Lipoproteins, Apoprotein A-1 containing particles and the development of atherosclerosis

This study concerns the measurement and composition of high density lipoprotein (HDL) and mechanisms through which HDL might exert the proposed protective effect against coronary heart disease (CHD). The composition of HDL, particularly in respect of particles containing apoA-I but not apoA-II (LpAI) was studied after precipitation of apoB-containing lipoproteins. Differences in HDL cholesterol, apoA-I and LpAI between commonly used methods were modest and those differences were not due to selective precipitation of LpAI. All precipitation methods were equally effective at precipitating Lp(a). Families with a history of premature CHD but without perceived risk factors showed an enhanced expression of two established risk associations - hypercholesterolaemia and/or reduced levels of HDL. LpAI concentrations were reduced in males with premature CHD and LpAI was marginally superior to apoA-I and HDL cholesterol in correctly discriminating between those with CHD and those without. Moderate exercise, sufficient to increase HDL cholesterol, could potentially reduce CHD risk. However twelve months of moderate exercise in previously sedentary, middle-aged men did not produce any significant changes in serum lipids or apolipoproteins suggesting that the potential for modifying lipoprotein profiles in men using low intensity exercise is modest. Exercise might produce changes in lipoprotein concentrations by modifying lipoprotein lipase (LPL) and hepatic lipase (HL) activity. In individuals with low and high HDL cholesterol, apoA-I and HDL cholesterol concentrations were correlated with post heparin plasma LPL and inversely correlated with HL. Furthermore HL, but not LPL, activities were inversely correlated with LpAI concentrations in both groups. The mechanisms by which HDL may protect against atherosclerosis are unclear. However the data reported here on the discriminating power of LpAI particles, the failure of exercise to influence LpAI particle concentrations and the factors which may influence LpAI concentrations in plasma, indicate that further work will be required before LpAI can truly be described as the anti-atherogenic fraction of HDL

Rapid, energy-efficient synthesis of the layered carbide, Al₄C₃

The phase-pure binary aluminium carbide, Al4C3 can be synthesised in vacuo from the elements in 30 minutes via microwave heating in a multimode cavity reactor. The success of the reaction is dependent on the use of finely divided aluminium and graphite starting materials, both of which couple effectively to the microwave field. The yellow-brown powder product was characterised by powder X-ray diffraction, scanning electron microscopy/energy dispersive X-ray spectroscopy thermogravimetric-differential thermal analysis and Raman spectroscopy. Powders were composed of hexagonal single crystallites tens of microns in diameter (rhombohedral space group R[3 with combining macron]m; Z = 3; a = 3.33813(5) Å, c = 25.0021(4) Å) and were stable to 1000 °C in air, argon and nitrogen. Equivalent microwave reactions of the elements in air led to the formation of the oxycarbide phases Al2OC and Al4O4C

Role of bacterial lipopolysaccharides in the modulation of platelet and megakaryocyte function

Cardiovascular disease is a group of inflammatory diseases that is responsible for the plurality of deaths in the world. In ischaemic diseases, aberrant platelet activation leads to thrombus formation and occlusion of key arteries. Furthermore, platelets are critically involved in sepsis, where thrombocytopaenia is correlated with worse patient outcomes. Therefore, immune receptors, such as Toll-like receptor 4 (TLR4), have been identified as potential targets for reducing platelet activation by decoupling immune functions from haemostasis. Multiple experimental techniques were used to determine the impact of ultrapure lipopolysaccharide (LPS) chemotypes on platelet and megakaryocyte function as LPS is a specific ligand for TLR4. To determine whether LPS modulates platelet activity, LPS and platelets were co-incubated under different conditions. We determined that ultrapure LPS derived from various species of Gram-negative bacteria cannot significantly modulate platelet activity, as measured by aggregation, fibrinogen binding, and P-selectin exposure, except under specific conditions. This is potentially due to activation dependent cell�surface expression of TLR4. Subsequently, to investigate whether LPS induces inflammatory signalling in megakaryocytes, a novel megakaryocyte reporter cell line, Meg-01R, was developed and used to determine that ultrapure LPS is not sufficient on its own to modulate megakaryocyte function. Characterisation of this cell line suggests that MyD88-dependent signalling pathways are active in Meg-01R cells but TLR4 is not present in sufficient quantities at the cell surface. As LL37 directly binds to LPS and is also a strong platelet agonist, we investigated the impact of LPS on LL37- induced platelet activation. Here, we discovered a TLR4-independent cell-independent formation of LL37-LPS micelles that prevents LL37-induced platelet activation. Based on these results, LPS and TLR4 are not sufficient, in vitro, to decouple the immune function of platelets from haemostasis or induce changes in megakaryocyte function however it may still play an important role in conjunction with other immune receptors

