Correlative chemical and elemental nano-imaging of morphology and disorder at the nacre-prismatic region interface in Pinctada margaritifera.

Understanding biomineralization relies on imaging chemically heterogeneous organic-inorganic interfaces across a hierarchy of spatial scales. Further, organic minority phases are often responsible for emergent inorganic structures from the atomic arrangement of different polymorphs, to nano- and micrometer crystal dimensions, up to meter size mollusk shells. The desired simultaneous chemical and elemental imaging to identify sparse organic moieties across a large field-of-view with nanometer spatial resolution has not yet been achieved. Here, we combine nanoscale secondary ion mass spectroscopy (NanoSIMS) with spectroscopic IR s-SNOM imaging for simultaneous chemical, molecular, and elemental nanoimaging. At the example of Pinctada margaritifera mollusk shells we identify and resolve ~ 50 nm interlamellar protein sheets periodically arranged in regular ~ 600 nm intervals. The striations typically appear ~ 15 µm from the nacre-prism boundary at the interface between disordered neonacre to mature nacre. Using the polymorph distinctive IR-vibrational carbonate resonance, the nacre and prismatic regions are consistently identified as aragonite ([Formula: see text] cm-1) and calcite ([Formula: see text] cm-1), respectively. We observe previously unreported morphological features including aragonite subdomains encapsulated in extensions of the prism-covering organic membrane and regions of irregular nacre tablet formation coincident with dispersed organics. We also identify a ~ 200 nm region in the incipient nacre region with less well-defined crystal structure and integrated organics. These results show with the identification of the interlamellar protein layer how correlative nano-IR chemical and NanoSIMS elemental imaging can help distinguish different models proposed for shell growth in particular, and how organic function may relate to inorganic structure in other biomineralized systems in general.

Agreement of Computerized QT and QTc Interval Measurements Between Both Bedside and Expert Nurses Using Electronic Calipers.

BACKGROUND: In hospitalized patients, QT/QTc (heart rate corrected) prolongation on the electrocardiogram (ECG) increases the risk of torsade de pointes. Manual measurements are time-consuming and often inaccurate. Some bedside monitors automatically and continuously measure QT/QTc; however, the agreement between computerized versus nurse-measured values has not been evaluated. OBJECTIVE: The aim of this study was to examine the agreement between computerized QT/QTc and bedside and expert nurses who used electronic calipers. METHODS: This was a prospective observational study in 3 intensive care units. Up to 2 QT/QTc measurements (milliseconds) per patient were collected. Bland-Altman test was used to analyze measurement agreement. RESULTS: A total of 54 QT/QTc measurements from 34 patients admitted to the ICU were included. The mean difference (bias) for QT comparisons was as follows: computerized versus expert nurses, -11.04 ± 4.45 milliseconds (95% confidence interval [CI], -2.3 to -19.8; P = .016), and computerized versus bedside nurses, -13.72 ± 6.70 (95% CI, -0.70 to -26.8; P = .044). The mean bias for QTc comparisons was as follows: computerized versus expert nurses, -12.46 ± 5.80 (95% CI, -1.1 to -23.8; P = .035), and computerized versus bedside nurses, -18.49 ± 7.90 (95% CI, -3.0 to -33.9; P = .022). CONCLUSION: Computerized QT/QTc measurements calculated by bedside monitor software and measurements performed by nurses were in close agreement; statistically significant differences were found, but differences were less than 20 milliseconds (on-half of a small box), indicating no clinical significance. Computerized measurements may be a suitable alternative to nurse-measured QT/QTc. This could reduce inaccuracies and nurse burden while increasing adherence to practice recommendations. Further research comparing computerized QT/QTc from bedside monitoring to standard 12-lead electrocardiogram in a larger sample, including non-ICU patients, is needed.

Blunted nutrient-response pathways in adipose tissue following high fat meals in men with metabolic syndrome: A randomized postprandial transcriptomic study.

