Proteomic profiling of neuromas reveals alterations in protein composition and local protein synthesis in hyper-excitable nerves

Neuropathic pain may arise following peripheral nerve injury though the molecular mechanisms associated with this are unclear. We used proteomic profiling to examine changes in protein expression associated with the formation of hyper-excitable neuromas derived from rodent saphenous nerves. A two-dimensional difference gel electrophoresis ( 2D-DIGE) profiling strategy was employed to examine protein expression changes between developing neuromas and normal nerves in whole tissue lysates. We found around 200 proteins which displayed a > 1.75-fold change in expression between neuroma and normal nerve and identified 55 of these proteins using mass spectrometry. We also used immunoblotting to examine the expression of low-abundance ion channels Nav1.3, Nav1.8 and calcium channel alpha 2 delta-1 subunit in this model, since they have previously been implicated in neuronal hyperexcitability associated with neuropathic pain. Finally, S(35)methionine in vitro labelling of neuroma and control samples was used to demonstrate local protein synthesis of neuron-specific genes. A number of cytoskeletal proteins, enzymes and proteins associated with oxidative stress were up-regulated in neuromas, whilst overall levels of voltage-gated ion channel proteins were unaffected. We conclude that altered mRNA levels reported in the somata of damaged DRG neurons do not necessarily reflect levels of altered proteins in hyper-excitable damaged nerve endings. An altered repertoire of protein expression, local protein synthesis and topological re-arrangements of ion channels may all play important roles in neuroma hyper-excitability.

Fatty acid compositions of inositol and choline phospholipids of breast tumours and normal breast tissue

The fatty acid compositions of the -choline and -inositol phospholipids of breast tumours of women undergoing surgery for treatment of breast disease (malignant n = 12; benign n = 10) and normal breast tissue of women undergoing breast reduction surgery (n = 6) were determined. The fatty acid compositions of erythrocyte phospholipids were also determined in the same subjects and in an additional number of normal healthy volunteers (n = 16). Levels of oleic acid were lower in both phospholipid fractions of erythrocytes of women with breast disease and in the phosphatidylcholine fraction of breast tumours compared with normal breast tissue. Significantly higher levels of linoleic acid were found in erythrocytes of tumour-bearing subjects and a similar trend was evident in the phosphatidylcholine fraction of tumour compared with normal breast tissues. Conversely, lower levels of two of the products of linoleic acid chain elongation and desaturation, dihomogamma-linolenic and arachidonic acids, were found in the erythrocyte phospholipids of tumour-bearing subjects and in the choline phospholipids of breast tumour tissues. These data suggest that in women with breast disease, there may be inhibition of 6-desaturase, and enhanced activity of 9-desaturase, enzymes which play an important role in determining membrane phospholipid fatty acid composition. This pattern of altered fatty acid composition characteristic of erythrocyte phospholipids of tumour-bearing subjects and phosphatidylcholine of breast tumour tissue was less evident in the case of the breast tumour phosphatidylinositol in which differences other than those described were seen.

Differential effects of two fermentable carbohydrates on central appetite regulation and body composition

BACKGROUND: Obesity is rising at an alarming rate globally. Different fermentable carbohydrates have been shown to reduce obesity. The aim of the present study was to investigate if two different fermentable carbohydrates (inulin and b-glucan) exert similar effects on body composition and central appetite regulation in high fat fed mice. METHODOLOGY/PRINCIPAL FINDINGS: Thirty six C57BL/6 male mice were randomized and maintained for 8 weeks on a high fat diet containing 0% (w/w) fermentable carbohydrate, 10% (w/w) inulin or 10% (w/w) b-glucan individually. Fecal and cecal microbial changes were measured using fluorescent in situ hybridization, fecal metabolic profiling was obtained by proton nuclear magnetic resonance (1H NMR), colonic short chain fatty acids were measured by gas chromatography, body composition and hypothalamic neuronal activation were measured using magnetic resonance imaging (MRI) and manganese enhanced MRI (MEMRI), respectively, PYY (peptide YY) concentration was determined by radioimmunoassay, adipocyte cell size and number were also measured. Both inulin and b-glucan fed groups revealed significantly lower cumulative body weight gain compared with high fat controls. Energy intake was significantly lower in b-glucan than inulin fed mice, with the latter having the greatest effect on total adipose tissue content. Both groups also showed an increase in the numbers of Bifidobacterium and Lactobacillus-Enterococcus in cecal contents as well as feces. b- glucan appeared to have marked effects on suppressing MEMRI associated neuronal signals in the arcuate nucleus, ventromedial hypothalamus, paraventricular nucleus, periventricular nucleus and the nucleus of the tractus solitarius, suggesting a satiated state. CONCLUSIONS/SIGNIFICANCE: Although both fermentable carbohydrates are protective against increased body weight gain, the lower body fat content induced by inulin may be metabolically advantageous. b-glucan appears to suppress neuronal activity in the hypothalamic appetite centers. Differential effects of fermentable carbohydrates open new possibilities for nutritionally targeting appetite regulation and body composition.

