Concentrations Of Iso Research Articles

The Interaction between Anesthetic Isoflurane and Model-Biomembrane Monolayer Using Simultaneous Quartz Crystal Microbalance (QCM) and Quartz Crystal Impedance (QCI) Methods.

The interaction between anesthetic Isoflurane (Iso) and model-biomembrane on the water surface has been investigated using quartz crystal microbalance (QCM) and quartz crystal impedance (QCI) methods. The model-biomembranes used were dipalmitoyl phosphatidyl choline (DPPC), DPPC-palmitic acid (PA) mixture (DPPC:PA = 8:2), DPPC-Alamethicin (Al) mixture (DPPC:Al = 39:1), and DPPC-β-Lactoglobulin (βLG) mixture (DPPC:βLG = 139:1) monolayers, respectively. The quartz crystal oscillator (QCO) was attached horizontally to each monolayer, and QCM and QCI measurements were performed simultaneously. It was found that Iso hydrate physisorbed on each monolayer/water interface from QCM and changed those interfacial viscosities from QCI. With an increase in Iso concentration, pure DPPC, DPPC-PA mixed, and DPPC-Al mixed monolayers showed a two-step process of Iso hydrate on both physisorption and viscosity, whereas it was a one-step for the DPPC-βLG mixed monolayer. The viscosity change in the DPPC-βLG mixed monolayer with the physisorption of Iso hydrate was much larger than that of other monolayers, in spite of the one-step process. From these results, the action mechanism of anesthetics and their relevance to the expression of anesthesia were discussed, based on the "release of interfacial hydrated water" hypothesis on the membrane/water interface.

Effects of direct-fed Bacillus subtilis and Bacillus licheniformis on production performance and milk fatty acid profile in dairy cows

The aim of the study was to determine the effect of a Bacillus-based direct-fed microbial on performance of mid-lactating Holstein dairy cows and on their milk fatty acid composition. Six multiparous cows fitted with a rumen cannula were used in a randomized replicated crossover design. Cows received 200 g/d of either whey powder as a control or BioPlus 2B (Chr. Hansen), a commercial direct-fed microbial providing Bacillus subtilis and Bacillus licheniformis, representing a daily dose of 6.4 × 1011 cfu, and using whey powder as a carrier. The 2 experimental periods lasted 14 d and were separated by a 7-d washout interval. Samples were collected on d 0, 13, and 14 of each period. Data from d 0 were used as covariate. Significance was declared at P ≤ 0.05 and tendency at 0.05 <P ≤ 0.10. There was a 10-fold increase in the relative concentration of bacteria from the Bacillus subtilis group in the rumen when feeding direct-fed Bacillus compared with control. Treatment did not affect ruminal pH, NH3-N, or concentrations of acetate, propionate, and butyrate. However, direct-fed Bacillus increased ruminal concentrations of isovalerate and isobutyrate (tendency). Treatments did not affect lactation performance. Supplying direct-fed Bacillus enhanced milk relative concentration of anteiso 13:0 by 27.3% and of anteiso 15:0 by 6.5% and tended to increase concentrations of iso 14:0 (+41.8%) relative to control. When expressed on a yield basis, direct-fed Bacillus increased the secretion of anteiso 13:0 and decreased that of 11:0, 15:0, 17:0 (tendency), and cis-9 17:1. These variations, although limited in magnitude, indicate that milk branched-chain fatty acid composition is sensitive to ruminal microbiota modifications without changes in chemical composition of the diet.

