Endothelial function and endogenous fibrinolysis in inflammation and ischaemic heart disease

The vascular endothelium plays a vital role in the control of blood flow, haemostasis, fibrinolysis and inflammation. Impairment of endothelial vasomotor and fibrinolytic function is implicated in the pathogenesis of ischaemic heart disease. Atherosclerosis is now widely recognised to be an inflammatory disease process, but the mechanistic links between inflammation, endothelial dysfunction and endogenous fibrinolysis remain poorly understood.In a series of studies, we explored the effects of inflammation on endothelial vasomotion and fibrinolytic capacity using an in vivo forearm model of systemic and local inflammation. Systemic inflammation stimulated by typhoid vaccination had no major effect on vasomotor tone. However, tumour necrosis factor-a induced local vascular inflammation was associated with impaired resistance vessel endotheliumdependent vasodilatation, possibly through the development of acute arterial injury. Both systemic and local inflammation were found to augment the acute release of endothelial tissue plasminogen activator. In addition, intra-arterial tumour necrosis factor-a administration resulted in a unique profile of substantial and sustained local increase in endogenous tissue plasminogen activator.We extended these investigations and assessed the role of endogenous fibrinolysis and endothelial dysfunction in the pathogenesis of prothrombotic conditions such as hyperhomocysteinaemia and coronary stent thrombosis or in-stent restenosis using this forearm model of endothelial function assessment. In patients with recent myocardial infarction, elevation of plasma homocysteine concentration was associated with impaired endothelium-dependent vasodilatation but not endogenous fibrinolysis. This vasomotor dysfunction was not rectified by vitamin supplementation. We also assessed three critical aspects of vascular function in patients who have undergone percutaneous coronary intervention and found no evidence that endothelial vasomotor, fibrinolytic or platelet function play a major role in the pathogenesis of acute stent thrombosis or in-stent restenosis.We conclude that there is complex interaction between inflammation, endothelial function and endogenous fibrinolysis. We have identified the unique role of systemic inflammation and specifically, tumour necrosis factor-a. Furthermore, our studies indicated that there were no significant associations between fibrinolysis and hyperhomocysteinaemia or coronary stent complications. Therefore modulating cytokine actions and their interaction with fibrinolysis may be critical in the prevention of thrombotic coronary occlusion and myocardial ischaemia as well as in the future development of anti-thrombotic therapies

Interpenetration as a Mechanism for Liquid-Liquid Phase Transitions

We study simple lattice systems to demonstrate the influence of interpenetrating bond networks on phase behavior. We promote interpenetration by using a Hamiltonian with a weakly repulsive interaction with nearest neighbors and an attractive interaction with second-nearest neighbors. In this way, bond networks will form between second-nearest neighbors, allowing for two (locally) distinct networks to form. We obtain the phase behavior from analytic solution in the mean-field approximation and exact solution on the Bethe lattice. We compare these results with exact numerical results for the phase behavior from grand canonical Monte Carlo simulations on square, cubic, and tetrahedral lattices. All results show that these simple systems exhibit rich phase diagrams with two fluid-fluid critical points and three thermodynamically distinct phases. We also consider including third-nearest-neighbor interactions, which give rise to a phase diagram with four critical points and five thermodynamically distinct phases. Thus the interpenetration mechanism provides a simple route to generate multiple liquid phases in single-component systems, such as hypothesized in water and observed in several model and experimental systems. Additionally, interpenetration of many such networks appears plausible in a recently considered material made from nanoparticles functionalized by single strands of DNA.Comment: 12 pages, 9 figures, submitted to Phys. Rev.