BACKGROUND: Excessive adipose tissue is central to disease burden posed by the Metabolic Syndrome (MetS). Whilst much is known of the altered transcriptomic regulation of adipose tissue under fasting conditions, little is known of the responses to high-fat meals. METHODS: Nineteen middle-aged males (mean ± SD 52.0 ± 4.6 years), consumed two isocaloric high-fat, predominately dairy-based or soy-based, breakfast meals. Abdominal subcutaneous adipose biopsies were collected after overnight fast (0 h) and 4 h following each meal. Global gene expression profiling was performed by microarray (Illumina Human WG-6 v3). RESULTS: In the fasted state, 13 genes were differently expressed between control and MetS adipose tissue (≥1.2 fold-difference, p < 0.05). In response to the meals, the control participants had widespread increases in genes related to cellular nutrient responses (≥1.2 fold-change, p < 0.05; 2444 & 2367 genes; dairy & soy, respectively). There was blunted response in the MetS group (≥1.2 fold-change, p < 0.05; 332 & 336 genes; dairy & soy, respectively). CONCLUSIONS: In middle-aged males with MetS, a widespread suppression of the subcutaneous adipose tissue nutrient responsive gene expression suggests an inflexibility in the transcriptomic responsiveness to both high-fat meals.

Actionable Ventricular Tachycardia During In-Hospital ECG Monitoring and Its Impact on Alarm Fatigue.

BACKGROUND: Ventricular tachycardia (V-tach) is the most common lethal arrhythmia, yet 90% of alarms are false and contribute to alarm fatigue. We hypothesize that some true V-tach also causes alarm fatigue because current criteria are too sensitive (i.e., ≥6 beats ≥100 beats/min [bpm]). PURPOSE: This study was designed to determine (1) the proportion of clinically actionable true V-tach events; (2) whether true actionable versus nonactionable V-tach differs in terms of heart rate and/or duration (seconds); and (3) if actionable V-tach is associated with adverse outcomes. METHODS: This was a secondary analysis in 460 intensive care unit (ICU) patients. Electronic health records were examined to determine if a V-tach event was actionable or nonactionable. Actionable V-tach was defined if a clinical action(s) was taken within 15 minutes of its occurrence (i.e., new and/or change of medication, defibrillation, and/or laboratory test). Maximal heart rate and duration for each V-tach event were measured from bedside monitor electrocardiography. Adverse patient outcomes included a code blue event and/or death. RESULTS: In 460 ICU patients, 50 (11%) had 151 true V-tach events (range 1-20). Of the 50 patients, 40 (80%) had only nonactionable V-tach (97 events); 3 (6%) had both actionable and nonactionable V-tach (32 events); and 7 patients (14%) had only actionable V-tach (23 events). There were differences in duration comparing actionable versus nonactionable V-tach (mean 56.19 ± 116.87 seconds vs. 4.28 ± 4.09 seconds; P = 0.001) and maximal heart rate (188.81 ± 116.83 bpm vs. 150.79 ± 28.26 bpm; P = 0.001). Of the 50 patients, 3 (6%) had a code blue, 2 died, and all were in the actionable V-tach group. CONCLUSIONS: In our sample, <1% experienced a code blue following true V-tach. Heart rate and duration for actionable V-tach were much faster and longer than that for nonactionable V-tach. Current default settings typically used for electrocardiographic monitoring (i.e., ≥6 beats ≥100 bpm) appear to be too conservative and can lead to crisis/red level nuisance alarms that contribute to alarm fatigue. A prospective study designed to test whether adjusting default settings to these higher levels is safe for patients is needed.

Glasgow Coma Scale: Generating Clinical Standards.

BACKGROUND: The Glasgow Coma Scale (GCS) is a tool used to aid in objectively measuring the neurological status of a patient. This study aimed to evaluate the limitations and discrepancies in GCS use among nurses in an academic medical center neurological intensive care unit and compile evidence for development of a standardized GCS educational program. METHODS: Twenty nurse participants completed a survey before attending an educational intervention. Participants then attended a 90-minute educational intervention. In follow-up, participants were asked to complete a postsurvey. RESULTS: The standardized GCS educational program significantly improved nurse knowledge of the GCS as measured by presurvey and postsurvey general GCS question scores. Educational programming improved application of the GCS as measured by presurvey and postsurvey GCS verbal component, motor component, and sum scores. GCS motor score performance was the least accurate component. CONCLUSION: Participants reported that the education has informed the unit culture and emboldened clinical nurses to speak to their practice with more authority. Educational interventions should be aimed toward applied transfer of knowledge to the case-based scenarios in the clinical setting.

