Antiviral signalling by a cyclic nucleotide activated CRISPR protease.

CRISPR defence systems such as the well-known DNA-targeting Cas9 and the RNA-targeting type III systems are widespread in prokaryotes1,2. The latter orchestrates a complex antiviral response that is initiated through the synthesis of cyclic oligoadenylates after recognition of foreign RNA3-5. Among the large set of proteins that are linked to type III systems and predicted to bind cyclic oligoadenylates6,7, a CRISPR-associated Lon protease (CalpL) stood out to us. CalpL contains a sensor domain of the SAVED family7 fused to a Lon protease effector domain. However, the mode of action of this effector is unknown. Here we report the structure and function of CalpL and show that this soluble protein forms a stable tripartite complex with two other proteins, CalpT and CalpS, that are encoded on the same operon. After activation by cyclic tetra-adenylate (cA4), CalpL oligomerizes and specifically cleaves the MazF homologue CalpT, which releases the extracytoplasmic function σ factor CalpS from the complex. Our data provide a direct connection between CRISPR-based detection of foreign nucleic acids and transcriptional regulation. Furthermore, the presence of a SAVED domain that binds cyclic tetra-adenylate in a CRISPR effector reveals a link to the cyclic-oligonucleotide-based antiphage signalling system.

Absolute proteomic quantification reveals design principles of sperm flagellar chemosensation.

Cilia serve as cellular antennae that translate sensory information into physiological responses. In the sperm flagellum, a single chemoattractant molecule can trigger a Ca2+ rise that controls motility. The mechanisms underlying such ultra-sensitivity are ill-defined. Here, we determine by mass spectrometry the copy number of nineteen chemosensory signaling proteins in sperm flagella from the sea urchin Arbacia punctulata. Proteins are up to 1,000-fold more abundant than the free cellular messengers cAMP, cGMP, H+ , and Ca2+ . Opto-chemical techniques show that high protein concentrations kinetically compartmentalize the flagellum: Within milliseconds, cGMP is relayed from the receptor guanylate cyclase to a cGMP-gated channel that serves as a perfect chemo-electrical transducer. cGMP is rapidly hydrolyzed, possibly via "substrate channeling" from the channel to the phosphodiesterase PDE5. The channel/PDE5 tandem encodes cGMP turnover rates rather than concentrations. The rate-detection mechanism allows continuous stimulus sampling over a wide dynamic range. The textbook notion of signal amplification-few enzyme molecules process many messenger molecules-does not hold for sperm flagella. Instead, high protein concentrations ascertain messenger detection. Similar mechanisms may occur in other small compartments like primary cilia or dendritic spines.

A family of hyperpolarization-activated channels selective for protons.

Proton (H+) channels are special: They select protons against other ions that are up to a millionfold more abundant. Only a few proton channels have been identified so far. Here, we identify a family of voltage-gated "pacemaker" channels, HCNL1, that are exquisitely selective for protons. HCNL1 activates during hyperpolarization and conducts protons into the cytosol. Surprisingly, protons permeate through the channel's voltage-sensing domain, whereas the pore domain is nonfunctional. Key to proton permeation is a methionine residue that interrupts the series of regularly spaced arginine residues in the S4 voltage sensor. HCNL1 forms a tetramer and thus contains four proton pores. Unlike classic HCN channels, HCNL1 is not gated by cyclic nucleotides. The channel is present in zebrafish sperm and carries a proton inward current that acidifies the cytosol. Our results suggest that protons rather than cyclic nucleotides serve as cellular messengers in zebrafish sperm. Through small modifications in two key functional domains, HCNL1 evolutionarily adapted to a low-Na+ freshwater environment to conserve sperm's ability to depolarize.

Spatiotemporal Resolution of Conformational Changes in Biomolecules by Combining Pulsed Electron-Electron Double Resonance Spectroscopy with Microsecond Freeze-Hyperquenching.