Structural basis for the identification of the N-terminal domain of coronavirus nucleocapsid protein as an antiviral target

Coronaviruses (CoVs) cause numerous diseases, including Middle East respiratory syndrome and severe acute respiratory syndrome, generating significant health-related and economic consequences. CoVs encode the nucleocapsid (N) protein, a major structural protein that plays multiple roles in the virus replication cycle and forms a ribonucleoprotein complex with the viral RNA through the N protein's N-terminal domain (N-NTD). Using human CoV-OC43 (HCoV-OC43) as a model for CoV, we present the 3D structure of HCoV-OC43 N-NTD complexed with ribonucleoside 5'-monophosphates to identify a distinct ribonucleotide-binding pocket. By targeting this pocket, we identified and developed a new coronavirus N protein inhibitor, N-(6-oxo-5,6-dihydrophenanthridin-2-yl)(N,N-dimethylamino)acetamide hydrochloride (PJ34), using virtual screening; this inhibitor reduced the N protein's RNA-binding affinity and hindered viral replication. We also determined the crystal structure of the N-NTD-PJ34 complex. On the basis of these findings, we propose guidelines for developing new N protein-based antiviral agents that target CoVs

Participation of annexin 1 in the response of Arabidopsis thaliana to lead exposure: potential for phytoremediation

Heavy metal pollution has become a serious public health and environmental concern. Lead (Pb) is one of the heavy metals known to bioaccumulate in plants. Phytoremediation is an emerging technology based on the ability of green plants to remove Pb from the environment in a cost-efficient and ecologically sound manner. Currently, an important research focus is to seek a better understanding of the mechanisms of Pb tolerance by plant cells, with the aim of genetically engineering plants with improved tolerance to Pb, and hence better phytoremediation capabilities in the near future. Annexin, a calcium-dependent membrane-binding protein is believed to play a role in many essential cellular processes. It has been shown that expression of annexin genes from Arabidopsis thaliana are differentially regulated in response to a variety of abiotic stresses. Thus annexins are likely be involved in the response of plants to heavy metal stress. This study aimed to obtain new insights into whether annexin 1 (AnnAt1), is involved in Pb tolerance in plant cells. Message levels of AnnAt1 were assessed in response to Pb treatments using quantitative real-time PCR. Expression results were analysed using REST 2008 and normalized against the mitosis protein YLS8. We found that Pb effect on AnnAt1 expression in plants exposed to lower Pb concentrations (25 µM, 50 µM, and 75 µM) was not significantly different from the controls. However, AnnAt1 message levels doubled (2.12-fold, S.E. range is 1.77 - 2.61, p < 0.001) in seedlings treated with 100 µM Pb, in comparison to the control plants. The relative contribution of AnnAt1 in defence against Pb stress will be discussed

Ribosomal oxygenases are structurally conserved from prokaryotes to humans

2-Oxoglutarate (2OG)-dependent oxygenases have important roles in the regulation of gene expression via demethylation of N-methylated chromatin components1,2 and in the hydroxylation of transcription factors3 and splicing factor proteins4. Recently, 2OG-dependent oxygenases that catalyse hydroxylation of transfer RNA5,6,7 and ribosomal proteins8 have been shown to be important in translation relating to cellular growth, TH17-cell differentiation and translational accuracy9,10,11,12. The finding that ribosomal oxygenases (ROXs) occur in organisms ranging from prokaryotes to humans8 raises questions as to their structural and evolutionary relationships. In Escherichia coli, YcfD catalyses arginine hydroxylation in the ribosomal protein L16; in humans, MYC-induced nuclear antigen (MINA53; also known as MINA) and nucleolar protein 66 (NO66) catalyse histidine hydroxylation in the ribosomal proteins RPL27A and RPL8, respectively. The functional assignments of ROXs open therapeutic possibilities via either ROX inhibition or targeting of differentially modified ribosomes. Despite differences in the residue and protein selectivities of prokaryotic and eukaryotic ROXs, comparison of the crystal structures of E. coli YcfD and Rhodothermus marinus YcfD with those of human MINA53 and NO66 reveals highly conserved folds and novel dimerization modes defining a new structural subfamily of 2OG-dependent oxygenases. ROX structures with and without their substrates support their functional assignments as hydroxylases but not demethylases, and reveal how the subfamily has evolved to catalyse the hydroxylation of different residue side chains of ribosomal proteins. Comparison of ROX crystal structures with those of other JmjC-domain-containing hydroxylases, including the hypoxia-inducible factor asparaginyl hydroxylase FIH and histone Nε-methyl lysine demethylases, identifies branch points in 2OG-dependent oxygenase evolution and distinguishes between JmjC-containing hydroxylases and demethylases catalysing modifications of translational and transcriptional machinery. The structures reveal that new protein hydroxylation activities can evolve by changing the coordination position from which the iron-bound substrate-oxidizing species reacts. This coordination flexibility has probably contributed to the evolution of the wide range of reactions catalysed by oxygenases

