PPARgamma activation induces CD36 expression and stimulates foam cell like changes in rVSMCs

The purpose of the present study was to determine the role of peroxisome proliferator-activated receptor gamma (PPARgamma) activation in smooth muscle cell (SMC) derived form cell formation. Wild and mutant type PPARgamma were delivered by adenovirus then activated with troglitazone. The result of Oil Red O staining and FACS analysis showed that PPARgamma activation induced lipid accumulation in rVSMCs. Furthermore, PPARgamma activation reduced SMC marker genes such as alpha-actin while induced adipocyte differentiation marker genes and lipid metabolism-related genes as evidenced by RT-PCR and fluorescent immunocytochemistry. All these data demonstrate that PPARgamma activation can drive foam cell like change in rVSMCs. Our results strongly suggest that PPARgamma expression induces CD36 expression and adipocyte differentiation gene activation in the process of atherosclerosis and might be one of the crucial events in SMC derived foam cell formation.     

New route for synthesizing poly(ethylene glycol)-acrylic acid hydrogels using γ-irradiation for drug delivery carriers

Biocompatible and pH-responsive poly(ethylene glycol) (PEG)-acrylic acid (AAc) hydrogels were prepared by new technique using γ-irradiation for controlled oral drug delivery. The gel fraction was over 80% and the equal amounts of PEG and AAc blended hydrogel had efficient insulin loading using equilibrium swelling. These hydrogels exhibited unique pH-responsive characteristics in which interpolymer complexes were formed in acidic media and dissociated in neutral or basic environments. The insulin release from the gel was significantly retarded in acidic media while rapid release occurred under neutral/basic conditions. At the high pH solution, the gels swelled rapidly and over 70% of the insulin loaded was released over a period of 10 h. Within 2 h of administration of the insulin-containing gels, significant blood glucose reduction effects were observed in diabetic rats. The blood glucose reduction lasted for up to 10 h following administration.     

Identification of a Novel Class of Membrane-Bound [NiFe]-Hydrogenases in Thermococcus onnurineus NA1 by In Silico Analysis

In silico analysis of group 4 [NiFe]-hydrogenases from a hyperthermophilic archaeon, Thermococcus onnurineus NA1, revealed a novel tripartite gene cluster consisting of dehydrogenase-hydrogenase-cation/proton antiporter subunits, which may be classified as the new subgroup 4b of [NiFe]-hydrogenases-based on sequence motifs.Hydrogenases are the key enzymes involved in the metabolism of H₂, catalyzing the following chemical reaction: 2H⁺ + 2e⁻ ↔ H₂. Hydrogenases can be classified into [NiFe]-hydrogenases, [FeFe]-hydrogenases, and [Fe]-hydrogenases, based on their distinctive functional core containing the catalytic metal center (11, 17).The genomic analysis of Thermococcus onnurineus NA1, a hyperthermophilic archaeon isolated from a deep-sea hydrothermal vent area, revealed the presence of several distinct gene clusters encoding seven [NiFe]-hydrogenases and one homolog similar to Mbx (membrane-bound oxidoreductase) from Pyrococcus furiosus (1, 6, 8, 12). According to the classification system of hydrogenases by Vignais et al. (17), three hydrogenases (one F₄₂₀-reducing and two NADP-reducing hydrogenases) belong to group 3 [NiFe]-hydrogenases, and four hydrogenases belong to group 4 [NiFe]-hydrogenases. The group 4 hydrogenases are widely distributed among bacteria and archaea (17), with Hyc and Hyf (hydrogenase 3 and 4, respectively) from Escherichia coli (19), Coo (CO-induced hydrogenase) from Rhodospirillum rubrum (4), Ech (energy-converting hydrogenase) from Methanosarcina barkeri (7), and Mbh (membrane-bound hydrogenase) from P. furiosus (6, 10, 12) being relatively well-characterized hydrogenases in this group. One of the four group 4 hydrogenases from T. onnurineus NA1 was found to be similar in sequence to that of P. furiosus Mbh (10).     

Activity-dependent α-Cleavage of Nectin-1 Is Mediated by A Disintegrin and Metalloprotease 10 (ADAM10)

Nectin-1 is known to undergo ectodomain shedding by α-secretase and subsequent proteolytic processing by γ-secretase. How secretase-mediated cleavage of nectin-1 is regulated in neuronal cells and how nectin-1 cleavage affects synaptic adhesion is poorly understood. We have investigated α-and γ-secretase-mediated processing of nectin-1 in primary cortical neurons and identified which protease acts as a α-secretase. We report here that NMDA receptor activation, but not stimulation of AMPA or metabotropic glutamate receptors, resulted in robust α- and γ-secretase cleavage of nectin-1 in mature cortical neurons. Cleavage of nectin-1 required influx of Ca²⁺ through the NMDA receptor, and activation of calmodulin, but was not dependent on calcium/calmodulin-dependent protein kinase II (CaMKII) activation. We found that ADAM10 is the major secretase responsible for nectin-1 ectodomain cleavage in neurons and the brain. These observations suggest that α- and γ-secretase processing of nectin-1 is a Ca²⁺/calmodulin-regulated event that occurs under conditions of activity-dependent synaptic plasticity and ADAM10 and γ-secretase are responsible for these cleavage events.     

