Apolipoprotein A-I Attenuates Palmitate-Mediated NF-κB Activation by Reducing Toll-Like Receptor-4 Recruitment into Lipid Rafts

While high-density lipoprotein (HDL) is known to protect against a wide range of inflammatory stimuli, its anti-inflammatory mechanisms are not well understood. Furthermore, HDL's protective effects against saturated dietary fats have not been previously described. In this study, we used endothelial cells to demonstrate that while palmitic acid activates NF-κB signaling, apolipoprotein A–I, (apoA-I), the major protein component of HDL, attenuates palmitate-induced NF-κB activation. Further, vascular NF-κB signaling (IL-6, MCP-1, TNF-α) and macrophage markers (CD68, CD11c) induced by 24 weeks of a diabetogenic diet containing cholesterol (DDC) is reduced in human apoA-I overexpressing transgenic C57BL/6 mice compared to age-matched WT controls. Moreover, WT mice on DDC compared to a chow diet display increased gene expression of lipid raft markers such as Caveolin-1 and Flotillin-1, and inflammatory Toll-like receptors (TLRs) (TLR2, TLR4) in the vasculature. However apoA-I transgenic mice on DDC show markedly reduced expression of these genes. Finally, we show that in endothelial cells TLR4 is recruited into lipid rafts in response to palmitate, and that apoA-I prevents palmitate-induced TLR4 trafficking into lipid rafts, thereby blocking NF-κB activation. Thus, apoA-I overexpression might be a useful therapeutic tool against vascular inflammation

Two new steroidal tetraglycosides from the rhizomes of Smilacina atropurpurea

Smilacina atropurpurea (family Convallariaceae) is a perennial herb used medicinally for the treatment of lung ailments, rheumatism, menstrual disorders, and cuts and bruises, by the local people of its growing areas in southwest China. The tender aerial part has also been used as a wild vegetable by Lisu, Naxi and Tibetan people. Chemical analysis of the rhizomes of this plant has allowed us to isolate two new minor steroidal tetraglycosides, specifically atropurosides H (1) and I (2), by repeated column chromatography over silica gel and reversed phase silica. The structures were determined to be (25R,S)-12β-hydroxy-spirost-5-ene-3β-yl-O-β-D-glucopyranosyl-(1→2)-[β-D-xylopyranosyl-(1→3)]-β-D-glucopyranosyl-(1→4)-β-D-galactopyranoside (1) and (25S)-5α-spirost-17α-hydroxy-3β-yl-O-β-D-glucopyranosyl-(1→2)-[β-D-xylopyranosyl-(1→3)]-β-D-glucopyranosyl-(1→4)-β-D-galacto pyranoside (2), respectively, on the basis of detailed spectroscopic analysis including 1D and 2D NMR techniques

Three amino acid substitutions contributing to thermostability of phosphoglucose isomerase in the Glanville fritillary butterfly

Abstract Temperature is one of the most important environmental factors that affect organisms, especially ectotherms, due to its effects on protein stability. Understanding the general rules that govern thermostability changes in proteins to adapt high-temperature environments is crucial. Here, we report the amino acid substitutions of phosphoglucose isomerase (PGI) related to thermostability in the Glanville fritillary butterfly (Melitaea cinxia, Lepidoptera: Nymphalidae). The PGI encoded by the most common allele in M. cinxia in the Chinese population (G3-PGI), which is more thermal-tolerant, is more stable under heat stress than that in the Finnish population (D1-PGI). There are five amino acid substitutions between G3-PGI and D1-PGI. Site-directed mutagenesis revealed that the combination of amino acid substitutions of H35Q, M49T and I64V may increase PGI thermostability. These substitutions alter the 3D structure to increase the interaction between two monomers of PGI. Through molecular dynamics simulations, it was found that the amino acid at site 421 is more stable in G3-PGI, confining the motion of the α-helix 420?441 and stabilizing the interaction between two PGI monomers. The strategy for high-temperature adaptation through these three amino acid substitutions is also adopted by other butterfly species (Boloria eunomia, Aglais urticae, Colias erate and Polycaena lua) concurrent with M. cinxia in the Tianshan Mountains of China, i.e., convergent evolution in butterflies. This article is protected by copyright. All rights reservedPeer reviewe

Suppression of Rap1 Impairs Cardiac Myofibrils and Conduction System in Zebrafish

<div><p>Numerous studies have revealed that Rap1 (Ras-proximate-1 or Ras-related protein 1), a small GTPase protein, plays a crucial role in mediating cAMP signaling in isolated cardiac tissues and cell lines. However, the involvement of Rap1 in the cardiac development <em>in vivo</em> is largely unknown. By injecting anti-sense morpholino oligonucleotides to knock down Rap1a and Rap1b in zebrafish embryos, and in combination with time-lapsed imaging, <em>in situ</em> hybridization, immunohistochemistry and transmission electron microscope techniques, we seek to understand the role of Rap1 in cardiac development and functions. At an optimized low dose of mixed <em>rap1a</em> and <em>rap1b</em> morpholino oligonucleotides, the heart developed essentially normally until cardiac contraction occurred. Morphant hearts showed the myocardium defect phenotypes, most likely due to disrupted myofibril assembly and alignment. <em>In vivo</em> heart electrocardiography revealed prolonged P-R interval and QRS duration, consistent with an adherens junction defect and reduced Connexons in cardiac myocytes of morphants. We conclude that a proper level of Rap1 is crucial for heart morphogenesis and function, and suggest that Rap1 and/or their downstream factor genes are potential candidates for genetic screening for human heart diseases.</p> </div

Phenotype summary of the <i>rap1</i>MO fish.

<p>Phenotype summary of the <i>rap1</i>MO fish.</p

Rap1 knock-down zebrafish have prolonged P-R intervals, resembling the first-degree atrio-ventricular block.

<p>Zebrafish heart electrocardiography (ECG) was measured in wild type larvae (A), and <i>rap1</i>MO hearts at 3 dpf (B to D).</p

Ultra-structural changes of cell junctions and sarcomeres in Rap1 knock-down zebrafish heart.

<p>Transmission electronic scan revealed that the myofibril units, regularly organized into hexagonal lattices with thick (green arrow) and thin (yellow arrows) filaments (A) with adherens junctions (AJs) along the membrane of adjacent cardiac myocytes (B). In <i>rap1</i>MO heart, thick filaments (green arrow) number was not markablely changed but thin filaments (yellow arrows) number was significantly and statistically reduced (A' and D), and less myofibrils attached to AJs (arrows in B and B'), with lager space between the membrane of adjacent cardiac myocytes (B'). Antibody against Connexin 43 stained a significantly and statistically less GJ signal (arrowheads in C and C') in <i>rap1</i>MO cardiomyocytes at 3 dpf, compared to a normal heart (C, C', and E). Scale bar, 50 nm in A and A'; 350 nm in B and B'; 50 µm in C and C'. Columns and error bars in D and E showed mean±S.D. n = 9 in filaments experiment (D) for each group, and n = 10 in C×43 antibody staining experiment (E) for each group. Unpaired two-tailed t-test was used to test the significance between two columns in each group in D. *** in statistic graph represent p<0.001 in the t-test.</p