Divergent shifts in lipid mediator profile following supplementation with n-3 docosapentaenoic acid and eicosapentaenoic acid.

In contrast to the well-characterized effects of specialized proresolving lipid mediators (SPMs) derived from eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA), little is known about the metabolic fate of the intermediary long-chain (LC) n-3 polyunsaturated fatty acid (PUFA) docosapentaenoic acid (DPA). In this double blind crossover study, shifts in circulating levels of n-3 and n-6 PUFA-derived bioactive lipid mediators were quantified by an unbiased liquid chromatography-tandem mass spectrometry lipidomic approach. Plasma was obtained from human subjects before and after 7 d of supplementation with pure n-3 DPA, n-3 EPA or placebo (olive oil). DPA supplementation increased the SPM resolvin D5n-3DPA (RvD5n-3DPA) and maresin (MaR)-1, the DHA vicinal diol 19,20-dihydroxy-DPA and n-6 PUFA derived 15-keto-PG E2 (15-keto-PGE2). EPA supplementation had no effect on any plasma DPA or DHA derived mediators, but markedly elevated monohydroxy-eicosapentaenoic acids (HEPEs), including the e-series resolvin (RvE) precursor 18-HEPE; effects not observed with DPA supplementation. These data show that dietary n-3 DPA and EPA have highly divergent effects on human lipid mediator profile, with no overlap in PUFA metabolites formed. The recently uncovered biologic activity of n-3 DPA docosanoids and their marked modulation by dietary DPA intake reveals a unique and specific role of n-3 DPA in human physiology.-Markworth, J. F., Kaur, G., Miller, E. G., Larsen, A. E., Sinclair, A. J., Maddipati, K. R., Cameron-Smith, D. Divergent shifts in lipid mediator profile following supplementation with n-3 docosapentaenoic acid and eicosapentaenoic acid.

Postprandial Responses to Lipid and Carbohydrate Ingestion in Repeated Subcutaneous Adipose Tissue Biopsies in Healthy Adults.

Adipose tissue is a primary site of meta-inflammation. Diet composition influences adipose tissue metabolism and a single meal can drive an inflammatory response in postprandial period. This study aimed to examine the effect lipid and carbohydrate ingestion compared with a non-caloric placebo on adipose tissue response. Thirty-three healthy adults (age 24.5 ± 3.3 year (mean ± standard deviation (SD)); body mass index (BMI) 24.1 ± 3.2 kg/m2, were randomised into one of three parallel beverage groups; placebo (water), carbohydrate (maltodextrin) or lipid (dairy-cream). Subcutaneous, abdominal adipose tissue biopsies and serum samples were collected prior to (0 h), as well as 2 h and 4 h after consumption of the beverage. Adipose tissue gene expression levels of monocyte chemoattractant protein-1 (MCP-1), interleukin 6 (IL-6) and tumor necrosis factor-α (TNF-α) increased in all three groups, without an increase in circulating TNF-α. Serum leptin (0.6-fold, p = 0.03) and adipose tissue leptin gene expression levels (0.6-fold, p = 0.001) decreased in the hours following the placebo beverage, but not the nutrient beverages. Despite increased inflammatory cytokine gene expression in adipose tissue with all beverages, suggesting a confounding effect of the repeated biopsy method, differences in metabolic responses of adipose tissue and circulating adipokines to ingestion of lipid and carbohydrate beverages were observed.

Postprandial Plasma Phospholipids in Men Are Influenced by the Source of Dietary Fat.