New self-assembling multifunctional templates for the biofabrication and controlled self-release of cultured tissue

The need to source live human tissues for research and clinical applications has been a major driving force for the development of new biomaterials. Ideally, these should elicit the formation of scaffold-free tissues with native-like structure and composition. In this study, we describe a biologically interactive coating that combines the fabrication and subsequent self-release of live purposeful tissues using template–cell–environment feedback. This smart coating was formed from a self-assembling peptide amphiphile comprising a proteasecleavable sequence contiguous with a cell attachment and signaling motif. This multifunctional material was subsequently used not only to instruct human corneal or skin fibroblasts to adhere and deposit discreet multiple layers of native extracellular matrix but also to govern their own self-directed release from the template solely through the action of endogenous metalloproteases. Tissues recovered through this physiologically relevant process were carrier-free and structurally and phenotypically equivalent to their natural counterparts. This technology contributes to a new paradigm in regenerative medicine, whereby materials are able to actively direct and respond to cell behavior. The novel application of such materials as a coating capable of directing the formation and detachment of complex tissues solely under physiological conditions can have broad use for fundamental research and in future cell and tissue therapies.

Low-grade inflammation, diet composition and health: current research evidence and its translation

The importance of chronic low-grade inflammation in the pathology of numerous age-related chronic conditions is now clear. An unresolved inflammatory response is likely to be involved from the early stages of disease development. The present position paper is the most recent in a series produced by the International Life Sciences Institute's European Branch (ILSI Europe). It is co-authored by the speakers from a 2013 workshop led by the Obesity and Diabetes Task Force entitled ‘Low-grade inflammation, a high-grade challenge: biomarkers and modulation by dietary strategies’. The latest research in the areas of acute and chronic inflammation and cardiometabolic, gut and cognitive health is presented along with the cellular and molecular mechanisms underlying inflammation–health/disease associations. The evidence relating diet composition and early-life nutrition to inflammatory status is reviewed. Human epidemiological and intervention data are thus far heavily reliant on the measurement of inflammatory markers in the circulation, and in particular cytokines in the fasting state, which are recognised as an insensitive and highly variable index of tissue inflammation. Potential novel kinetic and integrated approaches to capture inflammatory status in humans are discussed. Such approaches are likely to provide a more discriminating means of quantifying inflammation–health/disease associations, and the ability of diet to positively modulate inflammation and provide the much needed evidence to develop research portfolios that will inform new product development and associated health claims.

Osteoconductivity of modified fluorcanasite glass-ceramics for bone tissue augmentation and repair