Direct numerical simulation of double diffusive natural convection in a closed mixture cavity heated from below

Double diffusive natural convection is one of the most widely used study subjects in heat and mass transfer. In this present study, double diffusive natural convection heating from below the wall in a closed cavity was studied with direct numerical simulation method. The flow characteristics are presented as isotherms, iso concentrations, and streamlines. The results show that RaT has signification strong effects on average Nusselt number, with a more considerable RaT, the value of Nusselt number more higher. These were also found to increase with increasing buoyancy ratio for adding flow and decrease as buoyancy ratio decreases for opposing flows

A kinetic study of the photolysis of sulfamethoxazole with special emphasis on the photoisomer

The previously not studied photochemical degradation of sulfamethoxazole (SMX) to the isomer of SMX (ISO) was measured via a polychromatic (Xe) and a monochromatic (Hg) light source and accompanied by quantum chemical DFT calculations. In addition to the mathrm{p}K_mathrm{a} = ;7.0 pm 0.1 of ISO, tautomer-dependent properties such as the K_mathrm{OW} were measured and theoretically confirmed by DFT. The kinetics in solutions below and above the mathrm{p}K_mathrm{a} = 5.6 of SMX were studied for the available and quantifiable products SMX, ISO, 3-amino-5-methylisoxazole (AMI), 2-amino-5-methyloxazole (AMO), and sulfanilic acid (SUA). The quantum yields of the neutral (Phi _mathrm{N}) and anionic Phi _mathrm{A}) forms of SMX (Phi _mathrm{A} = 0.03 pm 0.001, Phi _mathrm{N} = 0.15 pm 0.01) and ISO (Phi _mathrm{A} = 0.05 pm 0.01 and Phi _mathrm{N} = 0.06 pm 0.02) were found to be wavelength-independent. In a competitive reaction to the formation of ISO from SMX, the degradation product TP271 is formed. Various proposed structures for TP271 described in the literature have been studied quantum mechanically and can be excluded for thermodynamic reasons. In real samples in a northern German surface water in summer 2021 mean concentrations of SMX were found in the range of 120 ng/L. In agreement with the pH-dependent yields, concentrations of ISO were low in the range of 8 ng/L.Graphical abstract

Analysis of Residual Chlorine and Total Coliform on Consumers of Perumdam Maja Tirta, Mojokerto City

This study aims to determine the concentration of residual chlorine and net total Coliform distributed by Perumdam Maja Tirta Water Supply Company and received by consumers. Water samples were taken from 7 locations on the distribution network a residential area of Perumdam Maja Tirta, the first sample taken at the reservoir of the Water Treatment Plant of Perumdam Maja Tirta and six samples from consumers. The residual chlorine concentration data and Total Coliform were mapped using Surfer 16 Software, producing an iso concentration map in the form of contour lines. The results showed that the concentration of residual water chlorine received by consumers of Perumdam Maja Tirta Mojokerto City ranged from &lt;LoQ 0.1 mg/L to 0.3 mg/L. The further away from the IPA, the concentration of residual water chlorine decreased. Total Coliform parameters ranged from &lt;1.8 – 240/100 ml in water received by consumers of Perumdam Maja Tirta Mojokerto City. There is a need to improve the quality of water services to consumers regarding water treatment and distribution.

Double-diffusive natural convection energy transfer in magnetically influenced Casson fluid flow in trapezoidal enclosure with fillets

The prime motive of this disquisition is to deal with mathematical analysis of natural convection energy transport driven by combined buoyancy effects of thermal and solutal diffusion in a trapezoidal enclosure. Casson fluid rheological constitutive model depicting attributes of viscoelastic liquids is envisioned. The influence of the inclined magnetic field governed by Lorentz field law is also considered. To raise the essence of the presently used computational and physical domain, an innovative structural design called fillet is capitalized at the edges of the cavity. A numerical solution of the leading formulation is sought through Galerkin finite element discretization. Momentum, temperature, and concentration equations are interpolated by quadratic polynomials, whereas pressure distribution is approximated by a linear interpolating function. Domain discretized version is evaluated in view of triangular and rectangular elements. Newton's scheme is employed to resolve the non-linearly discretized system and a matrix factorization-based non-linear solver renowned as PARADISO is used. Validation of results is ascertained by forming an agreement with existing studies. Streamlines, isothermal and iso concentration contours patterns are portrayed to evaluate variation inflow distributions. Kinetic energy, local heat, and mass fluxes are also divulged in graphical and tabular formats.

