Cigarette smoke concentrate increases 8-epi-PGF2alpha and TGFbeta1 secretion in rat mesangial cells

Epidemiological studies have shown that cigarette smoke, an oxidant agent, is a risk factor for the development of diabetic nephropathy (DN), in which pathogenesis transforming growth factor beta(1) (TGFbeta(1)) plays a key role. In our experimental model we exposed mesangial cell cultures to cigarette smoke concentrate (CSC) to study the effect of smoking on the pathogenesis of DN. Thus, we analyzed the effect of CSC on TGFbeta(1) and lipid peroxidation (8-epi-PGF(2alpha)) in rat mesangial cells. Furthermore, since the protein kinase C (PKC) pathway appears to be a key factor for the enhanced production of TGFbeta(1), we also analyzed the effect of the selective PKCbeta inhibitor LY379196 on TGFbeta(1) response to CSC. CSC induced an increase of both TGFbeta(1) and 8-epi-PGF(2) compared to basal conditions (5 mM glucose). The CSC-induced increase in TGFbeta(1) secretion was significantly suppressed by LY379196. These data suggest that smoking could increase TGFbeta(1) production, probably due to oxidative stress and PKCbeta activation. This finding supports the concept that smoking is a risk factor for DN development.     

Biochemical,physicochemical and molecular characterization of a genuine 2-Cys-peroxiredoxin purified from cowpea [Vigna unguiculata (L.) Walpers] leaves

Background

Peroxiredoxins have diverse functions in cellular defense-signaling pathways. 2-Cys-peroxiredoxins (2-Cys-Prx) reduce H₂O₂ and alkyl-hydroperoxide. This study describes the purification and characterization of a genuine 2-Cys-Prx from Vigna unguiculata (Vu-2-Cys-Prx).

Methods

Vu-2-Cys-Prx was purified from leaves by ammonium sulfate fractionation, chitin affinity and ion exchange chromatography.

Results

Vu-2-Cys-Prx reduces H₂O₂ using NADPH and DTT. Vu-2-Cys-Prx is a 44 kDa (SDS-PAGE)/46 kDa (exclusion chromatography) protein that appears as a 22 kDa molecule under reducing conditions, indicating that it is a homodimer linked intermolecularly by disulfide bonds and has a pI range of 4.56–4.72; its NH₂-terminal sequence was similar to 2-Cys-Prx from Phaseolus vulgaris (96%) and Populus tricocarpa (96%). Analysis by ESI-Q-TOF MS/MS showed a molecular mass/pI of 28.622 kDa/5.18. Vu-2-Cys-Prx has 8% α-helix, 39% β-sheet, 22% of turns and 31% of unordered forms. Vu-2-Cys-Prx was heat stable, has optimal activity at pH 7.0, and prevented plasmid DNA degradation. Atomic force microscopy shows that Vu-2-Cys-Prx oligomerized in decamers which might be associated with its molecular chaperone activity that prevented denaturation of insulin and citrate synthase. Its cDNA analysis showed that the redox-active Cys⁵² residue and the amino acids Pro⁴⁵, Thr⁴⁹ and Arg¹²⁸ are conserved as in other 2-Cys-Prx.

General significance

The biochemical and molecular features of Vu-2-Cys-Prx are similar to other members of 2-Cys-Prx family. To date, only one publication reported on the purification of native 2-Cys-Prx from leaves and the subsequent analysis by N-terminal Edman sequencing, which is crucial for construction of stromal recombinant 2-Cys-Prx proteins.     

Effect of the rate of increase in concentrate allowance with concentrates high or low in starch on the performance of dairy cows in early lactation

The effect of the rate of increase in concentrate allowance after calving with two concentrate mixes (A and B) differing in composition was evaluated using 64 Finnish Ayrshire cows during the first 100 days of lactation. After calving, the concentrate allowance of multiparous cows was increased stepwise from 4 to 17 kg/day, and of primiparous cows from 3 to 13.5 kg/day over 12 days (F rate of increase; multiparous 1.08 kg/day, primiparous 0.88 kg/day) or 24 days (S rate of increase; mutiparous 0.54 kg/day, primiparous 0.44 kg/day). The concentrates were formulated to have similar crude protein and metabolizable energy concentrations but differing starch and NDF concentrations. For concentrate A the starch and NDF concentrations were 421 and 167 g/kg dry matter (DM) and for concentrate B 258 and 251 g/kg DM. All cows received grass silage ad libitum. The higher concentrate intake during weeks 1 to 4 of lactation with F compared with the S rate of increase caused higher DM, energy and protein intake. The higher concentrate intake for F than for S treatment in early lactation did not cause a large decrease in silage intake (8.8 v. 8.3 kg DM/day). The intake of concentrate A and B after calving did not differ for S treatment. However, for F treatment the intake of fibrous concentrate B increased faster than starch-rich concentrate A during weeks 1 to 4 of lactation. The concentrate composition had no effect on energy-corrected milk (ECM) yield during weeks 1 to 4 of lactation for S treatments, but with F treatments the cows fed B concentrate produced more milk. The F rate of increase in concentrate allowance compared with the S rate increased the calculated energy balance after calving. The rate of increase in concentrate feeding post partum or concentrate composition had no effect on DM, energy or protein intake during the whole 100-day experiment. The average ECM yield over days 1 to 100 of lactation was higher for S than for F treatments and tended to be higher with concentrate B than A. Results of this study showed that by the fast rate of increase in concentrate allowance after calving on a grass silage diet, it was possible to improve the energy status of the cows in early lactation. This had, however, no effect on production later in lactation.     

