Nutritional and insulin regulation of fatty acid synthetase and leptin gene expression through ADD1/SREBP1.

The ability to regulate specific genes of energy metabolism in response to fasting and feeding is an important adaptation allowing survival of intermittent food supplies. However, little is known about transcription factors involved in such responses in higher organisms. We show here that gene expression in adipose tissue for adipocyte determination differentiation dependent factor (ADD) 1/sterol regulatory element binding protein (SREBP) 1, a basic-helix-loop-helix protein that has a dual DNA-binding specificity, is reduced dramatically upon fasting and elevated upon refeeding; this parallels closely the regulation of two adipose cell genes that are crucial in energy homeostasis, fatty acid synthetase (FAS) and leptin. This elevation of ADD1/SREBP1, leptin, and FAS that is induced by feeding in vivo is mimicked by exposure of cultured adipocytes to insulin, the classic hormone of the fed state. We also show that the promoters for both leptin and FAS are transactivated by ADD1/SREBP1. A mutation in the basic domain of ADD1/SREBP1 that allows E-box binding but destroys sterol regulatory element-1 binding prevents leptin gene transactivation but has no effect on the increase in FAS promoter function. Molecular dissection of the FAS promoter shows that most if not all of this action of ADD1/SREBP1 is through an E-box motif at -64 to -59, contained with a sequence identified previously as the major insulin response element of this gene. These results indicate that ADD1/SREBP1 is a key transcription factor linking changes in nutritional status and insulin levels to the expression of certain genes that regulate systemic energy metabolism.

Transfer insensitive labeling technique (TILT): application to multislice functional perfusion imaging.

Cerebral blood flow can be studied in a multislice mode with a recently proposed perfusion sequence using inversion of water spins as an endogenous tracer without magnetization transfer artifacts. The magnetization transfer insensitive labeling technique (TILT) has been used for mapping blood flow changes at a microvascular level under motor activation in a multislice mode. In TILT, perfusion mapping is achieved by subtraction of a perfusion-sensitized image from a control image. Perfusion weighting is accomplished by proximal blood labeling using two 90 degrees radiofrequency excitation pulses. For control preparation the labeling pulses are modified such that they have no net effect on blood water magnetization. The percentage of blood flow change, as well as its spatial extent, has been studied in single and multislice modes with varying delays between labeling and imaging. The average perfusion signal change due to activation was 36.9 +/- 9.1% in the single-slice experiments and 38.1 +/- 7.9% in the multislice experiments. The volume of activated brain areas amounted to 1.51 +/- 0.95 cm3 in the contralateral primary motor (M1) area, 0.90 +/- 0.72 cc in the ipsilateral M1 area, 1.27 +/- 0.39 cm3 in the contralateral and 1.42 +/- 0.75 cm3 in the ipsilateral premotor areas, and 0.71 +/- 0.19 cm3 in the supplementary motor area.

Stable one-step technetium-99m labeling of His-tagged recombinant proteins with a novel Tc(I)-carbonyl complex.

We have developed a technetium labeling technology based on a new organometallic chemistry, which involves simple mixing of the novel reagent, a 99m Tc(I)-carbonyl compound, with a His-tagged recombinant protein. This method obviates the labeling of unpaired engineered cysteines, which frequently create problems in large-scale expression and storage of disulfide-containing proteins. In this study, we labeled antibody single-chain Fv fragments to high specific activities (90 mCi/mg), and the label was very stable to serum and all other challenges tested. The pharmacokinetic characteristics were indistinguishable from iodinated scFv fragments, and thus scFV fragments labeled by the new method will be suitable for biodistribution studies. This novel labeling method should be applicable not only to diagnostic imaging with 99mTc, but also to radioimmunotherapy approaches with 186/188 Re, and its use can be easily extended to almost any recombinant protein or synthetic peptide.