BACKGROUND: Postprandial lipemia represents a risk factor for chronic diseases, including type 2 diabetes. Little is known about the effect of dietary fat on the plasma lipidome in the postprandial period. OBJECTIVE: The objective of this study was to assess the effect of dairy fat and soy oil on circulating postprandial lipids in men. METHODS: Men (40-60 y old, nonsmokers; n = 16) were randomly assigned in a crossover design to consume 2 breakfast meals of dairy-based or soy oil-based foods. The changes in the plasma lipidome during the 4-h postprandial period were analyzed with electrospray ionization tandem mass spectrometry and included 316 lipid species in 23 classes and subclasses, representing sphingolipids, phospholipids, glycerolipids, and sterols. RESULTS: Nonparametric Friedman tests showed significant changes in multiple plasma lipid classes, subclasses, and species in the postprandial period after both dairy and soy meals. No difference was found in triglyceridemia after each meal. However, 6 endogenous lipid classes increased after dairy but decreased after soy (P < 0.05), including ether-linked phospholipids and plasmalogens and sphingomyelin (not present in soy), dihexosylceramide, and GM3 ganglioside. Phosphatidylcholine and phosphatidylinositol were not affected by the soy meal but were significantly elevated after the dairy meal (8.3% and 16%, respectively; P < 0.05). CONCLUSIONS: The changes in postprandial plasma phospholipids in men relate to the diet composition and the relative size of the endogenous phospholipid pools. Despite similar lipemic responses as measured by changes in triglyceride concentrations, the differential responses to dairy and soy meals derived through lipidomic analysis of phospholipids suggest differences in the metabolism of soybean oil and dairy fat. The increased concentrations of plasmalogens, with potential antioxidant capacity, in the postprandial period after dairy but not soy meals may represent a further important difference in the response to these sources of fat. The trial was registered at www.anzctr.org.au as ACTRN12610000562077.

Muscle p70S6K phosphorylation in response to soy and dairy rich meals in middle aged men with metabolic syndrome: a randomised crossover trial.

BACKGROUND: The mammalian target of rapamycin (mTOR) pathway is the primary regulator of muscle protein synthesis. Metabolic syndrome (MetS) is characterized by central obesity and insulin resistance; little is known about how MetS affects the sensitivity of the mTOR pathway to feeding. METHODS: The responsiveness of mTOR pathway targets such as p706Sk to a high protein meal containing either dairy or soy foods was investigated in healthy insulin sensitive middle-aged men and those presenting with metabolic syndrome (MetS). Twenty male subjects (10 healthy controls, 10 MetS) participated in a single-blinded randomized cross-over study. In a random sequence, subjects ingested energy-matched breakfasts composed predominately of either dairy-protein or soy-protein foods. Skeletal muscle biopsies were collected in the fasted state and at 2 and 4 h post-meal ingestion for the analysis of mTOR- and insulin-signalling kinase activation. RESULTS: Phosphorylated Akt and Insulin receptor substrate 1 (IRS1) increased during the postabsorptive period with no difference between groups. mTOR (Ser448) and ribosomal protein S6 phosphorylation increased 2 h following dairy meal consumption only. p70S6K (Thr389) phosphorylation was increased after feeding only in the control subjects and not in the MetS group. CONCLUSIONS: These data demonstrate that the consumption of a dairy-protein rich but not a soy-protein rich breakfast activates the phosphorylation of mTOR and ribosomal protein S6, required for protein synthesis in human skeletal muscle. Unlike healthy controls, subjects with MetS did not increase muscle p70S6K(Thr389) phosphorylation in response to a mixed meal. TRIAL REGISTRATION: This trial was registered with the Australian New Zealand Clinical Trials Registry (ANZCTR) as ACTRN12610000562077.

Endothelial permeability in vitro and in vivo: protective actions of ANP and omapatrilat in experimental atherosclerosis.

Increased arterial endothelial cell permeability (ECP) is considered an initial step in atherosclerosis. Atrial natriuretic peptide (ANP) which is rapidly degraded by neprilysin (NEP) may reduce injury-induced endothelial cell leakiness. Omapatrilat represents a first in class of pharmacological agents which inhibits both NEP and angiotensin converting enzyme (ACE). We hypothesized that ANP prevents thrombin-induced increases of ECP in human aortic ECs (HAECs) and that omapatrilat would reduce aortic leakiness and atherogenesis and enhance ANP mediated vasorelaxation of isolated aortas. Thrombin induced ECP determined by I(125) albumin flux was assessed in HAECs with and without ANP pretreatment. Next we examined the effects of chronic oral administration of omapatrilat (12 mg/kg/day, n=13) or placebo (n=13) for 8 weeks on aortic leakiness, atherogenesis and ANP-mediated vasorelaxation in isolated aortas in a rabbit model of atherosclerosis produced by high cholesterol diet. In HAECs, thrombin-induced increases in ECP were prevented by ANP. Omapatrilat reduced the area of increased aortic leakiness determined by Evans-blue dye and area of atheroma formation assessed by Oil-Red staining compared to placebo. In isolated arterial rings, omapatrilat enhanced vasorelaxation to ANP compared to placebo with and without the endothelium. ANP prevents thrombin-induced increases in ECP in HAECs. Chronic oral administration of omapatrilat reduces aortic leakiness and atheroma formation with enhanced endothelial independent vasorelaxation to ANP. These studies support the therapeutic potential of dual inhibition of NEP and ACE in the prevention of increased arterial ECP and atherogenesis which may be linked to the ANP/cGMP system.