Effects of Lysine-Lysine Dipeptide on Serum Amino Acid Profiles, Intestinal Morphology, and Microbiome in Suckling Piglets.

AimsSmall peptides are more energy-saving and efficiently absorbed compared to amino acids. Our study aimed to evaluate the effect of the Lys-Lys dipeptide on the improvement of growth performance, amino acid metabolism, and gut development in suckling piglets.Methods and ResultsTwenty-eight newborn suckling piglets were orally administrated with 0.1%, 1%, and 5% Lys-Lys dipeptide for 21 days. Our results showed that the Lys-Lys dipeptide has no significant effect on growth performance and intestinal morphology compared with the control group. We also found that the 1% Lys-Lys dipeptide significantly increased the concentrations of serum Lys, Thr, Phe, and Pro while decreasing Cys compared to the control group. Similarly, the 5% Lys-Lys dipeptide markedly increased the concentrations of serum Lys, Iso, Thr, Asp, Glu, and Pro compared to the control group. Moreover, the Lys-Lys dipeptide downregulated the expression of jejunal Slc7a1, Slc7a2, and Slc15a1 and ileal Slc7a2. Additionally, the Lys-Lys dipeptide decreased the microbiota richness indices and relative abundance of Bacteroidales.ConclusionIn this study, we found that the Lys-Lys dipeptide contributes to the metabolism of amino acids but failed to affect the growth performance of piglets. Additionally, the Lys-Lys dipeptide decreased the relative abundance of Bacteroidales. These results provide a theoretical for the future application and research of Lys-Lys dipeptide in intestinal development of suckling piglets.

Fluorescence spectroscopy and molecular docking analysis of the binding of Lactobacillus acidophilus GIM1.208 β-glucosidase with quercetin glycosides

Lactobacillus acidophilus is an important probiotic. The β-glucosidase produced by L. acidophilus GIM1.208 can transform quercetin glycosides of Rosa roxburghii Tratt to release quercetin and improve the functional activity of raw materials. Understanding the interaction and the characteristics of the two will lay a theoretical foundation for the site-directed transformation and functional application of the catalytic active site of enzymes. In our study, using the heterologously expressed and highly stable, purified L. acidophilus GIM1.208 BGL as the strain, the representative quercetin in β-glucosidase and Rosa roxburghii Tratt was preliminarily predicted and explored using ultraviolet-visible absorption spectroscopy. Fluorescence spectroscopy combined with molecular docking was used to determine the interaction characteristics of the glycoside substrates, rutin (Rut) and isoquercitrin (Iso). Results from molecular docking showed that Asp159, Arg56, Iso294, Phe292, and Gly25 were the main residues of β-glucosidase and Rut. Arg56 was found to be the most crucial residue of β-glucosidase and isoquercitrin; the interaction between Rut and Iso and β-glucosidase was mainly driven by hydrogen bonding. The combined free energy of β-glucosidase and Iso was found to be -182.10 kcal/mol, while that of β-glucosidase and Rut was -32.37 kcal/mol. The results of fluorescence spectroscopy showed that the fluorescence intensity of β-glucosidase decreased with an increase in Rut and Iso concentrations. This interaction made β-glucosidase quench endogenous fluorescence, which was static quenching. The binding constants of Rut and Iso with β-glucosidase were determined to be 0.50×107 and 0.31×107 L/mol, respectively, indicating that rutin had a stronger affinity when interacting with β-glucosidase. These findings were consistent with our prediction results determined using molecular docking studies.