Percutaneous coronary intervention in asians- are there differences in clinical outcome?

<p>Abstract</p> <p>Background</p> <p>Ethnic differences in clinical outcome after percutaneous coronary intervention (PCI) have been reported. Data within different Asian subpopulations is scarce. We aim to explore the differences in clinical profile and outcome between Chinese, Malay and Indian Asian patients who undergo PCI for coronary artery disease (CAD).</p> <p>Methods</p> <p>A prospective registry of consecutive patients undergoing PCI from January 2002 to December 2007 at a tertiary care center was analyzed. Primary endpoint was major adverse cardiovascular events (MACE) of myocardial infarction (MI), repeat revascularization and all-cause death at six months.</p> <p>Results</p> <p>7889 patients underwent PCI; 7544 (96%) patients completed follow-up and were included in the analysis (79% males with mean age of 59 years ± 11). There were 5130 (68%) Chinese, 1056 (14%) Malays and 1001 (13.3%) Indian patients. The remaining 357 (4.7%) patients from other minority ethnic groups were excluded from the analysis. The primary end-point occurred in 684 (9.1%) patients at six months. Indians had the highest rates of six month MACE compared to Chinese and Malays (Indians 12% vs. Chinese 8.2% vs. Malays 10.7%; OR 1.55 95%CI 1.24-1.93, p < 0.001). This was contributed by increased rates of MI (Indians 1.9% vs. Chinese 0.9% vs. Malays 1.3%; OR 4.49 95%CI 1.91-10.56 p = 0.001), repeat revascularization (Indians 6.5% vs. Chinese 4.1% vs. Malays 5.1%; OR 1.64 95%CI 1.22-2.21 p = 0.0012) and death (Indians 11.4% vs. Chinese 7.6% vs. Malays 9.9%; OR 1.65 95%CI 1.23-2.20 p = 0.001) amongst Indian patients.</p> <p>Conclusion</p> <p>These data indicate that ethnic variations in clinical outcome exist following PCI. In particular, Indian patients have higher six month event rates compared to Chinese and Malays. Future studies are warranted to elucidate the underlying mechanisms behind these variations.</p

Bioactive Peptide Improves Diet-Induced Hepatic Fat Deposition and Hepatocyte Proinflammatory Response in SAMP8 Ageing Mice

Background/Aims: High-fat diet (HFD)-induced nonalcoholic fatty liver disease (NAFLD) poses therapeutic challenges in elderly subjects. Due to lack of efficient drug therapy, plant-based bioactive peptides have been studied as alternative strategy in NAFLD and for less toxicity in elderly. To mimic fatty liver in aging conditions, researchers highly commended the genetically engineered strains SAMP8 (senescence-accelerated mice prone 8). However, there is a paucity of reports about the anti-steatosis effects of bioactive peptides against fatty liver development under a combined action of high-fat diet exposure and aging process. This study was conducted to evaluate the activity of DIKTNKPVIF peptide synthesized from alcalase-generated potato protein hydrolysate (PH), on reducing HFD-driven and steatosis-associated proinflammatory reaction in ageing model. Methods: Five groups of six-month-old SAMP8 mice (n=4, each) were fed either a normal chow (NC group) for 14 weeks upon sacrifice, or induced with a 6-week HFD feeding, then treated without (HCO group) or with an 8-week simultaneous administration of peptide (HPEP group), protein (HPH group) or probucol (HRX group). Liver organs were harvested from each group for histological analysis and immunoblot assay. Results: In contrast to NC, extensive fat accumulation was visualized in the liver slides of HCO. Following the trends of orally administered PH, intraperitoneally injected peptide reduces hepatic fat deposition and causes at protein level, a significant decrease in HFD-induced proinflammatory mediators p-p38 MAPK, FGF-2, TNF-α, IL-6 with concomitant reactivation of AMPK. However, p-Foxo1 and PPAR-α levels were slightly changed. Conclusion: Oral supplementation of PH and intraperitoneal injection of derived bioactive peptide alleviate proinflammatory reaction associated with hepatosteatosis development in elderly subjects, through activation of AMPK