Evaluation of concentrate factors affecting silage intake of dairy cows: a development of the relative total diet intake index

The aim of this work was to develop an index describing the relative intake of the total diet by dairy cows, and hence the ability to predict intake responses to changes in both forage and concentrate variables. An evaluation of concentrate factors affecting silage dry matter (DM) intake of dairy cows was conducted based on dietary treatment means from milk production experiments. The data were divided into four subsets according to concentrate treatments used within the experiments: the amount of concentrate supplementation (n = 217), protein supplementation (n = 336), carbohydrate composition (n = 114) and fat concentration of the concentrate (n = 29). The data were subjected to mixed-model regression analysis. Increased concentrate DM intake (CDMI) decreased silage DM intake (SDMI) quadratically. The substitution rate (substitution of silage DM for concentrate DM) increased with improved silage intake potential. SDMI increased quadratically with concentrate protein intake, the response being negatively related to the effective protein degradability (EPD) of concentrates. Replacement of starchy concentrate ingredients with fibrous supplements had a small positive effect on silage intake, whereas increased concentrate fat concentration slightly decreased SDMI. The outcome of concentrate factors influencing total DM intake (TDMI) was used to create a relative CDMI index as follows: CDMI index = 100 + 10 × [(CDMI - 0.1629 × CDMI - 0.01882 × CDMI2 - 5.49) + ((0.9474 × CCPI - 0.4965 × CCPI2) - 2.017 × (CEPD - 0.74)) + 0.00225 × (CNDF - 250) - 0.0103 × (40 - Cfat) - 0.00058 × (CDMI - 8.0) × (SDMI index - 100)], where CDMI = concentrate DM intake (kg/day), CCPI = supplementary concentrate CP intake (kg/day; CP>170 g/kg DM), CEPD = concentrate EPD (g/g), CNDF = concentrate NDF concentration (g/kg DM), Cfat = concentrate fat concentration (g/kg DM) and SDMI index is the relative intake potential of silage (Huhtanen, Rinne and Nousiainen 2007. Animal 1, 758-770). TDMI index was calculated as SDMI index + CDMI index - 100 to describe the relative intake potential of the total diet. For the whole data set (n = 943), one TDMI index unit was equivalent to 0.095 kg/day DM intake, i.e. close to the default value of 0.100 kg. The CDMI index explained proportionally 0.88 of the variation in TDMI within a study with a 0.27 kg/day residual mean-square error (n = 616). The corresponding values for the TDMI index were 0.81 and 0.37 kg/day (n = 943), respectively. The residual mean-square errors in cross-validation were marginally higher. The developed TDMI index can be used to estimate the intake responses to diet changes. It provides an improved basis for practical dairy cow ration formulation and economic evaluation.     

Entamoeba histolytica: lipid rafts are involved in adhesion of trophozoites to host extracellular matrix components

Adhesion is an important virulence function for Entamoeba histolytica, the causative agent of amoebic dysentery. Lipid rafts, cholesterol-rich domains, function in compartmentalization of cellular processes. In E. histolytica, rafts participate in parasite-host cell interactions; however, their role in parasite-host extracellular matrix (ECM) interactions has not been explored. Disruption of rafts with a cholesterol extracting agent, methyl-β-cyclodextrin (MβCD), resulted in inhibition of adhesion to collagen, and to a lesser extent, to fibronectin. Replenishment of cholesterol in MβCD-treated cells, using a lipoprotein-cholesterol concentrate, restored adhesion to collagen. Confocal microscopy revealed enrichment of rafts at parasite-ECM interfaces. A raft-resident adhesin, the galactose/N-acetylgalactosamine-inhibitable lectin, mediates interaction to host cells by binding to galactose or N-acetylgalactosamine moieties on host glycoproteins. In this study, galactose inhibited adhesion to collagen, but not to fibronectin. Together these data suggest that rafts participate in E. histolytica-ECM interactions and that raft-associated Gal/GalNAc lectin may serve as a collagen receptor.     