Macro-, meso- and microplastic debris in the beaches of Tuticorin district, Southeast coast of India

This study investigates the mean concentration and characteristics of macroplastics (>2.5 cm), mesoplastics (5 mm - 2.5 cm) and microplastics (<5 mm) on eight sandy beaches along the shoreline of Tuticorin, Tamil Nadu. Prevalence of plastic litters varies among the study sites depending on the intensity of fishing and other human activities. Mean concentrations of macroplastics (1.38 ± 78 to 6.16 ± 94 items/m2), mesoplastics (2 ± 0.8 to 17 ± 0.11 items/m2) and microplastics (25 ± 1.58 to 83 ± 49 items/m2) were estimated in respect of the polymers composing them namely PE, PP, PET, NY, PS and PVC, of which PE is the most predominant polymer. At Sites 1, 2, 3, 5 and 8 there is correlation between the intensity of fishing activity and the concentrations of macro- (p = 0.02) and microplastics (p = 0.03). Sites 4, 6 and 7 there is correlation established between the degree of recreational activity and the concentrations of meso- (p = 0.02) and microplastics (p = 0.01).

Primary and secondary reactions in the synthesis of hydrocarbons from dimethyl ether over a Pd-Zn-HZSM-5/Al2O3 catalyst

The synthesis of hydrocarbons from dimethyl ether in an atmosphere of synthesis gas, over a Pd-Zn-HZSM-5/Al2O3 catalyst, under flow-circulation conditions, at a pressure of 10.0 MPa, was investigated. It was found that the resulting hydrocarbon fractions were characterized by a low concentration of aromatic compounds and high concentrations of iso- and cycloalkanes. The dependence of the yields of the main groups of products and individual components on the contact time was explained in terms of the occurrence of methylation, oligomerization, hydrogenation, H-transfer, and hydroisomerization reactions. The role of Pd and Zn in the formation of the products was evaluated.

Tolerance and Recovery of Ultralow-Loaded Platinum Anode Electrodes upon Carbon Monoxide and Hydrogen Sulfide Exposure.

The effects of carbon monoxide (CO) and hydrogen sulfide (H2S) in concentrations close to their respective limits in the Hydrogen Quality Standard ISO 14687-2:2012 on the performance of proton exchange membrane fuel cells (PEMFCs) with ultralow-loaded platinum anode catalyst layers (CLs) were investigated. The anodic loadings were 50, 25, and 15 µg/cm2, which represent the current state-of-the-art, target, and stretch target, respectively, for future automotive PEMFCs. Additionally, the effect of shut-down and start-up (SD/SU) processes on recovery from sulfur poisoning was investigated. CO at an ISO concentration of 0.2 ppm caused severe voltage losses of ~40–50% for ultralow-loaded anode CLs. When H2S was in the fuel, these anode CLs exhibited both a nonlinear decrease in tolerance toward sulfur and an improved self-recovery during shut-down and start-up (SD/SU) processes. This observation was hypothesized to have resulted from the decrease in the ratio between CL thickness and geometric cell area, as interfacial effects of water in the pores increasingly impacted the performance of ultrathin CLs. The results indicate that during the next discussions on the Hydrogen Quality Standard, a reduction in the CO limit could be a reasonable alternative considering future PEMFC anodic loadings, while the H2S limit might not require modification.

The effects of molecular structure and functional group of a rodlike Schiff base mesogen on blue phase stabilization in a chiral system

A wider blue phase (BP) range can be induced easily when two difluoro substituted and racemic rodlike Schiff base mesogens are doped with the appropriate concentration of chiral dopants S811 or ISO(6OBA)2.

Atomic layout of an orthodontic titanium mini-implant in human tissue: Insights into the possible mechanisms during osseointegration

