CaMKII Splice Variants in Vascular Smooth Muscle Cells: The Next Step or Redundancy?

Vascular smooth muscle cells (VSMCs) help to maintain the normal physiological contractility of arterial vessels to control blood pressure; they can also contribute to vascular disease such as atherosclerosis. Ca2+/calmodulin-dependent kinase II (CaMKII), a multifunctional enzyme with four isoforms and multiple alternative splice variants, contributes to numerous functions within VSMCs. The role of these isoforms has been widely studied across numerous tissue types; however, their functions are still largely unknown within the vasculature. Even more understudied is the role of the different splice variants of each isoform in such signaling pathways. This review evaluates the role of the different CaMKII splice variants in vascular pathological and physiological mechanisms, aiming to show the need for more research to highlight both the deleterious and protective functions of the various splice variants.

Incidence of malignancy in multiple sclerosis: A cohort study in the Danish Multiple Sclerosis Registry.

BACKGROUND: The association between multiple sclerosis and malignancy is controversial and a current appraisal is needed. OBJECTIVE: To determine the incidence of malignancy in patients with multiple sclerosis compared with the general population and in relation to disease-modifying therapy. METHODS: Patients with multiple sclerosis (1995 - 2015) were matched by birth year and sex to individuals without multiple sclerosis in the general population. Patients with multiple sclerosis initiating disease-modifying therapy were evaluated using landmark period analysis. Malignancy risk was assessed by incidence rates, incidence rate ratios, and standardised incidence ratios. RESULTS: The standardised incidence ratio of any malignancy (excluding non-melanoma skin cancer) in patients with multiple sclerosis (n = 10,557) was 0.96 (95% CI 0.88 - 1.06), and there was no increased incidence of specific malignancy types compared with the general population cohort (n = 103,761). At the 48-month landmark period, the age-adjusted incidence per 100,000 person-years of any malignancy (excluding non-melanoma skin cancer) was 436.7 (95% CI 361.0 - 512.4) in patients newly treated with immunomodulator-only and 675.1 (95% CI 130.4 - 1219.9) in patients newly treated with immunosuppressant-only. CONCLUSIONS: There was no increased incidence of malignancy overall or by type in patients with multiple sclerosis compared neither with the general population nor in relation to disease-modifying therapy.

Achieving a flat, wideband frequency response of a loudspeaker unit by numerical optimization with requirements on its directivity.

This paper demonstrates how significant improvement in frequency response and directivity of a loudspeaker may be obtained by optimizing the local properties of the materials for the diaphragm and surround. Performance is investigated as the considered frequency range and off-axis requirements are progressively expanded. The results are generated by optimizing the values and layout of stiffness, mass, and damping of both the speaker diaphragm and surround. This is accomplished using a density and gradient-based optimization technique in conjunction with a fully coupled finite element model of the loudspeaker and the surrounding acoustic domain. The targeted frequency range is from 600 Hz up to 10 kHz and the range for the directivity is from 0° to 30°. The results show that a completely flat on-axis response is achievable even for very broad frequency ranges and that a reasonably flat response over a wide directivity can be obtained as well. The results presented in this research assume that complete design and production freedom are available.

Clonal replacement of meticillin-resistant Staphylococcus aureus during repeated outbreaks in a long-term care facility.

Meticillin-resistant Staphylococcus aureus (MRSA) is common among residents of long-term care facilities (LTCFs). Analysing the spa types of 22 isolates, mostly bloodstream infections (BSI), revealed five temporally distinct clonal outbreaks occurring in one ward of our local LTCF between 2012 and 2019. Each clone caused episodes of BSI for several months until replaced by another clone. A high MRSA carriage rate of 32% among healthcare workers in this ward was documented during the investigation of the 2019 outbreak. Clonal replacement of MRSA and the role of healthcare workers in transmission are discussed.

Large-scale outdoor sound field control.