The function of proteins is linked to their conformations that can be resolved with several high-resolution methods. However, only a few methods can provide the temporal order of intermediates and conformational changes, with each having its limitations. Here, we combine pulsed electron-electron double resonance spectroscopy with a microsecond freeze-hyperquenching setup to achieve spatiotemporal resolution in the angstrom range and lower microsecond time scale. We show that the conformational change of the Cα-helix in the cyclic nucleotide-binding domain of the Mesorhizobium loti potassium channel occurs within about 150 µs and can be resolved with angstrom precision. Thus, this approach holds great promise for obtaining 4D landscapes of conformational changes in biomolecules.

The CatSper channel controls chemosensation in sea urchin sperm.

Sperm guidance is controlled by chemical and physical cues. In many species, Ca(2+) bursts in the flagellum govern navigation to the egg. In Arbacia punctulata, a model system of sperm chemotaxis, a cGMP signaling pathway controls these Ca(2+) bursts. The underlying Ca(2+) channel and its mechanisms of activation are unknown. Here, we identify CatSper Ca(2+) channels in the flagellum of A. punctulata sperm. We show that CatSper mediates the chemoattractant-evoked Ca(2+) influx and controls chemotactic steering; a concomitant alkalization serves as a highly cooperative mechanism that enables CatSper to transduce periodic voltage changes into Ca(2+) bursts. Our results reveal intriguing phylogenetic commonalities but also variations between marine invertebrates and mammals regarding the function and control of CatSper. The variations probably reflect functional and mechanistic adaptations that evolved during the transition from external to internal fertilization.

The CatSper channel mediates progesterone-induced Ca2+ influx in human sperm.

In the oviduct, cumulus cells that surround the oocyte release progesterone. In human sperm, progesterone stimulates a Ca(2+) increase by a non-genomic mechanism. The Ca(2+) signal has been proposed to control chemotaxis, hyperactivation and acrosomal exocytosis of sperm. However, the underlying signalling mechanism has remained mysterious. Here we show that progesterone activates the sperm-specific, pH-sensitive CatSper Ca(2+) channel. We found that both progesterone and alkaline pH stimulate a rapid Ca(2+) influx with almost no latency, incompatible with a signalling pathway involving metabotropic receptors and second messengers. The Ca(2+) signals evoked by alkaline pH and progesterone are inhibited by the Ca(v) channel blockers NNC 55-0396 and mibefradil. Patch-clamp recordings from sperm reveal an alkaline-activated current carried by mono- and divalent ions that exhibits all the hallmarks of sperm-specific CatSper Ca(2+) channels. Progesterone substantially enhances the CatSper current. The alkaline- and progesterone-activated CatSper current is inhibited by both drugs. Our results resolve a long-standing controversy over the non-genomic progesterone signalling. In human sperm, either the CatSper channel itself or an associated protein serves as the non-genomic progesterone receptor. The identification of CatSper channel blockers will greatly facilitate the study of Ca(2+) signalling in sperm and help to define further the physiological role of progesterone and CatSper.

A Quantitative Model for cAMP Binding to the Binding Domain of MloK1.

Ligand-protein binding processes are essential in biological systems. A well-studied system is the binding of cyclic adenosine monophosphate to the cyclic nucleotide binding domain of the bacterial potassium channel MloK1. Strikingly, the measured on-rate for cyclic adenosine monophosphate binding is two orders of magnitude slower than a simple Smoluchowski diffusion model would suggest. To resolve this discrepancy and to characterize the ligand-binding path in structural and energetic terms, we calculated 1100 ligand-binding molecular dynamics trajectories and tested two scenarios: In the first scenario, the ligand transiently binds to the protein surface and then diffuses along the surface into the binding site. In the second scenario, only ligands that reach the protein surface in the vicinity of the binding site proceed into the binding site. Here, a binding funnel, which increasingly confines the translational as well as the rotational degrees of freedom, determines the binding pathways and limits the on-rate. From the simulations, we identified five surface binding states and calculated the rates between these surface binding states, the binding site, and the bulk. We find that the transient binding of the ligands to the surface binding states does not affect the on-rate, such that this effect alone cannot explain the observed low on-rate. Rather, by quantifying the translational and rotational degrees of freedom and by calculating the binding committor, our simulations confirmed the existence of a binding funnel as the main bottleneck. Direct binding via the binding funnel dominates the binding kinetics, and only ∼10% of all ligands proceed via the surface into the binding site. The simulations further predict an on-rate between 15 and 40µs-1(mol/l)-1, which agrees with the measured on-rate.