Escarres et dénutrition: dépistage et évaluation de l'état nutritionnel [Pressure ulcers and undernutrition: screening and assessment of nutritional status]

Acquired pressure ulcer is associated with significant human, economic and functional consequences. Its prevalence varies between 3 and 23% in a community hospital and between 7 and 54% in an elderly home residency. Pressure ulcer healing is a complex process which involves numerous cellular and molecular mechanisms. An altered nutritional status is a contributing factor in the development of pressure ulcers and the delay in pressure ulcer healing. The key to management of undernutrition is screening and early intervention. According to the gravity of undernutrition, various degrees of intervention will be required. Systematic oral supplementation with various nutrients may provide benefit in the prevention of pressure ulcers, but further studies have to be completed in human subjects prior to being recommended for the treatment of pressure ulcers.

Liming influence on soil chemical properties, nutritional status and yield of alfalfa grown in acid soil

Alfalfa is an important forage crop with high nutritive value, although highly susceptible to soil acidity. Liming is one of the most efficient and prevailing practices to correct soil acidity and improve alfalfa yield. The objective of this study was to evaluate response to liming of alfalfa grown in a greenhouse on a Typic Quartzipsamment soil. The treatments consisted of four lime rates (0, 3.8, 6.6 and 10.3 Mg ha-1) and two cuts. Alfalfa dry matter increased quadratically with increasing lime rates. In general, dry matter yield was maximized by a lime rate of 8.0 Mg ha-1. Except for the control, the dry matter nutrient contents in the treatments were adequate. The positive linear correlation between root and nodule dry matter with lime rates indicated improvement of these plant traits with decreasing soil acidity. The soil acidity indices pH, base saturation, Ca2+ concentration, Mg2+ concentration, and H + Al were relevant factors in the assessment of alfalfa yield. The magnitude of influence of these soil acidity indices on yield as determined by the coefficient of determination (R²) varied and decreased in the order: base saturation, H + Al, pH, Ca and Mg concentrations. Optimum values of selected soil chemical properties were defined for maximum shoot dry matter; these values can serve as a guideline for alfalfa liming to improve the yield of this forage on acid soils.

DRIS formulas for evaluation of nutritional status of cupuaçu trees

DRIS, an Diagnosis and Recommendation Integrated System, is a tool to evaluate the nutritional status of plants. Different DRIS formulas have been proposed to improve the efficiency of the crop nutrition diagnoses. The objective of this study was to compare the nutritional diagnosis of the formulas of Beaufils (1973), of Jones (1981) and of Elwali and Gascho (1984), based on the degree of agreement in commercial orchards of Theobrama grandiflorum trees. Leaf samples of 5 to 18 year-old cupuaçu trees were collected from 153 commercial orchards in agroforestry and monoculture systems in the state of Rondonia, Brazil. Bivariate relationships between nutrition concentrations in healthy trees were used to calculate DRIS norms. DRIS indices were calculated based on the different formulas and interpreted by the Potential Fertilizer Response method, in five categories. The DRIS norms, DRIS index calculations and their interpretations were developed using the DRIS Cupuaçu computer program (www.dris.com.br). The different DRIS formulas resulted in similar diagnoses with a degree of agreement of > 90% for the nutrients N, P, K, Ca, and Mg.

Soil composition and nutritional status of apple as affected by long-term application of gypsum

Gypsum does not affect the soil negative charges and maintains sulfate in the soil solution, making it one of the cheapest products to increase Ca activity in soil solution, especially in the deeper soil layers. Higher Ca levels in the soil solution can increase the uptake of this nutrient by apple trees, reducing the risk of physiological disorders caused by Ca deficiency. This study assessed the effect of long-term gypsum application on some soil properties and on the chemical composition of leaves and fruits of an apple cultivar susceptible to fruit disorders associated with low Ca. The experiment was conducted in São Joaquim, in the South of Brazil, from 2001 to 2009. Gypsum rates of 0, 1.0, 2.0 and 3.0 t ha-1 were annually broadcast over the soil surface, without incorporation, in an apple orchard with cultivar ´Catarina´, planted in 1997. Gypsum application over eight consecutive years had no effect on soil exchangeable K and Al to a depth of 80 cm, but increased exchangeable Ca in the sampled layers (0-10, 10-20, 40-60 and 60-80 cm), while exchangeable Mg decreased only in the surface layer (0-20 cm). Gypsum did not affect the concentration of any nutrient in the fruits, including Ca. The same was verified in the leaves, except for Mg which decreased with increased gypsum rate. Despite increasing the availability of Ca in the soil profile to a depth of 80 cm, gypsum was not effective to increase the Ca content in leaves and fruits of an apple cultivar susceptible to Ca deficiency grown in an appropriately limed soil.