The feasibility and the performance of controlling low frequency sound of loudspeaker systems under varying atmospheric conditions is examined experimentally. In the experiment, a control subwoofer array is canceling the sound of a primary subwoofer array over long distances (∼100 m) and in large areas (∼320 m2) using the pressure-matching method. To avoid the measurement of the sound field over the entire control area, a sound propagation model is introduced that is fitted in situ to model the radiation properties of the loudspeakers and the variation of the speed of sound. The results show that the control system reduces the sound pressure levels by up to 15-20 dB over the subwoofers' frequency range. However, the reduction can vary considerably depending on the specific atmospheric condition. The model-based approach reduces the number of required measurements and achieves similar reduction performance to the control based on direct measurements with considerably fewer microphone locations while also being more robust. Additionally, the sound propagation model enables the reduction of acoustic energy in virtual control zones that are far away from the microphone location. The investigated methodology has a direct application in the mitigation of sound from outdoor concerts.

Heterologous production of the widely used natural food colorant carminic acid in Aspergillus nidulans.

The natural red food colorants carmine (E120) and carminic acid are currently produced from scale insects. The access to raw material is limited and current production is sensitive to fluctuation in weather conditions. A cheaper and more stable supply is therefore desirable. Here we present the first proof-of-concept of heterologous microbial production of carminic acid in Aspergillus nidulans by developing a semi-natural biosynthetic pathway. Formation of the tricyclic core of carminic acid is achieved via a two-step process wherein a plant type III polyketide synthase (PKS) forms a non-reduced linear octaketide, which subsequently is folded into the desired flavokermesic acid anthrone (FKA) structure by a cyclase and a aromatase from a bacterial type II PKS system. The formed FKA is oxidized to flavokermesic acid and kermesic acid, catalyzed by endogenous A. nidulans monooxygenases, and further converted to dcII and carminic acid by the Dactylopius coccus C-glucosyltransferase DcUGT2. The establishment of a functional biosynthetic carminic acid pathway in A. nidulans serves as an important step towards industrial-scale production of carminic acid via liquid-state fermentation using a microbial cell factory.

Active room compensation for sound reinforcement using sound field separation techniques.

This work investigates how the sound field created by a sound reinforcement system can be controlled at low frequencies. An indoor control method is proposed which actively absorbs the sound incident on a reflecting boundary using an array of secondary sources. The sound field is separated into incident and reflected components by a microphone array close to the secondary sources, enabling the minimization of reflected components by means of optimal signals for the secondary sources. The method is purely feed-forward and assumes constant room conditions. Three different sound field separation techniques for the modeling of the reflections are investigated based on plane wave decomposition, equivalent sources, and the Spatial Fourier transform. Simulations and an experimental validation are presented, showing that the control method performs similarly well at enhancing low frequency responses with the three sound separation techniques. Resonances in the entire room are reduced, although the microphone array and secondary sources are confined to a small region close to the reflecting wall. Unlike previous control methods based on the creation of a plane wave sound field, the investigated method works in arbitrary room geometries and primary source positions.

Characterization of a membrane-bound C-glucosyltransferase responsible for carminic acid biosynthesis in Dactylopius coccus Costa.

Carminic acid, a glucosylated anthraquinone found in scale insects like Dactylopius coccus, has since ancient times been used as a red colorant in various applications. Here we show that a membrane-bound C-glucosyltransferase, isolated from D. coccus and designated DcUGT2, catalyzes the glucosylation of flavokermesic acid and kermesic acid into their respective C-glucosides dcII and carminic acid. DcUGT2 is predicted to be a type I integral endoplasmic reticulum (ER) membrane protein, containing a cleavable N-terminal signal peptide and a C-terminal transmembrane helix that anchors the protein to the ER, followed by a short cytoplasmic tail. DcUGT2 is found to be heavily glycosylated. Truncated DcUGT2 proteins synthesized in yeast indicate the presence of an internal ER-targeting signal. The cleavable N-terminal signal peptide is shown to be essential for the activity of DcUGT2, whereas the transmembrane helix/cytoplasmic domains, although important, are not crucial for its catalytic function.

Investigating the effect of banning non-reduced ignition propensity cigarettes on fatal residential fires in Sweden.