Prospective Associations of Systemic and Urinary Choline Metabolites with Incident Type 2 Diabetes.

BACKGROUND: Several compounds in the choline oxidation pathway are associated with insulin resistance and prevalent diabetes; however, prospective data are scarce.We explored the relationships between systemic and urinary choline-related metabolites and incident type 2 diabetes in an observational prospective study among Norwegian patients. METHODS: We explored risk associations by logistic regression among 3621 nondiabetic individuals with suspected stable angina pectoris, of whom 3242 provided urine samples. Reclassification of patients was investigated according to continuous net reclassification improvement (NRI >0). RESULTS: After median (25th to 75th percentile) follow-up of 7.5 (6.4-8.7) years, 233 patients (6.4%) were registered with incident type 2 diabetes. In models adjusted for age, sex, and fasting status, plasma betaine was inversely related to new-onset disease [odds ratio (OR) per 1 SD, 0.72; 95% CI, 0.62-0.83; P < 0.00001], whereas positive associations were observed for urine betaine (1.25; 1.09-1.43; P = 0.001), dimethylglycine (1.22; 1.06-1.40; P = 0.007), and sarcosine (1.30; 1.13-1.49; P < 0.001). The associations were maintained in a multivariable model adjusting for body mass index, hemoglobin A1c, urine albumin-to-creatinine ratio, estimated glomerular filtration rate, C-reactive protein, HDL cholesterol, and medications. Plasma betaine and urine sarcosine, the indices most strongly related to incident type 2 diabetes, improved reclassification [NRI >0 (95% CI) 0.33 (0.19-0.47) and 0.16 (0.01-0.31), respectively] and showed good within-person reproducibility. CONCLUSIONS: Systemic and urinary concentrations of several choline metabolites were associated with risk of incident type 2 diabetes, and relevant biomarkers may improve risk prediction.

Serum trans fatty acids, asymmetric dimethylarginine and risk of acute myocardial infarction and mortality in patients with suspected coronary heart disease: a prospective cohort study.

BACKGROUND: Trans fatty acids (TFAs) have been found to impair flow mediated vasodilation and nitric oxide (NO) production. We sought to examine if serum TFA levels are associated with plasma levels of the NO inhibitor asymmetric dimethylarginine (ADMA) and if possible relationships between serum TFA and cardiovascular morbidity or mortality are mediated or modified by plasma ADMA levels. METHODS: The cohort included patients who underwent coronary angiography for suspected coronary heart disease in 2000-2001. Serum trans 16:1n7 and trans 18:1 isomers were determined by gas liquid chromatography and the summation of these two TFAs is reported as TFA (percentage by weight (wt%) or concentration). Associations between TFAs and ADMA were estimated by calculating the Spearman's rank correlation coefficient (ρ), and risk associations with AMI, cardiovascular death and all-cause mortality across quartiles of TFAs (wt% or concentration) were explored by Cox modeling. RESULTS: A total of 1364 patients (75 % men) with median (25(th),75(th) percentile) age 61 (54, 69) years, serum TFA 0.46 (0.36, 0.56) wt% and plasma ADMA 0.59 (0.50, 0.70) µmol/L were studied. Serum TFA levels (ρ = 0.21, p < 0.001), trans 16:1n7 (ρ = 0.22, p < 0.001) and trans 18:1 (ρ = 0.20, p < 0.001) levels were significantly correlated with plasma ADMA levels. During the median (25(th),75(th) percentile) follow-up time of 5.8 (4.5, 6.4) years, 129 (9.5 %) patients experienced an AMI, 124 (9.1 %) died, whereof 66 (53 %) due to cardiovascular causes. After multivariate adjustments no significant associations between serum TFA levels (wt% or concentration) and incident AMI, CV death and all-cause mortality were observed. Similar results were obtained when repeating the analyses with trans 16:1n7 and trans 18:1 individually. Plasma ADMA levels did not significantly modify the associations between TFA levels and outcomes. CONCLUSIONS: Serum TFA levels were positively correlated with plasma ADMA levels. After multivariate adjustments, TFAs were not associated with incident AMI or mortality, and associations were not influenced by ADMA. TRIAL REGISTRATION: Clinicaltrials.gov Identifier: NCT00354081.