Nutritional properties of potato protein concentrate compared with soybean meal as the main protein source in feed for the double-muscled Belgian Blue bull

The objective of this experiment was to compare the nutritional properties of potato protein concentrate, a by-product of the starch industry produced entirely in Europe, with that of soybean meal (SBM), for growing cattle. The experiment was conducted on double-muscled Belgian Blue bulls, fitted with rumen, duodenal and ileal cannulas, according to a 4 × 4 Latin square design. They were fed three different iso-N and iso-net energy diets formulated according to the Dutch feed evaluation system, differing in the nature of the main protein source, which was either SBM (‘SBM’ treatment), potato protein concentrate (PPC, ‘PPC’ treatment) or an iso-N mixture of these two protein sources (‘mixed’ treatment). A fourth treatment consisted of ‘PPC’ supplemented by 9.5% digestible proteins supplied by duodenal perfusion of sodium caseinate (CAS, ‘PPC + CAS’ treatment). No significant difference was observed in the ruminal fluid pH, whereas both ‘PPC’ and ‘PPC + CAS’ had the effect of reducing the ruminal ammonia nitrogen (N-NH₃) concentration. No significant difference was observed in the apparent intestinal digestibility of the dry matter (DM), organic matter (OM) or N. Outflows of non-NH₃-N, microbial proteins and dietary proteins from the rumen were similar for ‘PPC’, ‘SBM’ and ‘mixed’, and increased with CAS infusion by 20%, 17% and 27%, respectively. On the basis of in vivo observations, the degradability of SBM and PPC proteins was estimated at 0.60 and 0.43, respectively, corresponding to the values quoted in the literature. The supply of digestible essential amino acids (EAA) was significantly greater with ‘PPC + CAS’ and did not differ among ‘SBM’, ‘mixed’ and ‘PPC’. This illustrates the difficulty of altering the amino acid (AA) pattern of digestible protein by the nature of the protein of dietary origin when an animal is fed a high nutritional value diet. N retention was not affected by replacing SBM with PPC, but increased by 10% with CAS infusion. On the basis of the plasma AA pattern, the supply of digestible Met was probably limiting with ‘SBM’, ‘mixed’ and ‘PPC’. The CAS perfusion supplemented all AA, including Met, leading to increased N retention. This improvement was limited, however, and N utilisation remained unchanged between treatments. In conclusion, despite a more favourable EAA pattern, PPC offered no advantage compared with SBM for growing bulls when diets were formulated according to the Dutch feed evaluation system.     

饲料中土豆蛋白替代鱼粉对虹鳟摄食率、消化率和生长的影响

通过生长试验探讨了饲料中土豆蛋白浓缩物替代鱼粉后对虹鳟摄食率,消化率,生长和饲料利用的影响,对照饲料全部鱼粉为蛋白源,其它饲料分别含土豆蛋白２．２％,５．６％,８．９％和１１．１％,另一饲料含５．６％,土豆蛋白和１．７％蛋氨酸,在１２℃下饲养４周,试验结果表明,随着饲料中土豆蛋白使用比例的增加,虹鳟的摄食率下降,饲料的干物质,蛋白和灰分的表现消化率上升,脂肪的表现消化率无显著影响,当饲料中含５．６     

饲料中添加乌贼肝粉和鱼浆蛋白对中华鳖抗氧化能力的影响

本实验研究了饲料中添加乌贼肝粉和鱼浆蛋白对中华鳖抗氧化能力的影响。设计了3个实验组,对照组（白鱼粉组）、乌贼肝粉组和鱼浆蛋白组。3种实验饲料的营养成分相近,主要区别是乌贼肝粉组和鱼浆蛋白组的饲料中分别用乌贼肝粉和鱼浆蛋白取代了对照组饲料中5%的白鱼粉。用3种饲料分别养殖中华鳖（Pelodiscus sinensis）稚鳖8周,测定了稚鳖血清和肝组织的超氧化物歧化酶（SOD）活性、丙二醛含量（MDA）、总抗氧化能力（T-AOC）以及血清碱性磷酸酶（AKP）和溶菌酶（LZM）的活性。结果发现,乌贼肝粉组稚鳖血清和肝MDA含量显著低于对照组,T-AOC高于对照组;鱼浆蛋白组稚鳖血清和肝MDA含量、血清T-AOC与对照组没有显著差异,但肝T-AOC显著低于对照组。实验结果表明,饲料中添加5%的乌贼肝粉能提高稚鳖的抗氧化能力。     