BACKGROUND: Annually, 100 people die as a result of residential fires in Sweden and almost a third of the fatal fires are known to be caused by smoking. In an attempt to reduce the occurrence of these events, reduced ignition propensity (RIP) cigarettes have been developed. They are designed to reduce the risk of fire by preventing the cigarette from burning through the full length when left unattended. In November 2011, a ban was introduced, forbidding the production and sale of all non-RIP cigarettes in all member states of the European Union, including Sweden. METHODS: Monthly data on all recorded residential fires and associated fatalities in Sweden from January 2000 to December 2013 were analyzed using an interrupted time series design. The effect of the intervention [in relative risk (RR)] was quantified using generalised additive models for location, shape and scale. RESULTS: There were no statistically significant intervention effects on residential fires (RR 0.95 [95% CI: 0.89-1.01]), fatal residential fires (RR 0.99 [95% CI: 0.80-1.23]), residential fires where smoking was a known cause (RR 1.10 [95% CI: 0.95-1.28]) or fatal residential fires where smoking was a known cause (RR 0.92 [95% CI: 0.63-1.35]). CONCLUSION: No evidence of an effect of the ban on all non-RIP cigarettes on the risk of residential fires in Sweden was found. The results may not be generalisable to other countries.

Reproduction of nearby sources by imposing true interaural differences on a sound field control approach.

In anechoic conditions, the Interaural Level Difference (ILD) is the most significant auditory cue to judge the distance to a sound source located within 1 m of the listener's head. This is due to the unique characteristics of a point source in its near field, which result in exceptionally high, distance dependent ILDs. When reproducing the sound field of sources located near the head with line or circular arrays of loudspeakers, the reproduced ILDs are generally lower than expected, due to physical limitations. This study presents an approach that combines a sound field reproduction method, known as Pressure Matching (PM), and a binaural control technique. While PM aims at reproducing the incident sound field, the objective of the binaural control technique is to ensure a correct reproduction of interaural differences. The combination of these two approaches gives rise to the following features: (i) an accurate reproduction of ILDs is achieved at the head positions considered by the method, (ii) the ILD variations in the vicinity of those positions are smoothed, thus lowering the ILD error, and (iii) the true wavefront is preserved. Given the properties of the presented method, intended distance and directional perception is expected.

De novo biosynthesis of vanillin in fission yeast (Schizosaccharomyces pombe) and baker's yeast (Saccharomyces cerevisiae).

Vanillin is one of the world's most important flavor compounds, with a global market of 180 million dollars. Natural vanillin is derived from the cured seed pods of the vanilla orchid (Vanilla planifolia), but most of the world's vanillin is synthesized from petrochemicals or wood pulp lignins. We have established a true de novo biosynthetic pathway for vanillin production from glucose in Schizosaccharomyces pombe, also known as fission yeast or African beer yeast, as well as in baker's yeast, Saccharomyces cerevisiae. Productivities were 65 and 45 mg/liter, after introduction of three and four heterologous genes, respectively. The engineered pathways involve incorporation of 3-dehydroshikimate dehydratase from the dung mold Podospora pauciseta, an aromatic carboxylic acid reductase (ACAR) from a bacterium of the Nocardia genus, and an O-methyltransferase from Homo sapiens. In S. cerevisiae, the ACAR enzyme required activation by phosphopantetheinylation, and this was achieved by coexpression of a Corynebacterium glutamicum phosphopantetheinyl transferase. Prevention of reduction of vanillin to vanillyl alcohol was achieved by knockout of the host alcohol dehydrogenase ADH6. In S. pombe, the biosynthesis was further improved by introduction of an Arabidopsis thaliana family 1 UDP-glycosyltransferase, converting vanillin into vanillin beta-D-glucoside, which is not toxic to the yeast cells and thus may be accumulated in larger amounts. These de novo pathways represent the first examples of one-cell microbial generation of these valuable compounds from glucose. S. pombe yeast has not previously been metabolically engineered to produce any valuable, industrially scalable, white biotech commodity.