Postprandial metabolism of docosapentaenoic acid (DPA, 22:5n-3) and eicosapentaenoic acid (EPA, 20:5n-3) in humans.

The study of the metabolism of docosapentaenoic acid (DPA, 22:5n-3) in humans has been limited by the unavailability of pure DPA and the fact that DPA is found in combination with eicosapentaenoic acid (EPA, 20:5n-3) and docosahexaenoic acid (DHA, 22:6n-3) in natural products. In this double blind cross over study, pure DPA and EPA were incorporated in meals served to healthy female volunteers. Mass spectrometric methods were used to study the chylomicron lipidomics. Plasma chylomicronemia was significantly reduced after the meal containing DPA compared with the meal containing EPA or olive oil only. Both EPA and DPA were incorporated into chylomicron TAGs, while there was less incorporation into chylomicron phospholipids. Lipidomic analysis of the chylomicron TAGs revealed the dynamic nature of chylomicron TAGs. The main TAG species that EPA and DPA were incorporated into were EPA/18:1/18:1, DPA/18:1/16:0 and DPA/18:1/18:1. There was very limited conversion of DPA and EPA to DHA and there were no increases in EPA levels during the 5h postprandial period after the DPA meal. In conclusion, EPA and DPA showed different metabolic fates, and DPA hindered the digestion, ingestion or incorporation into chylomicrons of the olive oil present in the meal.

A short-term n-3 DPA supplementation study in humans.

PURPOSE: Despite the detailed knowledge of the absorption and incorporation of eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) into plasma lipids and red blood cells (RBC) in humans, very little is known about docosapentaenoic acid (DPA, 22:5 n-3). The aim of this study was to investigate the uptake and incorporation of pure DPA and EPA into human plasma and RBC lipids. METHODS: Ten female participants received 8 g of pure DPA or pure EPA in randomized crossover double-blinded manner over a 7-day period. The placebo treatment was olive oil. Blood samples were collected at days zero, four and seven, following which the plasma and RBC were separated and used for the analysis of fatty acids. RESULTS: Supplementation with DPA significantly increased the proportions of DPA in the plasma phospholipids (PL) (by twofold) and triacylglycerol (TAG) fractions (by 2.3-fold, day 4). DPA supplementation also significantly increased the proportions of EPA in TAG (by 3.1-fold, day 4) and cholesterol ester (CE) fractions (by 2.0-fold, day 7) and of DHA in TAG fraction (by 3.1-fold, day 4). DPA proportions in RBC PL did not change following supplementation. Supplementation with EPA significantly increased the proportion of EPA in the plasma CE and PL fractions, (both by 2.7-fold, day 4 and day 7) and in the RBC PL (by 1.9-fold, day 4 and day 7). EPA supplementation did not alter the proportions of DPA or DHA in any lipid fraction. These results showed that within day 4 of supplementation, DPA and EPA demonstrated different and specific incorporation patterns. CONCLUSION: The results of this short-term study suggest that DPA may act as a reservoir of the major long-chain n-3 fatty acids (LC n-3 PUFA) in humans.

Lipidomic profiling of chylomicron triacylglycerols in response to high fat meals.

