, Italy. Population 295 HT pregnant women (≥18 years) suffering from ID/IDA. Methods Women were enrolled in Arm A or B in accordance with their personal choice. In Arm A, 156 women received oral administration of 100 mg of bLf twice a day; in Arm B, 139 women received 520 mg of ferrous sulphate once a day. Therapies lasted until delivery. Main outcome measures Red blood cells, haemoglobin, total serum iron, serum ferritin (haematological parameters) were assayed before and every 30 days during therapy until delivery. Serum IL-6, key factor in inflammatory and iron homeostasis disorders, was detected at enrolment and after therapy at delivery. Possible maternal, foetal, and neonatal adverse effects were assessed. Results Haematological parameters were significantly higher in Arm A than in Arm B pregnant women (P ≤ 0.0001). Serum IL-6 significantly decreased in bLf-treated women and increased in ferrous sulphate-treated women. BLf did not exert any adverse effect. Adverse effects in 16.5 % of ferrous sulphate-treated women were recorded. Arm A women experienced no miscarriage compared to five miscarriages in Arm B women. Conclusions Differently from ferrous sulphate, bLf is safe and effective in curing ID/IDA associated with a consistent decrease of serum IL-6. The absence of miscarriage among bLf-treated women provided an unexpected benefit. Trial registration: ClinicalTrials.gov Identifier NCT01221844.