Improved spiral tube assembly for high-speed counter-current chromatography.

The original spiral tube support (STS) assembly is improved by changing the shape of the tubing, with 1-cm presses perpendicularly along the length. This modification interrupts the laminar flow of the mobile phase. The tubing in the four return grooves to the center of the rotor is flattened by a specially made pressing tool to increase the number of spiral layers and decrease the dead space volume, thus increasing the column efficiency. The performance of this spiral tube assembly was tested in separations of dipeptides and proteins with suitable polar two-phase solvent systems. The results revealed that the present system yields high partition efficiency with a satisfactory level of stationary phase retention in a short elution time. The present high-speed counter-current chromatographic (HSCCC) system will be efficiently applied to a broad spectrum of two-phase solvent systems including aqueous-aqueous polymer phase systems (TPAS) which are used for separation of biopolymers such as proteins and nucleic acids.

Spiral Tube Assembly for High-Speed Countercurrent Chromatography: Choice of Elution Modes for Four Typical Two-Phase Solvent Systems.

Optimal elution modes were determined for four typical two-phase solvent systems each with different physical parameters to achieve the best peak resolution and retention of the stationary phase by spiral tube high-speed countercurrent chromatography using a suitable set of test samples. Both retention of the stationary phase and partition efficiency are governed by an interplay between two forces, i.e., Archimedean Screw force and radial centrifugal force gradient of the spiral channel. In the polar solvent system represented by 1-butanol./acetic acid/water (4:1:5, v/v/v) with settling time of over 30 s, the effect by the radial centrifugal gradient force dominates giving the best separation of dipeptides either by pumping the lower phase from the inner terminal or the upper phase from the outer terminal of the spiral channel. In the moderately hydrophobic two-phase solvent system represented by hexane/ethyl acetate/methanol/0.1 M HCl (1:1:1:1) with settling time of 19 s, and two hydrophobic solvent systems of hexane/ethanol/water (5:4:1, v/v/v) and non-aqueous binary system of hexane/acetonitrile both having settling time of 9, the effect of the Archimedean screw force play a major role in hydrodynamic equilibrium, giving the best separations by pumping the lower phase from the head or the upper phase from the tail of the spiral channel.

Fasting in healthy subjects is associated with intrahepatic accumulation of lipids as assessed by 1H-magnetic resonance spectroscopy.

The impact of fasting on IHL (intrahepatic lipid) content in human subjects has not been investigated previously, but results indicate that it may change rapidly in response to metabolic cues. The aim of the present study was to measure IHL content after fasting and to correlate this with circulating lipid intermediates. A total of eight healthy non-obese young males were studied before and after 12 or 36 h of fasting. IHL content was assessed by (1)H-magnetic resonance spectroscopy, and blood samples were drawn after the fasting period. IHL content increased significantly after the 36 h fasting period [median increase 156% (range, 4-252%); P<0.05]. Furthermore, a significant positive correlation between this increase and 3-hydroxybutyrate concentration was detected (P=0.03). No significant change in IHL content was demonstrated after the 12 h fasting period. The baseline median inter-individual variation in IHLs was 0.51% (range, 0.25-0.72%). The coefficient of variation of IHL measurements was 11.6%; 25-30% of the variation was of analytical origin and the remaining 70-75% was attributed to repositioning. In conclusion, IHL content increases in healthy male subjects during fasting, which demonstrates that nutritional status should be accounted for when assessing IHLs in clinical studies. Moreover, the increase in IHLs was positively correlated with the concentration of 3-hydroxybutyrate.

Postnatal adrenalectomy impairs urinary concentrating ability by increased COX-2 and leads to renal medullary injury.