Regulation of Son of sevenless by the membrane-actin linker protein ezrin.

Receptor tyrosine kinases participate in several signaling pathways through small G proteins such as Ras (rat sarcoma). An important component in the activation of these G proteins is Son of sevenless (SOS), which catalyzes the nucleotide exchange on Ras. For optimal activity, a second Ras molecule acts as an allosteric activator by binding to a second Ras-binding site within SOS. This allosteric Ras-binding site is blocked by autoinhibitory domains of SOS. We have reported recently that Ras activation also requires the actin-binding proteins ezrin, radixin, and moesin. Here we report the mechanism by which ezrin modulates SOS activity and thereby Ras activation. Active ezrin enhances Ras/MAPK signaling and interacts with both SOS and Ras in vivo and in vitro. Moreover, in vitro kinetic assays with recombinant proteins show that ezrin also is important for the activity of SOS itself. Ezrin interacts with GDP-Ras and with the Dbl homology (DH)/pleckstrin homology (PH) domains of SOS, bringing GDP-Ras to the proximity of the allosteric site of SOS. These actions of ezrin are antagonized by the neurofibromatosis type 2 tumor-suppressor protein merlin. We propose an additional essential step in SOS/Ras control that is relevant for human cancer as well as all physiological processes involving Ras.

CRIS-a novel cAMP-binding protein controlling spermiogenesis and the development of flagellar bending.

The second messengers cAMP and cGMP activate their target proteins by binding to a conserved cyclic nucleotide-binding domain (CNBD). Here, we identify and characterize an entirely novel CNBD-containing protein called CRIS (cyclic nucleotide receptor involved in sperm function) that is unrelated to any of the other members of this protein family. CRIS is exclusively expressed in sperm precursor cells. Cris-deficient male mice are either infertile due to a lack of sperm resulting from spermatogenic arrest, or subfertile due to impaired sperm motility. The motility defect is caused by altered Ca(2+) regulation of flagellar beat asymmetry, leading to a beating pattern that is reminiscent of sperm hyperactivation. Our results suggest that CRIS interacts during spermiogenesis with Ca(2+)-regulated proteins that--in mature sperm--are involved in flagellar bending.

Cooperative and uncooperative cyclic-nucleotide-gated ion channels.

Ion channels gated by cyclic nucleotides serve multiple functions in sensory signaling in diverse cell types ranging from neurons to sperm. Newly discovered members from bacteria and marine invertebrates provide a wealth of structural and functional information on this channel family. A hallmark of classical tetrameric cyclic-nucleotide-gated channels is their cooperative activation by binding of several ligands. By contrast, the new members seem to be uncooperative, and binding of a single ligand molecule suffices to open these channels. These new findings provide a fresh look at the mechanism of allosteric activation of ion channels.

Prognostic value of energy loss index in asymptomatic aortic stenosis.

BACKGROUND: Aortic valve area index adjusted for pressure recovery (energy loss index [ELI]) has been suggested as a more accurate measure of aortic stenosis (AS) severity, but its prognostic value has not been determined in a prospective study. METHODS AND RESULTS: The relation between baseline ELI and rate of aortic valve events and combined total mortality and hospitalization for heart failure resulting from the progression of AS was assessed by multivariate Cox regression and reclassification analysis in 1563 patients with initial asymptomatic AS in the Simvastatin and Ezetimibe in Aortic Stenosis (SEAS) study. During 4.3 years follow-up, a total of 498 aortic valve events and 181 combined total mortalities and hospitalizations for heart failure caused by the progression of AS occurred. In Cox regression analyses, 1-cm(2)/m(2) lower baseline ELI predicted a 2-fold higher risk both for aortic valve events and for combined total mortality and hospitalization for heart failure independently of baseline peak aortic jet velocity or mean aortic gradient and independently of aortic root size (all P<0.05). In reclassification analysis, ELI improved the prediction of aortic valve events by 13% (95% confidence interval, 5-19), whereas the prediction of combined total mortality and hospitalization for heart failure resulting from the progression of AS did not improve significantly. CONCLUSIONS: In asymptomatic AS patients without known atherosclerotic disease or diabetes mellitus, ELI provides independent and additional prognostic information to that derived from conventional measures of AS severity, suggesting that ELI should be measured in such patients. CLINICAL TRIAL REGISTRATION INFORMATION: URL: http://www.clinicaltrials.gov. Unique identifier: NCT00092677.