Using lipidomic methodologies the impact that meal lipid composition and metabolic syndrome (MetS) exerts on the postprandial chylomicron triacylglycerol (TAG) response was examined. Males (9 control; 11 MetS) participated in a randomised crossover trial ingesting two high fat breakfast meals composed of either dairy-based foods or vegetable oil-based foods. The postprandial lipidomic molecular composition of the TAG in the chylomicron-rich (CM) fraction was analysed with tandem mass spectrometry coupled with liquid chromatography to profile CM TAG species and targeted TAG regioisomers. Postprandial CM TAG concentrations were significantly lower after the dairy-based foods compared with the vegetable oil-based foods for both control and MetS subjects. The CM TAG response to the ingested meals involved both significant and differential depletion of TAG species containing shorter- and medium-chain fatty acids (FA) and enrichment of TAG molecular species containing C16 and C18 saturated, monounsaturated and diunsaturated FA. Furthermore, there were significant changes in the TAG species between the food TAG and CM TAG and between the 3- and 5-h postprandial samples for the CM TAG regioisomers. Unexpectedly, the postprandial CM TAG concentration and CM TAG lipidomic responses did not differ between the control and MetS subjects. Lipidomic analysing of CM TAG molecular species revealed dynamic changes in the molecular species of CM TAG during the postprandial phase suggesting either preferential CM TAG species formation and/or clearance.

Resistance exercise increases NF-κB activity in human skeletal muscle.

Intense resistance exercise causes a significant inflammatory response. NF-κB has been identified as a prospective key transcription factor mediating the postexercise inflammatory response. The purpose of this study was to determine whether a single bout of intense resistance exercise regulates NF-κB signaling in human skeletal muscle. Muscle biopsy samples were obtained from the vastus lateralis of five recreationally active, but not strength-trained, males (21.9 ± 1.3 yr) prior to, and at 2 and 4 h following, a single bout of intense resistance exercise. A further five subjects (4 males, 1 female) (23 ± 0.89 yr) were recruited as a nonexercise control group to examine the effect of the muscle biopsy protocol on key markers of skeletal muscle inflammation. Protein levels of IκBα and phosphorylated NF-κB (p65), as well as the mRNA expression of inflammatory myokines monocyte chemoattractant protein 1 (MCP-1), IL-6, and IL-8 were measured. Additionally, NF-κB (p65) DNA binding to the promoter regions of MCP-1, IL-6, and IL-8 was investigated. IκBα protein levels decreased, while p-NF-κB (p65) protein levels increased 2 h postexercise and returned to near-baseline levels by 4-h postexercise. Immunohistochemical data verified these findings, illustrating an increase in p-NF-κB (p65) protein levels, and nuclear localization at 2 h postexercise. Furthermore, NF-κB DNA binding to MCP-1, IL-6, and IL-8 promoter regions increased significantly 2 h postexercise as did mRNA levels of these myokines. No significant change was observed in the nonexercise control group. These novel data provide evidence that intense resistance exercise transiently activates NF-κB signaling in human skeletal muscle during the first few hours postexercise. These findings implicate NF-κB in the transcriptional control of myokines known to be central to the postexercise inflammatory response.

Impact of resistance exercise training on interleukin-6 and JAK/STAT in young men.

The JAK/STAT signaling pathway is essential for myogenic regeneration and is regulated by a diverse range of ligands, including interleukin-6 (IL-6) and platelet-derived growth factor-BB (PDGF-BB). Our aim was to evaluate the responsiveness of IL-6 and PDGF-BB to intense exercise, along with STAT3 activation, before and after 12 weeks of resistance training. In young men, IL-6 and PDGF-BB protein concentrations were quantified in biopsied muscle and increased at 3 h post-exercise (17.5-fold and 3-fold, respectively). The response was unaltered by 12 weeks of training. Similarly, STAT3 phosphorylation was elevated post-exercise (12.5-fold), irrespective of training status, as was the expression of downstream targets c-MYC (8-fold), c-FOS (4.5-fold), and SOCS3 (2.3-fold). Thus, intense exercise transiently increases IL-6 and PDGF-BB proteins, and STAT3 phosphorylation is increased. These responses are preserved after intense exercise. This suggests they are not modified by training and may be an essential component of the adaptive responses to intense exercise.

The role and regulation of MAFbx/atrogin-1 and MuRF1 in skeletal muscle atrophy.