To evaluate nanoscale molecular interactions in the interface between human bone and orthodontic titanium implants. An orthodontic implant (sandblasted with large grit and with an acid-etched surface treated with Ti6A14V alloy) retrieved from the mandible of human after 2 months of healing was used to analyze the molecular interactive mechanism between the implant and the surrounding bone tissue. To preserve the natural state of the sample as much as possible, cryofixation and scanning electron microscope/focused ion beam milling without any chemical treatment were used during sample preparation. Atom probe tomography was used to investigate the chemical composition and structure at the interface between the implant and human bone tissue. Three-dimensional (3D) reconstruction of the whole sample revealed a 20 × 50-nm2 plate-like bony element diffusion layer in the sample. The iso concentration analysis of the diffusion layer indicated that the bony element, calcium, and the implant element, titanium oxide, were interspersed with each other. Detailed ionic distribution was illustrated by 3D reconstruction with partial region of interest and one-dimensional concentration profiles of the implant-bone interface. The study results advance nanoscale understanding of osseointegration and suggest a potential nanostructure for increasing bond strength of biomaterials to bone.

Ca2+-dependent calcineurin/NFAT signaling in β-adrenergic-induced cardiac hypertrophy.

Ca2+ is an important mediator in the β-adrenergic-induced cardiac hypertrophy. The β-adrenergic stimulation alters the Ca2+ transient characteristics including its oscillation frequency, diastolic and systolic levels which lead to the CaN activation and subsequent NFAT-dependent hypertrophic genes transcription. Moreover, β-adrenergic-induced alterations in PKA and GSK3β kinase activities in both the cytosol and the nucleus regulate NFAT nuclear translocation and contribute in its hypertrophic response. Due to the complex nature of CaN/NFAT signaling in cardiac cells, we use a computational approach to investigate the β-adrenergic-induced CaN/NFAT activation in the cardiac myocytes. The presented model predicts well the main physiological characteristics of CaN/NFAT signaling in accordance with the experimental observations. The presented model establishes the previous experimental and mathematical results on the principal role of Ca2+ oscillation frequency in the CaN/NFAT signaling and shows that increase in Ca2+ oscillation frequency enhances CaN activity and its sensitivity to low ISO concentrations. The model illustrates that in addition to the known ISO effect on Ca2+ transient amplitude, ISO-induced alterations in Ca2+ oscillation frequency, PKA and GSK3β kinase activities also greatly affect the β-adrenergic-induced NFAT activity. We also found that PKA has both pro-hypertrophic and anti-hypertrophic effects on NFAT activation and is the main kinase in ISO-induced NFAT activation.

Isoimperatorin ameliorates osteoarthritis by downregulating the mammalian target of rapamycin C1 signaling pathway.

Osteoarthritis (OA) is the most common disease of the joints, and is characterized by the breakdown of cartilage and degradation of the extracellular matrix. OA causes a high level of patient suffering and incurs large societal costs; however, the current strategies for treating OA are restricted due to limited understanding of the underlying molecular and cellular mechanisms. In the present study, the beneficial effects of isoimperatorin (Iso) were investigated using an experimental mouse model of OA, and its mechanism of action on primary chondrocytes was elucidated. Destabilization of the medial meniscus was performed on 8‑week‑old male mice to induce OA in the knees. Iso (500mg/g/day) was intragastrically administered for 4weeks. Degeneration of articular cartilage was assessed by histology using the Osteoarthritis Research Society International scoring system. The expression of matrix metalloproteinase (MMP)13, Runt‑related transcription factor (Runx)2, type X collagen (Col X) and vascular endothelial growth factor (VEGF) in the knee joints was examined by immunohistochemistry. Invitro, murine primary chondrocytes were treated with various concentrations of Iso, followed by 10ng/ml interleukin‑1. The mRNA expression levels of MMP13, Runx2, Col X and VEGF were determined by reverse transcription‑quantitative polymerase chain reaction. The levels of autophagy and mammalian target of rapamycin (mTOR) signaling were determined by western blotting. Iso significantly ameliorated the severity of articular cartilage degradation in mice with experimental OA. The expression levels of MMP13, Runx2, Col X and VEGF were reduced in Iso‑treated mice. In murine primary chondrocytes, Iso also reduced MMP13, Runx2, Col X and VEGF expression, and activated autophagy by downregulating the mTOR complex1 (mTORC1) signaling pathway. Therefore, the results of the present study demonstrated that Iso ameliorates OA‑induced pathological alterations by delaying chondrocyte deterioration, activating autophagy and inhibiting mTORC1, which suggests that Iso may have therapeutic potential for attenuating articular cartilage degradation and treating OA.