Modified fluorcanasite glasses were fabricated by either altering the molar ratios of Na(2)O and CaO or by adding P(2)O(5) to the parent stoichiometric glass compositions. Glasses were converted to glass-ceramics by a controlled two-stage heat treatment process. Rods (2 mm x 4 mm) were produced using the conventional lost-wax casting technique. Osteoconductive 45S5 bioglass was used as a reference material. Biocompatibility and osteoconductivity were investigated by implantation into healing defects (2 mm) in the midshaft of rabbit femora. Tissue response was investigated using conventional histology and scanning electron microscopy. Histological and histomorphometric evaluation of specimens after 12 weeks implantation showed significantly more bone contact with the surface of 45S5 bioglass implants when compared with other test materials. When the bone contact for each material was compared between experimental time points, the Glass-Ceramic 2 (CaO rich) group showed significant difference (p = 0.027) at 4 weeks, but no direct contact at 12 weeks. Histology and backscattered electron photomicrographs showed that modified fluorcanasite glass-ceramic implants had greater osteoconductivity than the parent stoichiometric composition. Of the new materials, fluorcanasite glass-ceramic implants modified by the addition of P(2)O(5) showed the greatest stimulation of new mineralized bone tissue formation adjacent to the implants after 4 and 12 weeks implantation. (C) 2010 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 94A: 760-768, 2010

Fatty acid composition of intramuscular fat from Nellore steers fed dry or high moisture corn and calcium salts of fatty acids

The purpose of this work was to evaluate the fatty acid composition of the Longissimus muscle from carcasses of Nellore steers fed diets with calcium salts of fatty acids (CSFA) and high moisture corn. Forty eight steers were fed during 70 days four diets containing dry corn (DC), high moisture corn (HM). dry corn plus CSFA (DC-CSFA) or high moisture corn plus CSFA (HM-CSFA). Fatty acid composition of the Longissimus muscle was determined by gas chromatography. Corn type had no effect on the ether extract percentage and in the content of the majority of the fatty acids, although steers fed HMC showed higher levels of polyunsaturated fatty acids and polyunsaturated/saturated ratio. Feeding CSFA increased ether extract percentage but had no effect on total of saturated, unsaturated and saturated: unsaturated ratio. Both high moisture corn and calcium salts of fatty acids increased CIA (cis9, trans11) and total CIA concentrations in intramuscular fat (C) 2008 Elsevier B.V. All rights reserved.

Anatomical and biochemical changes in the composition of developing seed coats of annatto (Bixa orellana L.)

Seeds of Bixa orellana (L.) have a sclerified palisade cell layer, which constitutes a natural barrier to water uptake. In fact, newly fully developed B. orellana seeds are highly impermeable to water and thereby dormant. The purpose of this work is to investigate, from a developmental point of view, the histochemical and physical changes in the cell walls of the seed coat that are associated with the water impermeability. Seed coat samples were analyzed by histochemical and polarization microscopy techniques, as well as by fractionation/HPAEC-PAD. For histochemical analysis the tissue samples were fixed, dehydrated, embedded in paraffin and the slides were dewaxed and tested with appropriate stains for different cell wall components. Throughout the development of B. orellana seeds, there was a gradual thickening of the seed coat at the palisade region. This thickening was due to the deposition of cellulose and hemicelluloses in the palisade layer cell walls, which resulted in a highly water impermeable seed coat. The carbohydrate composition of the cell walls changed dramatically at the late developmental stages due to the intense deposition of hemicelluloses. Hemicelluloses were mainly deposited in the outer region of the palisade layer cell walls and altered the birefringent pattern of the walls. Xylans were by far the most abundant hemicellulosic component of the cell walls. Deposition of cellulose and hemicelluloses, especially xylans, could be responsible for the impermeability to water observed in fully developed B. orellana seeds.

Enhanced visualization of histological samples with an adjustable RGB contrast system with application for tissue used in photodynamic therapy

The analysis of histological sections has long been a valuable tool in the pathological studies. The interpretation of tissue conditions, however, relies directly on visual evaluation of tissue slides, which may be difficult to interpret because of poor contrast or poor color differentiation. The Chromatic Contrast Visualization System (CCV) combines an optical microscope with electronically controlled light-emitting diodes (LEDs) in order to generate adjustable intensities of RGB channels for sample illumination. While most image enhancement techniques rely on software post-processing of an image acquired under standard illumination conditions, CCV produces real-time variations in the color composition of the light source itself. The possibility of covering the entire RGB chromatic range, combined with the optical properties of the different tissues, allows for a substantial enhancement in image details. Traditional image acquisition methods do not exploit these visual enhancements which results in poorer visual distinction among tissue structures. Photodynamic therapy (PDT) procedures are of increasing interest in the treatment of several forms of cancer. This study uses histological slides of rat liver samples that were induced to necrosis after being exposed to PDT. Results show that visualization of tissue structures could be improved by changing colors and intensities of the microscope light source. PDT-necrosed tissue samples are better differentiated when illuminated with different color wavelengths, leading to an improved differentiation of cells in the necrosis area. Due to the potential benefits it can bring to interpretation and diagnosis, further research in this field could make CCV an attractive technique for medical applications.

