Effect of simvastatin on high density lipoprotein subfractions and apolipoproteins in Type IIa hypercholesterolemia

Summary Changes in plasma concentrations of high density lipoproteins (HDL) and triglycerides may partly explain the ability of cholesterol-lowering drugs to decrease the incidence of coronary heart disease. We measured the response of fasting plasma lipids, lipoproteins, and apolipoproteins in 46 subjects with Type IIa hypercholesterolemia treated with simvastatin for 3 months. The initial dose of simvastatin (10 mg/day) was subsequently increased up to 40 mg/day if the plasma cholesterol concentration had not fallen below 5.2 mmol/l. Plasma concentrations of HDL cholesterol and of the apolipoproteins AI and AII were increased by simvastatin. The increase in HDL cholesterol (9%) was due to increases in both subfractions (HDL₂ 17%; HDL₃ 7%), changes that would be consistent with a beneficial effect on cardiovascular risk. Simvastatin decreased plasma triglyceride concentrations by 25%. Plasma total cholesterol concentrations fell by 35% after 3 months of treatment; this fall was proportional to the initial concentration and was due almost entirely to a 45% fall in low density lipoprotein cholesterol. In contrast, plasma concentrations of lipoprotein Lp(a) were not affected by simvastatin.     

Low-dose levalbuterol in children with asthma: Safety and efficacy in comparison with placebo and racemic albuterol

BACKGROUND: Racemic albuterol (RAC) is an equal mixture of (R)-albuterol and (S)-albuterol. Only the (R)-isomer, levalbuterol (LEV), is therapeutically active. Lower doses of LEV, devoid of (S)-albuterol, have demonstrated efficacy comparable to that of higher doses of the (R)-isomer administered as a component of RAC. OBJECTIVE: The purpose of this study was to determine whether LEV results in improved safety and efficacy in children. METHODS: Asthmatic children aged 4 to 11 years (n = 338; FEV(1), 40% to 85% of predicted) participated in this multicenter, randomized, double-blinded study and received 21 days of 3-times-a-day treatment with nebulized LEV (0.31 or 0.63 mg), RAC (1.25 or 2.5 mg), or placebo. The primary endpoint was FEV(1) (peak percent change). Adverse events, clinical laboratory test results, vital signs, and electrocardiograms were evaluated for safety. RESULTS: All active treatments significantly improved the primary endpoint in comparison with placebo (P < .001). Significant differences in FEV(1) were noted immediately after nebulization (median change, 2.0%, 19.0%, 18.1%, 12.4%, and 15.6% for placebo, LEV 0.31 and 0.63, RAC 1.25 and 2.5 mg, respectively; P < .05 vs placebo; P < .05 for LEV 0.31 and 0.63 vs RAC 1.25 mg). LEV 0.31 mg was the only treatment not different from placebo for changes in ventricular heart rate, QT(c) interval, and glucose (P > .05). All active treatments decreased serum potassium (range, -0.3 to -0.6; P < .002 vs placebo), and RAC 2.5 mg caused the greatest change (P < .005 vs other actives). In a patient subset with severe asthma, a dose-response relationship was observed for levalbuterol, indicating that higher doses were more effective. CONCLUSION: LEV was clinically comparable to 4- to 8-fold higher doses of RAC, and it demonstrated a more favorable safety profile. LEV 0.31 mg should be used as the starting dose in 4-11 year old children with mild to moderate persistent asthma. Patients with severe disease might benefit from higher doses.     

Preexposure of murine macrophages to CpG oligonucleotide results in a biphasic tumor necrosis factor alpha response to subsequent lipopolysaccharide challenge