Toll-like receptor 4 signalling and its impact on platelet function, thrombosis, and haemostasis

Platelets are non-nucleated blood cells that participate in a wide range of physiological and pathological functions. Their major role is mediating haemostasis and thrombosis. In addition to these classic functions, platelets have emerged as important players in the innate immune system. In particular, they interact with leukocytes, secrete pro- and anti-inflammatory factors, and express a wide range of inflammatory receptors including Toll-like receptors (TLRs) e.g. Toll-like receptor 4 (TLR4). TLR4, which is the most extensively studied TLR in nucleated cells, recognises lipopolysaccharides (LPS) that are compounds of the outer surface of Gram-negative bacteria. Unlike other TLRs, TLR4 is able to signal through both the MyD88-dependent and -independent signalling pathways. Notably, despite both pathways culminating in activation of transcription factors, TLR4 has a prominent functional impact on platelet activity, haemostasis, and thrombosis. In this review, we summarise the current knowledge on the TLR4 signalling in platelets, critically discuss its impact on platelet function, and highlight the open questions in this area

Development and characterisation of a novel NF-κB reporter cell line for investigation of neuroinflammation

Aberrant activation of the transcription factor NF-κB, as well as uncontrolled inflammation has been linked to autoimmune diseases, development and progression of cancer and neurological disorders like Alzheimer’s disease. Reporter cell lines are a valuable state-of-the art tool for comparative analysis of in vitro drug screening. However, a reporter cell line for the investigation of NF-κB-driven neuroinflammation has not yet been available. Thus, we developed a stable neural NF-κB-reporter cell line to assess the potency of pro-inflammatory molecules and peptides, as well as anti-inflammatory pharmaceuticals. We used lentivirus to transduce the glioma cell line U251-MG with a tandem NF-κB reporter construct containing GFP and firefly luciferase allowing an assessment of NF-κB activity via fluorescence microscopy, flow cytometry and luminometry. We observed a robust activation of NF-κB after exposure of the reporter cell line to Tumour 2 necrosis factor alpha (TNFα) and amyloid-β peptide [1-42] as well as to LPS derived from Salmonella minnesota and Escherichia coli. Finally, we demonstrate that the U251-NF-κB-GFP-Luc reporter cells can be used for assessing the anti-inflammatory potential of pharmaceutical compounds using Bay11-7082 and IMD0354. In summary, our newly generated cell line is a robust and cost-efficient tool to study pro- and anti-inflammatory potential of drugs and biologicals in neural cells

Risk of myocardial infarction and stroke after acute infection or vaccination.

BACKGROUND: There is evidence that chronic inflammation may promote atherosclerotic disease. We tested the hypothesis that acute infection and vaccination increase the short-term risk of vascular events. METHODS: We undertook within-person comparisons, using the case-series method, to study the risks of myocardial infarction and stroke after common vaccinations and naturally occurring infections. The study was based on the United Kingdom General Practice Research Database, which contains computerized medical records of more than 5 million patients. RESULTS: A total of 20,486 persons with a first myocardial infarction and 19,063 persons with a first stroke who received influenza vaccine were included in the analysis. There was no increase in the risk of myocardial infarction or stroke in the period after influenza, tetanus, or pneumococcal vaccination. However, the risks of both events were substantially higher after a diagnosis of systemic respiratory tract infection and were highest during the first three days (incidence ratio for myocardial infarction, 4.95; 95 percent confidence interval, 4.43 to 5.53; incidence ratio for stroke, 3.19; 95 percent confidence interval, 2.81 to 3.62). The risks then gradually fell during the following weeks. The risks were raised significantly but to a lesser degree after a diagnosis of urinary tract infection. The findings for recurrent myocardial infarctions and stroke were similar to those for first events. CONCLUSIONS: Our findings provide support for the concept that acute infections are associated with a transient increase in the risk of vascular events. By contrast, influenza, tetanus, and pneumococcal vaccinations do not produce a detectable increase in the risk of vascular events