Health in Madeira: a comprehensive study of aging, body composition, physical activity and functional fitness

Physical Activity (PA) and functional fitness (FF) are predictors of a healthy and independent lifestyle in older adults. The purpose of this study was: (1) to construct reference values for FF; (2) to describe sex- and age-related changes in FF, balance, gait, PA, body composition, and bone health/strength; and (3) to determine their variation and co-variation with respect to PA. This cross-sectional study included 401 males and 401 females aged 60-79 years old. FF was assessed using the Senior Fitness test and balance by the Fullerton Advance Balance scale (FAB). Gait parameters: gait velocity (GV), stride length (SL), cadence and gait stability ratio (GSR) were measured. Femoral strength index (FSI) and bone mineral density (BMD) of the total body, lumbar spine, hip region and total lean tissue mass (TLTM) and total fat mass (TFM) were determined by dual-energy x-ray absorptiometry-DXA. PA was assessed during face-to-face interviews using the Baecke questionnaire. Demographic and health history information were obtained by structured telephone interview. In both sexes, a significant main effect for age-group was found for FF parameters, balance scores, gait performances, TLTM and hip, LS and total BMD and FSI. Likewise there were significant main effects for age-group for total PA in women and sports related PA in men. Men scored significantly better than women in FF (except in upper- and lower-body flexibility), balance, GV, SL, GSR and had higher BMD and TLTM compared with women. Active subjects scored better in FF, balance, and gait than their average and non-active peers. PA and FF exerted only a minor influence in the differentiation of BMD and FSI among the elderly while constitutive factors like age, height, body mass, TLTM and TFM entered as the most significant contributors. This study gives scientific support to public policies at the community level, targeted to increase PA, FF and TLTM, thereby contributing to improved quality of life in older adults.

Evaluation of muscle tissue growth in Young Mediterranean Buffaloes slaughtered at different weights

This study evaluated muscle, adipose and bone tissue growth in young Mediterranean buffaloes slaughtered with different weights. Twenty eight non-castrated males, approximately 14 months old and 330 kilos, were distributed in four experimental and one control groups. Received the same diet during a non-fixed period and when reached the pre-established weights (450 kg, 480 kg, 510 kg, 540 kg) were fasted for 16 hours and slaughtered. NRC 1996, level 2, ruminal simulation program for non-castrated animals with a daily weight gain of 1.40 kg determined percentage composition of experimental diet (dry matter): coast cross hay (20.6%), corn silage (7.8%), cotton seed (8.2%), humid corn silage (46%) and commercial mineral concentrate (17.4%), representing 13% of crude protein and 2,68% of metabolizable energy (Mcal/kg). Feed was offered ad libitum twice daily. Slaughter of control group occurred after 30 days of an adaptation period. The other ones were weighted each 28 days until reached pre-established weight. Carcasses were chilled for 18 hours under -5 degrees C. Ninth, 10 degrees and 11 degrees ribs of the half left carcass were submitted to HH section and according to the statistical regression analysis (SAS, 1996), allometric coefficient value was negative (b<1) demonstrating the early growth of muscle in relation to empty carcass weight.