Bacterial DNA and synthetic oligonucleotides containing CpG sequences (CpG-DNA and CpG-ODN) provoke a proinflammatory cytokine response (tumor necrosis factor alpha [TNF-alpha], interleukin-12 [IL-12], and IL-6) and increased mortality in lipopolysaccharide (LPS)-challenged mice via a TNF-alpha-mediated mechanism. It was hypothesized that preexposure of macrophages to CpG-ODN would result in an increased TNF-alpha response to subsequent LPS challenge in vitro. Using the murine macrophage cell line RAW 264.7, we demonstrated both a rapid proinflammatory cytokine response (TNF-alpha) and a delayed inhibitory cytokine response (IL-10) with CpG-ODN. Preexposure of macrophages to CpG-ODN for brief periods (1 to 3 h) augmented TNF-alpha secretion and mRNA accumulation following subsequent LPS challenge (1 microg/ml). However, prolonged preexposure to CpG-ODN (6 to 9 h) resulted in suppression of the TNF-alpha protein and mRNA response to LPS. The addition of anti-IL-10 antibody to CpG-ODN during preexposure resulted in an increase in the LPS-induced TNF-alpha response over that induced by CpG-ODN preexposure alone. Thus, while brief preexposure of macrophages to CpG-ODN augments the proinflammatory cytokine response to subsequent LPS challenge, prolonged preexposure elicits IL-10 production, which inhibits the TNF-alpha response. Although the initial proinflammatory effects of CpG-DNA are well established, the immune response to CpG-DNA may also include autocrine or paracrine feedback mechanisms, leading to a complex interaction of proinflammatory and inhibitory cytokines.     

Effect of Metronidazole Resistance on Bacterial Eradication of Helicobacter pylori in Infected Children

A prospective study was performed with 23 Helicobacter pylori-infected children (mean age, 9.5 ± 4.4 years) with clinical symptoms of gastritis and positive results of culture and histologic examination of gastric biopsy specimens to evaluate the influence of antibiotic resistance on eradication. Positive children were treated for 4 weeks with lansoprazole and for 2 weeks with either amoxicillin-metronidazole or spiramycin (a macrolide)-metronidazole. At endoscopy 1 month after the discontinuation of therapy, the eradication rate and improvement of histologically related gastritis were significantly dependent on the susceptibility or the resistance of the infecting organism to metronidazole (83 versus 17% and 88 versus 16.6%, respectively). Pretreatment determination of the susceptibility is appropriate in any anti-H. pylori regimen, including one with metronidazole.     

Tupuseleiamides and basiliskamides,new acyldipeptides and antifungal polyketides produced in culture by a Bacillus laterosporus isolate obtained from a tropical marine habitat

Laboratory cultures of PNG 276, a Bacillus laterosporus isolate obtained from coastal waters off Papua New Guinea, have been shown to produce the novel metabolites basiliskamide A (1), basiliskamide B (2), tupuseleiamide A (3), and tupusleiamide B (4). The structures of 1 to 4 were elucidated by analysis of spectroscopic data and chemical degradation. Basiliskamides A (1) and B (2) show potent in vitro anti-Candida activity.     

Early-stage dynamics of chloride ion–pumping rhodopsin revealed by a femtosecond X-ray laser

Chloride ion–pumping rhodopsin (ClR) in some marine bacteria utilizes light energy to actively transport Cl⁻ into cells. How the ClR initiates the transport is elusive. Here, we show the dynamics of ion transport observed with time-resolved serial femtosecond (fs) crystallography using the Linac Coherent Light Source. X-ray pulses captured structural changes in ClR upon flash illumination with a 550 nm fs-pumping laser. High-resolution structures for five time points (dark to 100 ps after flashing) reveal complex and coordinated dynamics comprising retinal isomerization, water molecule rearrangement, and conformational changes of various residues. Combining data from time-resolved spectroscopy experiments and molecular dynamics simulations, this study reveals that the chloride ion close to the Schiff base undergoes a dissociation–diffusion process upon light-triggered retinal isomerization.