Influence of cell confluence on the cAMP signalling pathway in vascular smooth muscle cells

The influence of cell confluence on the β-adrenoceptor (β-AR)/cAMP/phosphodiesterase (PDE) pathway was investigated in cultured rat aortic smooth muscle cells (RASMCs).Cells were plated either at low density (LD: 3·103cells/cm2) or high density (HD: 3·104cells/cm2) corresponding to non-confluent or confluent cells, respectively, on the day of experiment.β-AR-stimulated cAMP was monitored in real-time using the fluorescence resonance energy transfer (FRET)-based cAMP sensor, Epac2-camps. A brief application (15s) of the β-AR agonist isoprenaline (Iso) induced a typical transient FRET signal, reflecting cAMP production followed by its rapid degradation. The amplitude of this response, which increased with the concentration of Iso (10 or 100nM), was higher in HD than in LD cells, whatever the Iso concentration used. However, activation of adenylyl cyclase by L-858051 (100μM) induced a similar saturating response in both LD and HD cells. A β1-AR antagonist (CGP 20712A, 100nM) reduced the Iso (100nM) response in HD but not LD cells, whereas a β2-AR antagonist (ICI 118,551, 5nM) reduced this response in HD cells and almost abolished it in LD cells. Competitive [125I]-ICYP binding experiments with betaxolol, a β-AR ligand, identified two binding sites in HD cells, corresponding to β1- and β2-ARs with a proportion of 11% and 89%, respectively, but only one binding site in LD cells, corresponding to β2-ARs. Total cAMP-PDE activity (assessed by a radioenzymatic assay) was increased in HD cells compared to LD cells. This increase was associated with a rise in mRNA expression of five cAMP-PDEs subtypes (PDE1A, 3A, 4A, 4B and 7B) in HD cells, and a decrease in basal [cAMP]i (assessed by an EIA assay). PDE4 inhibition with Ro-20-1724 (10μM) strongly prolonged the Iso response in LD and HD cells, whereas PDE3 inhibition with cilostamide (1μM) slightly prolonged Iso response only in LD cells. Interestingly, inhibition of PDE4 unmasked an effect of PDE3 in HD cells.Our results show that in cultured RASMCs, the β-AR/cAMP/PDE signalling pathway is substantially modulated by the cell density. In HD cells, Iso response involves both β1- and β2-AR stimulation and is mainly controlled by PDE4, PDE3 being recruited only after PDE4 inhibition. In LD cells, Iso response involves only β2-AR stimulation and is controlled by PDE4 and to a lower degree by PDE3. This low density state is associated with an absence of membrane expression of the β1-AR, a lower cAMP-PDE activity and a higher basal [cAMP]i. This study highlights the critical role of the cellular environment in controlling the vascular β-AR signalling.