The response of agitated behavior to pain management in persons with dementia.

OBJECTIVES: Behavioral disturbances and pain are common in nursing home (NH) patients with dementia. An association between pain and increased agitation has been suggested, and recently a significant reduction of agitation has been demonstrated by pain treatment in patients with moderate to severe dementia. We now examined which specific agitated behaviors respond to individualized pain treatment. DESIGN: Cluster randomized clinical trial. SETTING: 60 clusters (i.e., clusters defined as single independent NH units) in 18 NHs within five municipalities of Western Norway. PARTICIPANTS: 352 patients with moderate to severe dementia and clinically significant behavioral disturbances. INTERVENTION: The control group received usual treatment and care. According to a predefined scheme for 8 weeks, all patients in the intervention group received individual daily pain treatment with acetaminophen, extended release morphine, buprenorphine transdermal patch, and/or pregabaline. MEASUREMENTS: Cohen-Mansfield Agitation Inventory subscales and items. RESULTS: Analyses demonstrated that Factor 3 (Verbally agitated behaviors) showed the largest significant difference (DF = 1204.0, t = -4.308, p <0.001), followed by Factor 2 (Physically non-aggressive behaviors) (DF = 1198.0, t = -2.672, p = 0.008), and Factor 1 (Aggressive behaviors) (DF = 1196.0, t = -2.093, p = 0.037) after 8 weeks, by a linear random intercept mixed model in two-way repeated-measures configuration with adjustment for heteroscedasticity. CONCLUSION: We found that verbal agitation behaviors such as complaining, negativism, repetitious sentences and questions, constant request for attention, and cursing or verbal aggression responded to pain treatment. In addition, restlessness and pacing were sensible to analgesics. Such behaviors should therefore lead to an assessment of pain, and pain treatment. Further studies comparing how pain treatment should be balanced against other strategies including psychotropic drugs are needed.

Plasma dimethylglycine and risk of incident acute myocardial infarction in patients with stable angina pectoris.

OBJECTIVE: Dimethylglycine is linked to lipid metabolism, and increased plasma levels may be associated with adverse prognosis in patients with coronary artery disease. We evaluated the relationship between plasma dimethylglycine and risk of incident acute myocardial infarction in a large prospective cohort of patients with stable angina pectoris, of whom approximately two thirds were participants in a B-vitamin intervention trial. Model discrimination and reclassification when adding plasma dimethylglycine to established risk factors were obtained. We also explored temporal changes and the test-retest reliability of plasma dimethylglycine. APPROACH AND RESULTS: Four thousand one hundred fifty patients (72% men; median age 62 years) were included. Plasma dimethylglycine was associated with several traditional coronary artery disease risk factors. During a median follow-up of 4.6 years, 343 (8.3%) patients experienced an acute myocardial infarction. The hazard ratio (95% confidence interval) for acute myocardial infarction was 1.95 (1.42-2.68; P<0.001) when comparing plasma dimethylglycine quartile 4 to 1 in a Cox regression model adjusted for age, sex, and fasting status. Adjusting for traditional coronary artery disease risk factors only slightly modified the estimates, which were particularly strong among nonsmokers and among patients with serum triglyceride or apolipoprotein B100 levels ≤ median (P for interaction=0.004, 0.004, and 0.03, respectively). Plasma dimethylglycine improved discrimination and reclassification and had high test-retest reliability. CONCLUSIONS: Plasma dimethylglycine is independently related to incident acute myocardial infarction and enhances risk prediction in patients with stable angina pectoris. Our results motivate further studies on the relationship between 1-carbon metabolism and atherothrombosis. A potential interplay with lipid and energy metabolism merits particular attention.