Repurposing cancer drugs, batimastat and marimastat, to inhibit the activity of a group I metalloprotease from the venom of the Western Diamondback rattlesnake, Crotalus atrox

Snakebite envenomation causes over 140,000 deaths every year predominantly in developing countries. As a result, it is one of the most lethal neglected tropical diseases. It is associated with an incredibly complex pathophysiology due to the vast number of unique toxins/proteins found in the venoms of diverse snake species found worldwide. Here, we report the purification and functional characteristics of a group I metalloprotease (CAMP-2) from the venom of the western diamondback rattlesnake, Crotalus atrox. Its sensitivity to matrix metalloprotease inhibitors (batimastat and marimastat) was established using specific in vitro experiments and in silico molecular docking analysis. CAMP-2 shows high sequence homology to atroxase from the venom of Crotalus atrox and exhibits collagenolytic, fibrinogenolytic and mild haemolytic activities. It exerts a mild inhibitory effect on agonist-induced platelet aggregation in the absence of plasma proteins. Its collagenolytic activity was completely inhibited by batimastat and marimastat. Zinc chloride also inhibits the collagenolytic activity of CAMP-2 by around 75% at 50 M, while it is partially potentiated by calcium chloride. Molecular docking studies demonstrate that batimastat and marimastat are able to bind strongly to the active site residues of CAMP-2. This study demonstrates the impact of matrix metalloprotease inhibitors in the modulation of a purified, group I metalloprotease activities in comparison to the whole venom. By improving our understanding of snake venom metalloproteases and their sensitivity to small molecule inhibitors, we can begin to develop novel and improved treatment strategies for snakebites

Synthetic flavonoids as novel modulators of platelet function and thrombosis

Cardiovascular diseases represent a major cause of mortality and morbidity in the world and thrombotic conditions such as heart attacks and strokes are caused by unwarranted activation of platelets and subsequent formation of blood clots (thrombi) within the blood vessels during pathological circumstances. Therefore, platelets act as a primary therapeutic target to treat and prevent thrombotic conditions. Current treatments are limited due to intolerance and they are associated with severe side effects such as bleeding complications. Hence, the development of novel therapeutic strategies for thrombotic diseases is an urgent priority. Flavonoids are naturally occurring plant-derived molecules that exert numerous beneficial effects in humans through modulating the functions of distinct cell types. However, naturally occurring flavonoids suffer from several issues such as poor solubility, lipophilicity, and bioavailability, which hinder their efficacy and potency. Despite this, flavonoids act as versatile templates for the design and synthesis of novel molecules for various therapeutic targets. Indeed, several synthetic flavonoids have recently been developed to improve their stability, bioavailability and efficacy including for the modulation of platelet function. Here, we provide insight into the actions of certain natural flavonoids along with the advantages of synthetic flavonoids in the modulation of platelet function, haemostasis and thrombosis

The 2 × 2 model of perfectionism and school‐ and community‐based sport participation.

The authors adopted the 2 × 2 model of perfectionism to examine the unique and interactive effects of two dimensions of perfectionism (personal standards perfectionism PSP and evaluative concerns perfectionism ECP) on personal and interpersonal indicators of participant experience in youth sport (enjoyment, physical self‐worth, and friendship quality). Participants (N = 219, M age = 15.12, SD = 2.02) were recruited from various school‐ and community‐based sports and completed a multi‐section questionnaire. Consideration of main and interaction effects indicated that pure PSP (high PSP/low ECP) was associated with the most positive sport experience and pure ECP (low PSP/high ECP) was associated with the least positive sport experience. The findings suggest that subtypes of perfectionism from the 2 × 2 model are predictive of differing experiences in youth sport participation. (PsycINFO Database Record (c) 2015 APA, all rights reserved). (journal abstract