Apoptotic activity of isoespintanol derivatives in human polymorphonuclear cells

Background: Inflammation is a complex physiopathologic response to different stimuli. Recently, some pharmacological strategies have been proposed that could be used for resolution of inflammation by enhancing apoptosis of inflammatory cells. Objectives: To study in vitro apoptotic activity of isoespintanol [ISO] and of two semi-synthetic derivatives, bromide isoespintanol [BrI] and demethylated isoespintanol [DMI], in human polymorphonuclear (PMN) cells. Methods: PMN were exposed to the different concentrations of ISO, BrI and DMI for 30 min in phosphate-buffered saline pH 7.4 containing 1 mg/mL glucose, 0.4 mM Mg2+, and 1.20 mM Ca2+. Viability was assessed by dimethylthiazol diphenyl tetrazolium bromide (MTT). To distinguish between the two modes of cell death, apoptosis and necrosis, we examined differences in morphological and biochemical changes of cells stained with annexin V- FITC (An) and/or propidium iodide (PI) using two different assays based on flow cytometry Results: The MTT assay revealed the ability of cells to reduce MTT salt to formazan. In the presence of BrI and DMI a significant concentration-dependent decrease of cell viability was observed. The annexin V- FITC binding assay showed a high proportion of apoptotic cells for those treated with BrI (An+/ PI-: 62.3 ± 8.2% vs. 2.1 ± 0.5% of control, P<0.05). The population of PMN treated with DMI produced the highest percentage (An+/IP+: 43.4 ± 5.2 % vs. 0.4 ± 0.3 % of control, P<0.05) of necrotic cells. Apoptotic nuclei were analyzed by PI staining. The cell population in the sub G0/G1 region represents cells with hypodiploidal DNA, an indicator of apoptosis. When cells were incubated with 50 and 100 μM of BrI, the cell population in the sub G0/G1 region increased, suggesting a dose-dependent increase in the population of apoptotic cells. The presence of the pan-inhibitor of caspases (Z-VAD-fmk) showed a significant reduction in cell population in the sub G0/G1 region, indicating less degradation of DNA. Conclusions: Bromide isoespintanol [BrI] induces an apoptotic process in PMN, mediated –at least in part– by activation of caspases, although this compound may probably act through other caspase-independent mechanisms as well.

Effect of lipid supplementation on milk odd- and branched-chain fatty acids in dairy cows

Eight ruminally fistulated, multiparous Holstein cows were arranged in a double 4×4 Latin square with 14-d periods to investigate the effects of lipid supplementation on performance, rumen parameters, the milk odd- and branched-chain fatty acid (OBCFA) profile, and the relationships between milk OBCFA and rumen parameters. Lipid supplementation is known to inhibit microbial growth in the rumen, decrease de novo microbial fatty acid synthesis, and increase the uptake of circulating fatty acids by the mammary gland; treatments were selected to isolate these effects on the milk OBCFA profile. The 4 treatments were (1) a lipid-free emulsion medium infused in the rumen (CTL), (2) soybean oil as a source of polyunsaturated fatty acids infused in the rumen (RSO), (3) saturated fatty acids (38% 16:0, 40% 18:0) infused in the rumen (RSF), and (4) saturated fatty acids infused in the abomasum (ASF). Fat supplements were provided continuously as emulsions at a rate of 450g/d. Preplanned contrasts compared CTL to RSO, RSO to RSF, and RSF to ASF. Infusing RSO slightly decreased ruminal pH, but did not affect volatile fatty acids profile and milk fat concentration as compared with CTL. The yields of energy-corrected milk, fat, and protein were greater with RSF compared with RSO. The concentration of odd-chain fatty acids was decreased by RSO, whereas even-chain iso fatty acids were not affected. Milk fat concentration of 17:0 + cis-9 17:1 was higher for RSF than for RSO, due to the saturated fatty acids supplement containing 2% 17:0 + cis-9 17:1. Limited differences were observed in the milk OBCFA profile between RSF and ASF. A multiple regression analysis yielded the following equation for predicting rumen pH based on milk fatty acids: pH=6.24 – (0.56×4:0) + (1.67 × iso 14:0) + (4.22 × iso 15:0) + (9.41×22:0). Rumen propionate concentration was negatively correlated with milk fat concentration of iso 14:0 and positively correlated with milk 15:0, whereas the acetate-to-propionate ratio gave the opposite correlations with milk iso 14:0 and 15:0. Milk fat concentration of 17:0 + cis-9 17:1 was not related to rumen propionate or to acetate-to-propionate ratio, due to the presence of 17:0 and cis-9 17:1 in the saturated fatty acids supplement. The results suggest that although lipid supplementation can affect the profile of milk OBCFA, the promise remains of using these milk fatty acids to evaluate rumen function.