Association of body mass index with risk of acute myocardial infarction and mortality in Norwegian male and female patients with suspected stable angina pectoris: a prospective cohort study.

BACKGROUND: A number of previous studies have suggested that overweight or obese patients with coronary artery disease (CAD) may have lower morbidity and mortality than their leaner counterparts. Few studies have addressed possible gender differences, and the results are conflicting. We examined the association between body mass index (BMI) and risk of acute myocardial infarction (AMI), cardiovascular (CV) death and all-cause mortality in men and women with suspected stable angina pectoris. METHOD: The cohort included 4164 patients with suspected stable angina undergoing elective coronary angiography between 2000 and 2004. Events were registered until the end of 2006. Hazard ratios (HR) (95% confidence intervals) were estimated using Cox regression by comparing normal weight (18.5-24.9 kg/m2) with overweight (25-29.9 kg/m2) and obese (≥30 kg/m2) patients. Underweight (<18.5 kg/m2) patients were excluded from the study. RESULTS: Of 4131 patients with complete data, 72% were males and 75% were diagnosed with significant CAD. The mean (standard deviation (SD)) age in the total population was 62 (10) years. Mean (SD) BMI was 26.8 (3.9) kg/m2, 34% was normal weight, 48% overweight and 19% obese. During follow up, a total of 337 (8.2%) experienced an AMI and 302 (7.3%) patients died, of whom 165 (4.0%) died from cardiovascular causes. We observed a significant interaction between BMI groups and gender with regards to risk of AMI (p = 0.011) and CV death (p = 0.031), but not to risk of all-cause mortality; obese men had a multivariate adjusted increased risk of AMI (HR 1.80 (1.28, 2.52)) and CV death (HR 1.60 (1.00, 2.55)) compared to normal weight men. By contrast, overweight women had a decreased risk of AMI (HR 0.56 (0.33, 0.98)) compared to normal weight women. The risk of all-cause mortality did not differ between BMI categories. CONCLUSION: Compared with normal weight subjects, obese men had an increased risk of AMI and CV death, while overweight women had a decreased risk of AMI. These findings may potentially explain some of the result variation in previous studies reporting on the obesity paradox.

Urinary excretion of kynurenine and tryptophan, cardiovascular events, and mortality after elective coronary angiography.

AIMS: Kynurenine is a potent endothelium-derived vasodilator. Its synthesis from tryptophan is stimulated by interferon γ and may represent an important compensatory pathway for the regulation of vascular function in inflammatory conditions. We assessed associations of urine kynurenine to tryptophan ratio (KTR) levels to incident major coronary events (MCEs), acute myocardial infarction (AMI), and ischaemic stroke and mortality in patients with suspected stable coronary artery disease (CAD). METHODS AND RESULTS: A total of 3224 patients (mean age 62 years, 69% men) underwent urine and blood sampling prior to elective coronary angiography and were subsequently followed up for median 55 months. A total of 8.4% experienced an MCE, 7.8% suffered an AMI, and 7.6% died. In age- and gender-adjusted analyses, the hazard ratios [HRs; 95% confidence intervals (CI)] of MCE, AMI, and all-cause mortality were 1.43 (1.29-1.59), 1.44 (1.29-1.59), and 1.38 (1.23-1.54) per standard deviation increment of the (log-transformed) urinary KTR, respectively. These estimates were only minimally attenuated after adjustment for potential confounders. The addition of the urine KTR to a model of conventional risk factors significantly improved goodness of fit, discrimination, and risk classification for these clinical endpoints. No association was seen between the urine KTR and the risk of incident ischaemic stroke. CONCLUSION: A novel urinary inflammation marker, KTR, is strongly associated with adverse prognosis in patients with suspected stable CAD. Underlying pathomechanisms should be further elucidated.

Kinetics of ligand-receptor interaction reveals an induced-fit mode of binding in a cyclic nucleotide-activated protein.