An Abundant Dysfunctional Apolipoprotein A1 in Human Atheroma

Recent studies have indicated that high-density lipoproteins (HDLs) and their major structural protein, apolipoprotein A1 (apoA1), recovered from human atheroma are dysfunctional and are extensively oxidized by myeloperoxidase (MPO). In vitro oxidation of either apoA1 or HDL particles by MPO impairs their cholesterol acceptor function. Here, using phage display affinity maturation, we developed a high-affinity monoclonal antibody that specifically recognizes both apoA1 and HDL that have been modified by the MPO-H2O2-Cl− system. An oxindolyl alanine (2-OH-Trp) moiety at Trp72 of apoA1 is the immunogenic epitope. Mutagenesis studies confirmed a critical role for apoA1 Trp72 in MPO-mediated inhibition of the ATP-binding cassette transporter A1 (ABCA1)-dependent cholesterol acceptor activity of apoA1 in vitro and in vivo. ApoA1 containing a 2-OH-Trp72 group (oxTrp72-apoA1) is in low abundance within the circulation but accounts for 20% of the apoA1 in atherosclerosis-laden arteries. OxTrp72-apoA1 recovered from human atheroma or plasma is lipid poor, virtually devoid of cholesterol acceptor activity and demonstrated both a potent proinflammatory activity on endothelial cells and an impaired HDL biogenesis activity in vivo. Elevated oxTrp72-apoA1 levels in subjects presenting to a cardiology clinic (n = 627) were associated with increased cardiovascular disease risk. Circulating oxTrp72-apoA1 levels may serve as a way to monitor a proatherogenic process in the artery wall

An Abundant Dysfunctional Apolipoprotein A1 in Human Atheroma

Recent studies have indicated that high-density lipoproteins (HDLs) and their major structural protein, apolipoprotein A1 (apoA1), recovered from human atheroma are dysfunctional and are extensively oxidized by myeloperoxidase (MPO). In vitro oxidation of either apoA1 or HDL particles by MPO impairs their cholesterol acceptor function. Here, using phage display affinity maturation, we developed a high-affinity monoclonal antibody that specifically recognizes both apoA1 and HDL that have been modified by the MPO-H2O2-Cl− system. An oxindolyl alanine (2-OH-Trp) moiety at Trp72 of apoA1 is the immunogenic epitope. Mutagenesis studies confirmed a critical role for apoA1 Trp72 in MPO-mediated inhibition of the ATP-binding cassette transporter A1 (ABCA1)-dependent cholesterol acceptor activity of apoA1 in vitro and in vivo. ApoA1 containing a 2-OH-Trp72 group (oxTrp72-apoA1) is in low abundance within the circulation but accounts for 20% of the apoA1 in atherosclerosis-laden arteries. OxTrp72-apoA1 recovered from human atheroma or plasma is lipid poor, virtually devoid of cholesterol acceptor activity and demonstrated both a potent proinflammatory activity on endothelial cells and an impaired HDL biogenesis activity in vivo. Elevated oxTrp72-apoA1 levels in subjects presenting to a cardiology clinic (n = 627) were associated with increased cardiovascular disease risk. Circulating oxTrp72-apoA1 levels may serve as a way to monitor a proatherogenic process in the artery wall