Differential Regulation of Cytoplasmic and Nuclear PKA Activities by β1- and β2-Adrenoceptors in Adult Cardiac Myocytes

Functional differences between β1-adrenoceptors (β1-ARs) and β2-ARs in the regulation of cardiac excitation-contraction coupling have been widely studied. Much less is known regarding differences between β1- and β2-ARs in the regulation of gene expression and the mechanisms involved. Here, we studied the mechanisms regulating nuclear versus bulk cytoplasmic PKA activation by β1- and β2-ARs and the functional consequences on gene expression.PKA activity was measured using targeted FRET-based A-kinase activity reporters. For specific β1- or β2-AR stimulation, isoprenaline (Iso) was used in combination with the β2-AR antagonist ICI118551 or with the β1-AR antagonist CGP20712A, respectively. At all Iso concentrations tested, β1-AR stimulation was more efficient than β2-AR stimulation to increase cytoplasmic and nuclear PKA activities. For a similar activation of cytoplasmic PKA, nuclear PKA activity was 3-fold higher with β1-AR than with β2-AR stimulation. Inhibition of Gi protein, caveolae disruption, and inhibition of GRK2-mediated desensitization potentiated β2-AR-induced cytoplasmic but not nuclear PKA activity. PDE4 inhibition strongly potentiated cytoplasmic and nuclear PKA responses to both β1- and β2-AR stimulation. In contrast, PDE3 inhibition had no significant effect on β1-AR induced PKA activation in both compartments, while it increased cytoplasmic but not nuclear PKA activity upon β2-AR stimulation. Downregulation of mAKAP, a nuclear envelope associated scaffold protein, decreased β1-AR stimulation of nuclear PKA activity. Consistently, β1-ARs but not β2-ARs were able to induce the expression of the PKA-regulated pro-apoptotic gene, ICER. These results show that i) β1- and β2-ARs differentially regulate cytoplasmic versus nuclear PKA activities, ii) nuclear PKA activation can be dissociated from bulk cytoplasmic PKA activity upon β2-AR stimulation; and iii) PDE4 and mAKAP are critical components of nuclear PKA signalling.

Isosakuranetin, a 4'-O-methylated flavonoid, stimulates melanogenesis in B16BL6 murine melanoma cells.

The beneficial effects of 4'-O-methylated flavonoids on induction of melanogenesis are well established. Here, we report the effect of isosakuranetin (Iso) on melanogenesis in B16BL6 melanoma cells and an analysis of the signaling pathways involved in this activity. B16BL6 melanoma cells were treated with several concentrations of Iso and melanin content was measured. Activation and expression of factors involved in melanogenesis were assessed via western blotting. Iso (15 and 30μmol/L) strongly stimulated melanogenesis in a dose-dependent manner. Iso increased tyrosinase activity and up-regulated tyrosinase (Tyr), tyrosinase related protein 1 (TRP1), and tyrosinase related protein 2 (TRP2) in a time-dependent manner. Iso decreased B16 cell proliferation at a concentration above 45μmol/L, and had no effect on cell viability as revealed by MTT and trypan blue assays. Iso up-regulated expression of microphthalmia transcription factor (MITF), with a maximum effect after 12h. H89, a specific inhibitor of PKA, showed that MITF up-regulation is mediated through PKA/CREB activation. Furthermore, Iso decreased phosphorylation of MITF at Ser73 after 24h and 48h of exposure, activating MITF and leading to up-regulation of Tyr, TRP1, and TRP2. Iso inhibited phosphorylation and activation of ERK1/2 after 12h, while no significant effects on p38 and JNK phosphorylation were observed. Iso inhibited AKT phosphorylation and led to activation of GSK3β. Iso stimulates melanogenesis in B16 melanoma cells via up-regulation of MITF. Furthermore, Iso-induced inhibition of ERK1/2 and PI3K/AKT signaling pathways activate MITF and subsequent expression of Tyr, TRP1, and TRP2.