Many receptors and ion channels are activated by ligands. One key question concerns the binding mechanism. Does the ligand induce conformational changes in the protein via the induced-fit mechanism? Or does the protein preexist as an ensemble of conformers and the ligand selects the most complementary one, via the conformational selection mechanism? Here, we study ligand binding of a tetrameric cyclic nucleotide-gated channel from Mesorhizobium loti and of its monomeric binding domain (CNBD) using rapid mixing, mutagenesis, and structure-based computational biology. Association rate constants of ∼10(7) M(-1) s(-1) are compatible with diffusion-limited binding. Ligand binding to the full-length CNG channel and the isolated CNBD differ, revealing allosteric control of the CNBD by the effector domain. Finally, mutagenesis of allosteric residues affects only the dissociation rate constant, suggesting that binding follows the induced-fit mechanism. This study illustrates the strength of combining mutational, kinetic, and computational approaches to unravel important mechanistic features of ligand binding.

Dietary intake of n-3 long-chain polyunsaturated fatty acids and risk of myocardial infarction in coronary artery disease patients with or without diabetes mellitus: a prospective cohort study.

BACKGROUND: A beneficial effect of a high n-3 long-chain polyunsaturated fatty acid (LCPUFA) intake has been observed in heart failure patients, who are frequently insulin resistant. We investigated the potential influence of impaired glucose metabolism on the relation between dietary intake of n-3 LCPUFAs and risk of acute myocardial infarction (AMI) in patients with coronary artery disease. METHODS: This prospective cohort study was based on the Western Norway B-Vitamin Intervention Trial and included 2,378 patients with coronary artery disease with available baseline glycosylated hemoglobin (HbA1c) and dietary data. Patients were sub-grouped as having no diabetes (HbA1c <5.7%), pre-diabetes (HbA1c ≥5.7%), or diabetes (previous diabetes, fasting baseline serum glucose ≥7.0, or non-fasting glucose ≥11.1 mmol/L). AMI risk was evaluated by Cox regression (age and sex adjusted), comparing the upper versus lower tertile of daily dietary n-3 LCPUFA intake. RESULTS: The participants (80% males) had a mean age of 62 and follow-up of 4.8 years. A high n-3 LCPUFA intake was associated with reduced risk of AMI (hazard ratio 0.38, 95%CI 0.18, 0.80) in diabetes patients (median HbA1c = 7.2%), whereas no association was observed in pre-diabetes patients. In patients without diabetes a high intake tended to be associated with an increased risk (hazard ratio1.45, 95%CI 0.84, 2.53), which was significant for fatal AMI (hazard ratio 4.79, 95%CI 1.05, 21.90) and associated with lower HbA1c (mean ± standard deviation 4.55 ±0.68 versus 4.92 ±0.60, P = 0.02). No such differences in HbA1c were observed in those with pre-diabetes or diabetes. CONCLUSIONS: A high intake of n-3 LCPUFAs was associated with a reduced risk of AMI, independent of HbA1c, in diabetic patients, but with an increased risk of fatal AMI and lower HbA1c among patients without impaired glucose metabolism. Further studies should investigate whether patients with diabetes may benefit from having a high intake of n-3 LCPUFAs and whether patients with normal glucose tolerance should be careful with a very high intake of these fatty acids. TRIAL REGISTRATION: This trial is registered at clinicaltrials.gov as NCT00354081.

Efficient burst mode amplifier for ultra-short pulses based on cryogenically cooled Yb³âº:CaF2.

We present a novel approach for the amplification of high peak power femtosecond laser pulses at a high repetition rate. This approach is based on an all-diode pumped burst mode laser scheme. In this scheme, pulse bursts with a total duration between 1 and 2 ms are be generated and amplified. They contain 50 to 2000 individual pulses equally spaced in time. The individual pulses have an initial duration of 350 fs and are stretched to 50 ps prior to amplification. The amplifier stage is based on Yb3+:CaF2 cooled to 100 K. In this amplifier, a total output energy in excess of 600 mJ per burst at a repetition rate of 10 Hz is demonstrated. For lower repetition rates the total output energy per burst can be scaled up to 915 mJ using a longer pump duration. This corresponds to an efficiency as high as 25% of extracted energy from absorbed pump energy. This is the highest efficiency, which has so far been demonstrated for a pulsed Yb3+:CaF2 amplifier.