Impact of cow strain and concentrate supplementation on grazing behaviour,milk yield and metabolic state of dairy cows in an organic pasture-based feeding system

As ruminants are able to digest fibre efficiently and assuming that competition for feed v. food use would intensify in the future, cereals and other field crops should primarily be destined to cover the dietary needs of humans and monogastric animals such as poultry and pigs. Farming systems with a reduced or absent concentrate supplementation, as postulated by organic agriculture associations, require adapted dairy cows. The aim of this experiment was to examine the impact of concentrate supplementation on milk production, grazing and rumination behaviour, feed intake, physical activity and blood traits with two Holstein-Friesian cow strains and to conclude the consequences for sustainable and organic farming. The experiment was a cross-over study and took place on an organic farm in Switzerland. In all, 12 Swiss Holstein-Friesian (HCH) cows and 12 New Zealand Holstein-Friesian (HNZ) cows, which were paired according to lactation number, days in milk and age for primiparous cows, were used. All cows grazed full time and were supplemented either with 6 kg/day of a commercial, organic cereal-grain mix or received no supplement. After an adaptation period of 21 days, a measurement period of 7 days followed, where milk yield and composition, pasture dry matter intake estimated with the n-alkane double-indicator technique, physical activity based on pedometer measurements, grazing behaviour recorded by automatic jaw movement recorder and blood samples were investigated. Non-supplemented cows had a lower milk yield and supplemented HCH cows produced more milk than supplemented HNZ cows. Grazing time and physical activity were greater for non-supplemented cows. Supplementation had no effect on rumination behaviour, but HNZ cows spent longer ruminating compared with HCH cows. Pasture dry matter intake decreased with the concentrate supplementation. Results of blood analysis did not indicate a strong negative energy balance for either non-supplemented or supplemented cows. Minor differences between cow strains in this short-term study indicated that both cow strains are equally suited for an organic pasture-based production system with no concentrate supplementation. Many factors such as milk yield potential, animal welfare and health, efficiency, grazing behaviour and social aspects influence the decision to supplement grazing dairy cows with concentrates.     

Effects of concentrate type and chromium propionate on insulin sensitivity,productive and reproductive parameters of lactating dairy cows consuming excessive energy

This experiment compared insulin sensitivity parameters, milk production and reproductive outcomes in lactating dairy cows consuming excessive energy, and receiving in a 2×2 factorial arrangement design: (1) concentrate based on ground corn (CRN; n=13) or citrus pulp (PLP; n=13), and (2) supplemented (n=14) or not (n=12) with 2.5 g/day of chromium (Cr)-propionate. During the experiment (day 0 to 182), 26 multiparous, non-pregnant, lactating Gir×Holstein cows (initial days in milk=80±2) were offered corn silage for ad libitum consumption, and individually received concentrate formulated to allow diets to provide 160% of their daily requirements of net energy for lactation. Cow BW and body condition score (BCS) were recorded weekly. Milk production was recorded daily and milk samples collected weekly. Blood samples were collected weekly before the morning concentrate feeding. Glucose tolerance tests (GTT; 0.5 g of glucose/kg of BW) were performed on days −3, 60, 120 and 180. Follicle aspiration for in vitro embryo production was performed via transvaginal ovum pick-up on days −1, 82 and 162. No treatment differences were detected (P⩾0.25) for BW and BCS change during the experiment. Within weekly blood samples, concentrations of serum insulin and glucose, as well as insulin : glucose ratio were similar among treatments (P⩾0.19), whereas CRN had less (P<0.01) non-esterified fatty acid concentrations compared with PLP (0.177 v. 0.215 mmol/l; SEM=0.009). During the GTT, no treatment differences were detected (P⩾0.16) for serum glucose concentration, glucose clearance rate, glucose half-life and insulin : glucose ratio. Serum insulin concentrations were less (P=0.04) in CRN supplemented with Cr-propionate compared with non-supplemented CRN (8.2 v. 13.5 µIU/ml, respectively; SEM=1.7), whereas Cr-propionate supplementation did not impact (P=0.70) serum insulin within PLP cows. Milk production, milk fat and solid concentrations were similar (P⩾0.48) between treatments. However, CRN had greater (P<0.01) milk protein concentration compared with PLP (3.54% v. 3.14%, respectively; SEM=0.08). No treatment differences were detected (P⩾0.35) on number of viable oocytes collected and embryos produced within each aspiration. In summary, feeding a citrus pulp-based concentrate to lactating dairy cows consuming excessive energy did not improve insulin sensitivity, milk production and reproductive outcomes, whereas Cr-propionate supplementation only enhanced insulin sensitivity in cows receiving a corn-based concentrate during a GTT.