Chloride ion (Cl⁻) concentration in some bacterial cells is regulated by rhodopsin proteins, generally known as halorhodopsin, or hR. These proteins use light energy to pump Cl⁻ into cells (1, 2). Light is harvested by a molecule of retinal, covalently linked to an essential lysine residue in the seventh transmembrane helix of GPCR-like (G protein–coupled receptor) proteins. Light activation causes retinal to isomerize from the all-trans to the 13-cis configuration. This change triggers subsequent conformational changes throughout the rhodopsin molecule and releases chloride into the cytoplasm. Retinal thermally relaxes to the all-trans configuration within milliseconds and is then ready for the next photocycle. Cl⁻ ions are transported from the extracellular (EC) side to the cytoplasmic (CP) side during each photocycle (3, 4).Light-driven ion-pumping rhodopsin can be used to develop artificial solar energy harvesting and optogenetics (5–8), but the molecular mechanism must be understood in detail for such applications. Despite the importance of hR, our current experimental data concerning the structure and dynamics of the protein remain very limited. A related protein, proton (H⁺)-pumping bacteriorhodopsin (bR) discovered in the early 1970s, has been extensively studied by multiple methods, including time-resolved spectroscopy, crystallography, mutagenesis, and computer simulation (9–12). In particular, recent studies using time-resolved serial femtosecond crystallography (TR-SFX) methods performed at X-ray free-electron laser (XFEL) facilities allow three-dimensional (3D) visualization of retinal isomerization and associated local conformational changes. These changes are accompanied by movement of protons from a donor aspartate group to an acceptor aspartate (13–15). However, the central component of this process, the transported H⁺, is difficult to observe by X-ray crystallography and could not be directly traced in bR TR-SFX studies. Recently, a breakthrough was reported in a study on the sodium-pumping rhodopsin KR2 (K. eikastus rhodopsin 2), in which electron density signals of Na⁺ uptake were observed at Δt = 1 ms after laser illumination (16).Cl⁻, a strong X-ray scatterer, can be directly observed from electron density maps. These maps provide first-hand information on the movement of ions as being transported within short timescales after light activation. Furthermore, hR and bR presumably share a common molecular mechanism despite transporting ions in opposite directions. A close relationship is strongly implied by the interconversion of the function of two rhodopsins. Outward H⁺-pumping bR can be converted to an inward Cl⁻ pump by changing a single residue (D85T) (17), while hR from the cyanobacterium, Mastigocladopsis repens, is reported to pump protons after a single mutation (T74D) (18). The chloride pump can therefore serve as a system analogous to the proton transporter and provide valuable information that is difficult to obtain directly from bR.In this study, we focus on chloride ion–pumping rhodopsin (ClR) from the marine flavobacterium Nonlabens marinus S1-08T (19). The conserved DTD motif (Asp85-Thr89-Asp96) of the bR family, residues 85, 89, and 96, is replaced by an NTQ motif (Asn98- Thr102-Gln109) in ClR (Fig. 1). The sequence identity of ClR and canonical bR from Halobacterium salinarum is only 27%, but the two proteins, nevertheless, have highly similar structures, including the disposition of the retinal chromophore. ClR structures at cryogenic and room temperatures clearly reveal an architecture composed of seven transmembrane helices (TM A to G) (2, 20, 21). The retinal is covalently linked to the Nζ atom of the Lys235 located on TM-G. Anomalous diffraction signals of the Br⁻ identify a stable binding site near the protonated Schiff base (PSB) and a plausible exit site on the CP side (Fig. 1A). Buried water molecules and locations of cavities inside ClR suggest a pathway for Cl⁻ uptake on the EC side, but the molecular mechanism for light-triggered Cl⁻ pumping remains obscure. Upon light activation, the Cl⁻ tightly held near the PSB must break free from its hydrogen bonding network (Fig. 1B). It then passes through a hydrophobic region to reach the CP side (Fig. 1C). Crystal structures of ClR were previously determined with crystals under continuous illumination of visible laser light. Intriguingly, these steady-state models revealed unexpected movement of the retinal, without indication of photo-isomerization (22). Steady-state measurements, which show averages of mixed states, are thus of limited use in deciphering the molecular mechanism of light-driven Cl⁻ pumping.Open in a separate windowFig. 1.Structure of ClR and a plausible pathway of Cl⁻ transport. (A) Cross-sections of ClR with the backbone structure shown in cartoon representation. Transmembrane helices are marked using letters A through G, and the C-terminal helix H in the cytoplasm is also indicated. Surfaces are clipped to show the cross-section colored in yellow and the model being sliced and then opened about the axis near the helix E. Water molecules and Cl⁻ ions are shown as red- and green-colored spheres. Blue curves indicate the path of ion entering ClR and the principal pumping direction after passing retinal. (B) Key residues near the Cl⁻ ion and retinal, together with the NTQ motif shown in stick representation. (C) Residues that form a hydrophobic region between the retinal and the cytoplasm are highlighted in ball-and-stick representation. The red arrow points to a major barrier that Cl⁻ needs to overcome. ClR backbone is shown in cartoon representation, with residues colored based on hydrophobicity (the blue to red spectrum corresponds to the hydrophobicity scale from hydrophilic to hydrophobic).