ANIBAL, stable isotope-based quantitative proteomics by aniline and benzoic acid labeling of amino and carboxylic groups

Identification and relative quantification of hundreds to thousands of proteins within complex biological samples have become realistic with the emergence of stable isotope labeling in combination with high throughput mass spectrometry. However, all current chemical approaches target a single amino acid functionality (most often lysine or cysteine) despite the fact that addressing two or more amino acid side chains would drastically increase quantifiable information as shown by in silico analysis in this study. Although the combination of existing approaches, e.g. ICAT with isotope-coded protein labeling, is analytically feasible, it implies high costs, and the combined application of two different chemistries (kits) may not be straightforward. Therefore, we describe here the development and validation of a new stable isotope-based quantitative proteomics approach, termed aniline benzoic acid labeling (ANIBAL), using a twin chemistry approach targeting two frequent amino acid functionalities, the carboxylic and amino groups. Two simple and inexpensive reagents, aniline and benzoic acid, in their (12)C and (13)C form with convenient mass peak spacing (6 Da) and without chromatographic discrimination or modification in fragmentation behavior, are used to modify carboxylic and amino groups at the protein level, resulting in an identical peptide bond-linked benzoyl modification for both reactions. The ANIBAL chemistry is simple and straightforward and is the first method that uses a (13)C-reagent for a general stable isotope labeling approach of carboxylic groups. In silico as well as in vitro analyses clearly revealed the increase in available quantifiable information using such a twin approach. ANIBAL was validated by means of model peptides and proteins with regard to the quality of the chemistry as well as the ionization behavior of the derivatized peptides. A milk fraction was used for dynamic range assessment of protein quantification, and a bacterial lysate was used for the evaluation of relative protein quantification in a complex sample in two different biological states

Hyperpolarized 13C lactate as a substrate for in vivo metabolic studies in skeletal muscle

Resting skeletal muscle has a preference for the oxidation of lipids compared to carbohydrates and a shift towards carbohydrate oxidation is observed with increasing exercise. Lactate is not only an end product in skeletal muscle but also an important metabolic intermediate for mitochondrial oxidation. Recent advances in hyperpolarized MRS allow the measurement of substrate metabolism in vivo in real time. The aim of this study was to investigate the use of hyperpolarized 13C lactate as a substrate for metabolic studies in skeletal muscle in vivo. Carbohydrate metabolism in healthy rat skeletal muscle at rest was studied in different nutritional states using hyperpolarized [1-13C]lactate, a substrate that can be injected at physiological concentrations and leaves other oxidative processes undisturbed. 13C label incorporation from lactate into bicarbonate in fed animals was observed within seconds but was absent after an overnight fast, representing inhibition of the metabolic flux through pyruvate dehydrogenase (PDH). A significant decrease in 13C labeling of alanine was observed comparing the fed and fasted group, and was attributed to a change in cellular alanine concentration and not a decrease in enzymatic flux through alanine transaminase. We conclude that hyperpolarized [1-13C]lactate can be used to study carbohydrate oxidation in resting skeletal muscle at physiological levels. The herein proposed method allows probing simultaneously both PDH activity and variations in alanine tissue concentration, which are associated with metabolic dysfunctions. A simple alteration of the nutritional state demonstrated that the observed pyruvate, alanine, and bicarbonate signals are indeed sensitive markers to probe metabolic changes in vivo.

Renal arteries: navigator-gated balanced fast field-echo projection MR angiography with aortic spin labeling: initial experience.

A cardiac-triggered free-breathing three-dimensional balanced fast field-echo projection magnetic resonance (MR) angiographic sequence with a two-dimensional pencil-beam aortic labeling pulse was developed for the renal arteries. For data acquisition during free breathing in eight healthy adults and seven consecutive patients with renal artery disease, real-time navigator technology was implemented. This technique allows high-spatial-resolution and high-contrast renal MR angiography and visualization of renal artery stenosis without exogenous contrast agent or breath hold. Initial promising results warrant larger clinical studies.