Histomorphometric analysis of tissue responses to bioactive glass implants in critical defects in rat calvaria

The aim of this study was to evaluate the osteogenic behavior of two chemically similar bioactive glass products (Biogran (R) and Perioglas (R)) implanted in critical bone defects in rat calvaria. Thirty-six transfixed bone defects of 8 mm diameter were made surgically in adult male Wistar rats. The animals were distributed equally into three groups: Biogran (GI), Perioglas (GII) and without implant material (control; GIII). The morphology and composition of both bioactive glasses were analyzed by scanning electron microscopy and energy-dispersive spectrometry. Tissue specimens were analyzed at the biological time points of 15, 30 and 60 days by optical microscopy and morphometry, demonstrating biocompatibility for the tested materials with moderate chronic inflammation involving their particles. Bone neoformation resulted only as a reparative reaction to an intentionally produced defect and was limited to the defect's edges. No statistically significant differences among the groups were observed. At the scar interstice, abundant deposits of collagenous fibers enveloping the particles were noted. The present results indicated that the bioactive glasses, under the experimental conditions analyzed, did not show osteogenic behavior. Copyright (c) 2007 S. Karger AG, Basel.

Stable carbon (delta(13)C) and nitrogen (delta(15)N) isotopes as natural indicators of live and dry food in Piaractus mesopotamicus (Holmberg, 1887) larval tissue

This study proposed the use of the stable isotope technique to track the type of food utilized by pacu Piaractus mesopotamicus larvae during their development, and to identify the moment when the larvae start using nutrients from the dry diet by retaining its carbon and nitrogen atoms in their body tissues. Five-day-old pacu larvae at the onset of exogenous feeding were fed Artemia nauplii or formulated diet exclusively; nauplii+formulated diet during the entire period; or were weaned from nauplii to a dry diet after 3, 6 or 12 days after the first feeding. delta(13)C and delta(15)N values for Artemia nauplii were -15.1 parts per thousand and 4.7 parts per thousand, respectively, and -25.0 parts per thousand and 7.4 parts per thousand for the dry diet. The initial isotopic composition of the larval tissue was -20.2 parts per thousand and 9.5 parts per thousand for delta(13)C and delta(15)N respectively. Later, at the end of a 42-day feeding period, larvae fed Artemia nauplii alone reached values of -12.7 parts per thousand and 7.0 parts per thousand for delta(13)C and delta(15)N respectively. Larvae that received the formulated diet alone showed values of -22.7 parts per thousand for delta(13)C and 9.6 parts per thousand for delta(15)N. The stable isotope technique was precise, and the time at which the larvae utilized Artemia nauplii, and later dry diet as a food source could be clearly defined.

Behavior of plant tissue in osmotic solutions

The effect of the concentration of sucrose solutions on the cellular structure of potato tissue in equilibrium at 27 degreesC was Studied. Two different methods of investigation were used to determine the volume of the different phases composing the cellular tissue of the potato when in equilibrium with the solutions. one based on data of the concentration itself and the overall volume of 2 mm slices after 48 h at equilibrium, and the other on microscopic images of cells in thin slices of fresh tissue stained with neutral red after an hour in equilibrium to show protoplasts, vacuoles and plasmolysis spaces. The results of these methods were compared with those obtained by a predictive thermodynamic approach considering the semipermeability of cell membranes. Phase volume data obtained from microscopic analysis were more similar to what was predicted by the theoretical model than those obtained by means of composition measurement. where the long equilibrium time apparently led to the loss of semi permeability of the cell membranes, since total volumes calculated without consideration of the cell membranes were similar to those measured. This suggests that the length of time of osmotic dehydration brings about a change in cell structure and the consequent involvement of a different mechanism in mass transfer. (C) 2002 Elsevier B.V. Ltd. All rights reserved.

Carbon isotope composition and leaf anatomy as a tool to characterize the photosynthetic mechanism of Artemisia annua L.

Leaves of Artemisia annua L. are a plentiful source of artemisinin, a drug with proven effectiveness against malaria. The aim of this study was to classify the photosynthetic mechanism of A. annua through studies of the carbon isotope composition (δ 13C) and the leaf anatomy. A. annua presented a δ 13C value of - 31.76 ± 0.07, which characterizes the plants as a typical species of the C3 photosynthethic mechanism, considering that the average δ 13C values for C3 and C4 species are -28 and -14, respectively. The leaf anatomy studies were consistent with the δ 13C results, where, in spite of the existence of parenchymatic cells forming a sheath surrounding the vascular tissue, the cells do not contain chloroplasts or starch. This characteristic is clearly different from that of the Kranz anatomy found in C4 species.