Factors governing loss and rescue of DNA binding upon single and double mutations in the p53 core domain

The mutation of R273→H in the p53 core domain (p53-CD) is one of the most common mutations found in human cancers. Although the 273H p53-CD retains the wild-type conformation and stability, it lacks sequence-specific DNA binding, a transactivation function and growth suppression. However, mutating T284→R in the 273H p53-CD restores the DNA binding affinity, and transactivation and tumour suppressor functions. Since X-ray/NMR structures of DNA-free or DNA-bound mutant p53-CD molecules are unavailable, the factors governing the loss and rescue of sequence-specific DNA binding in the 273H and 273H+284R p53-CD, respectively, are unclear. Hence, we have carried out molecular dynamics (MD) simulations of the wild-type, single mutant and double mutant p53-CD, free and DNA bound, in the presence of explicit water molecules. Based on the MD structures, the DNA-binding free energy of each p53 molecule has been computed and decomposed into component energies and contributions from the interface residues. The wild-type and mutant p53-CD MD structures were found to be consistent with the antibody-binding, X-ray and NMR data. The predicted DNA binding affinity and specificity of both mutant p53-CDs were also in accord with experimental data. The non-detectable DNA binding of the 273H p53-CD is due mainly to the disruption of a hydrogen-bonding network involving R273, D281 and R280, leading to a loss of major groove binding by R280 and K120. The restoration of DNA binding affinity and specificity of the 273H+284R p53-CD is due mainly to the introduction of another DNA-binding site at position 284, leading to a recovery of major groove binding by R280 and K120. The important role of water molecules and the DNA major groove conformation as well as implications for structure-based linker rescue of the 273H p53-CD DNA-binding affinity are discussed.     

Atrial Arrhythmia in Ageing Spontaneously Hypertensive Rats: Unraveling the Substrate in Hypertension and Ageing

Background

Both ageing and hypertension are known risk factors for atrial fibrillation (AF) although the pathophysiological contribution or interaction of the individual factors remains poorly understood. Here we aim to delineate the arrhythmogenic atrial substrate in mature spontaneously hypertensive rats (SHR).

Methods

SHR were studied at 12 and 15 months of age (n = 8 per group) together with equal numbers of age-matched normotensive Wistar-Kyoto control rats (WKY). Electrophysiologic study was performed on superfused isolated right and left atrial preparations using a custom built high-density multiple-electrode array to determine effective refractory periods (ERP), atrial conduction and atrial arrhythmia inducibility. Tissue specimens were harvested for structural analysis.

Results

Compared to WKY controls, the SHR demonstrated: Higher systolic blood pressure (p<0.0001), bi-atrial enlargement (p<0.05), bi-ventricular hypertrophy (p<0.05), lower atrial ERP (p = 0.008), increased atrial conduction heterogeneity (p = 0.001) and increased atrial interstitial fibrosis (p = 0.006) & CD68-positive macrophages infiltration (p<0.0001). These changes resulted in higher atrial arrhythmia inducibility (p = 0.01) and longer induced AF episodes (p = 0.02) in 15-month old SHR. Ageing contributed to incremental bi-atrial hypertrophy (p<0.01) and atrial conduction heterogeneity (p<0.01) without affecting atrial ERP, fibrosis and arrhythmia inducibility. The limited effect of ageing on the atrial substrate may be secondary to the reduction in CD68-positive macrophages.

Conclusions

Significant atrial electrical and structural remodeling is evident in the ageing spontaneously hypertensive rat atria. Concomitant hypertension appears to play a greater pathophysiological role than ageing despite their compounding effect on the atrial substrate. Inflammation is pathophysiologically linked to the pro-fibrotic changes in the hypertensive atria.     

Enterovirus‐A71 exploits peripherin and Rac1 to invade the central nervous system

Enterovirus‐A71 (EV‐A71) has been associated with severe neurological forms of hand, foot, and mouth disease (HFMD). EV‐A71 infects motor neurons at neuromuscular junctions (NMJs) to invade the central nervous system (CNS). Here, we investigate the role of peripherin (PRPH) during EV‐A71 infection, a type III intermediate neurofilament involved in neurodegenerative conditions. In mice infected with EV‐A71, PRPH co‐localizes with viral particles in the muscles at NMJs and in the spinal cord. In motor neuron‐like and neuroblastoma cell lines, surface‐expressed PRPH facilitates viral entry, while intracellular PRPH influences viral genome replication through interactions with structural and non‐structural viral components. Importantly, PRPH does not play a role during infection with coxsackievirus A16, another causative agent of HFMD rarely associated with neurological complications, suggesting that EV‐A71 ability to exploit PRPH represents a unique attribute for successful CNS invasion. Finally, we show that EV‐A71 also exploits some of the many PRPH‐interacting partners. Of these, small GTP‐binding protein Rac1 represents a potential druggable host target to limit neuroinvasion of EV‐A71.