Skeletal muscle atrophy occurs in many chronic diseases and disuse conditions. Its severity reduces patient recovery, independence and quality of life. The discovery of two muscle-specific E3 ubiquitin ligases, MAFbx/atrogin-1 and Muscle RING Finger-1 (MuRF1), promoted an expectation of these molecules as targets for therapeutic development. While numerous studies have determined the conditions in which MAFbx/atrogin-1 and MuRF1 mRNA levels are regulated, few studies have investigated their functional role in skeletal muscle. Recently, studies identifying new target substrates for MAFbx/atrogin-1 and MuRF1, outside of their response to the initiation of muscle atrophy, suggest that there is more to these proteins than previously appreciated. This review will highlight our present knowledge of MAFbx/atrogin-1 and MuRF1 in skeletal muscle atrophy, the impact of potential therapeutics and their known regulators and substrates. Finally, we will comment on new approaches that may expand our knowledge of these two molecules in their control of skeletal muscle function.

Effects of conjugated linoleic acid on myogenic and inflammatory responses in a human primary muscle and tumor coculture model.

The antiproliferative and anti-inflammatory properties of conjugated linoleic acid (CLA) make it a potentially novel treatment in chronic inflammatory muscle wasting disease, particularly cancer cachexia. Human primary muscle cells were grown in coculture with MIA PaCa-2 pancreatic tumor cells and exposed to varying concentrations of c9,t11 and t10,c12 CLA. Expression of myogenic (Myf5, MyoD, myogenin, and myostatin) and inflammatory genes (CCL-2, COX-2, IL-8, and TNF-alpha) were measured by real-time PCR. The t10,c12 CLA isomer, but not the c9,t11 isomer, significantly decreased MIA PaCa-2 proliferation by between 15% and 19%. There was a marked decrease in muscle MyoD and myogenin expression (78% and 62%, respectively), but no change in either Myf5 or myostatin, in myotubes grown in coculture with MIA PaCa-2 cells. CLA had limited influence on these responses. A similar pattern of myogenic gene expression changes was observed in myotubes treated with TNF-alpha alone. Several-fold significant increases in CCL-2, COX-2, IL-8, and TNF-alpha expression in myotubes were observed with MIA PaCa-2 coculture. The c9,t11 CLA isomer significantly decreased basal expression of TNF-alpha in myotubes and could ameliorate its tumor-induced rise. The study provides insight into the anti-inflammatory and antiproliferative actions of CLA and its application as a therapeutic agent in inflammatory disease states.

Brucella regulators: self-control in a hostile environment.

Brucella is an important zoonotic pathogen for which no human vaccine exists. In an infected host, Brucella resides in macrophages but must coordinate expression of multiple virulence factors for successful cell entry and trafficking to acquire this replicative niche. Brucella responds to environmental signals to regulate virulence strategies that circumvent or blunt the host immune response. The Brucella quorum sensing system is a nexus of control for several Brucella virulence factors including flagellar genes and the type IV secretion system. Other sensory transduction systems, such as BvrRS and the newly described LOV-HK, sense environmental factors to control virulence. Here, we examine the contributions of various regulatory systems to Brucella virulence.

Putative quorum-sensing regulator BlxR of Brucella melitensis regulates virulence factors including the type IV secretion system and flagella.

Brucella melitensis is an intracellular pathogen that establishes a replicative niche within macrophages. While the intracellular lifestyle of Brucella is poorly understood and few virulence factors have been identified, components of a quorum-sensing pathway in Brucella have recently been identified. The LuxR-type regulatory protein, VjbR, and an N-acylhomoserine lactone signaling molecule are both involved in regulating expression of the virB-encoded type IV secretion system. We have identified a second LuxR-type regulatory protein (BlxR) in Brucella. Microarray analysis of a blxR mutant suggests that BlxR regulates the expression of a number of genes, including those encoding the type IV secretion system and flagella. Confirming these results, deletion of blxR in B. melitensis reduced the transcriptional activities of promoters for the virB operon, flagellar genes, and another putative virulence factor gene, bopA. Furthermore, our data suggested that both BlxR and VjbR are positively autoregulated and cross-regulate the expression of each other. The blxR deletion strain exhibited reduced growth in macrophages, similar to that observed for a vjbR deletion strain. However, unlike the vjbR deletion, the blxR deletion did not fully attenuate virulence in mice. More strikingly, bioluminescent imaging revealed that dissemination of the blxR mutant was similar to that of wild-type B. melitensis, while the vjbR mutant was defective for systemic spread in IRF-1(-/-) mice, suggesting that these regulators are not functionally redundant but that they converge in a common pathway regulating bacterial processes.