We hypothesized that aldosterone promotes development of the renal medulla in the postnatal period and that cyclooxygenase-2 (COX-2) activity contributes to renal dysfunction after impaired aldosterone signaling. To test these hypotheses, rat pups underwent either sham operation or adrenalectomy at postnatal day 10. Adrenalectomized rats were divided into no steroid substitution (ADX), corticosterone replacement (ADX-C), and corticosterone and DOCA substitution (ADX-CD) groups that received subcutaneous pellets with steroids. Without replacement, pups failed to thrive and exhibited impaired urinary-concentrating ability. The renal medulla was significantly smaller, and the medullary interstitial osmolality was lower in the ADX group, whereas COX-2 and PGE2 tissue levels were significantly elevated compared with levels shown in sham animals. Substitution with DOCA and corticosterone corrected these changes, whereas corticosterone replacement alone improved survival but not weight gain and urinary-concentrating ability. Administration of a COX-2 inhibitor to ADX rats (parecoxib, 5 mg.kg(-1).day(-1), days 17-20) increased weight gain, urinary-concentrating ability, and papillary osmolality. After fluid deprivation, parecoxib attenuated weight loss and the increase in plasma Na+ concentration and osmolality. It is concluded that mineralocorticoid is required for normal postnatal development of the renal medulla. COX-2 contributes to impaired urine-concentrating ability, NaCl loss, and extracellular volume depletion in postnatal mineralocorticoid deficiency.

Dynamics of proviral HIV-1 184M/V in circulating CD4+ T-cells: a 90 days follow-up study after initiation/switch of HAART.

HIV-1 viral load falls rapidly on initiation of HAART. This phase of decreasing yet substantial viral production in the presence of antiretroviral drugs could generate resistant HIV-1. Whether switching a drug from a failing regime changes the demography of the mutations associated with it in the CD4+ T-cell compartment is not well-defined. We investigated the presence/absence and quantity of 184M and 184V in the CD4+ T-cell compartment of naïve patients initiated to HAART (group I), and patients who shifted to a non-lamivudine therapy (group II). We initiated a prospective 90 d follow-up study of 11 patients to detect and quantity proviral HIV-1 184M and 184V in the CD4+ T-cell compartment with a sensitive real time PCR assay. Results showed that the 184V was not detected in the CD4+ T-cell compartment of any of the 7 naïve patients who started on HAART. Three out of the 4 patients in group II experienced a fall in the percentage of 184V, with reduction to below detection limits in 2 patients. It can be concluded that initiation of HAART does not allow the archiving of the lamivudine associated mutation, 184V, in the CD4+ T-cell compartment. Reduction in the quantity of 184V when therapy is switched to an effective non-lamivudine regime indicates that the mutation in this compartment is dynamic.

HIV-1 reverse transcriptase gene 103K/N and 184M/V combinations in tandem: detection and quantification of HIV-1 populations in the CD45RO+ T-cell compartment by a double-ARMS real-time PCR assay.

BACKGROUND: The proviral HIV-1 reverse transcriptase gene for the 103K/N and 184M/V combinations were studied in tandem. The CD45RO T (memory) cell compartment was investigated. METHODS: A new double-ARMS (amplification refractory mutation system) real-time polymerase chain reaction assay was developed to detect and quantify 4 populations (103K-184M, 103K-184V, 103N-184M, and 103N-184V) in the CD45RO T-cell compartment. Twenty-one patients, 18 lamivudine and efavirenz/nevirapine experienced, were enrolled in a cross-sectional study. RESULTS: None of the mutation combinations were detected in patients on highly active antiretroviral therapy (HAART) (naive at start) with viremia suppression below detection limits. Conversely, all patients exposed to mono- or dual therapy (prior to HAART) carried at least 1 mutation combination regardless of viral load. In 9 patients, 17 mutations were detected in a mosaic of combinations. This study provides definite evidence of the existence of 103N and 184V mutation quasi-populations in tandem, and separately in combination with the wild-type codons, 184M and 103K, in the CD45RO T-cell compartment. CONCLUSIONS: The initiation and continuation of potent antiretroviral therapy effectively hinders the appearance of 103N and 184V mutations alone or in tandem in memory cells. When switching therapies because of failure, caution should be exercised with drugs associated with single-mutation threshold; they can appear in tandem with contemporary resistant virus populations, leading to multidrug resistance.