Farmers without borders-genetic structuring in century old barley (Hordeum vulgare).

The geographic distribution of genetic diversity can reveal the evolutionary history of a species. For crop plants, phylogeographic patterns also indicate how seed has been exchanged and spread in agrarian communities. Such patterns are, however, easily blurred by the intense seed trade, plant improvement and even genebank conservation during the twentieth century, and discerning fine-scale phylogeographic patterns is thus particularly challenging. Using historical crop specimens, these problems are circumvented and we show here how high-throughput genotyping of historical nineteenth century crop specimens can reveal detailed geographic population structure. Thirty-one historical and nine extant accessions of North European landrace barley (Hordeum vulgare L.), in total 231 individuals, were genotyped on a 384 single nucleotide polymorphism assay. The historical material shows constant high levels of within-accession diversity, whereas the extant accessions show more varying levels of diversity and a higher degree of total genotype sharing. Structure, discriminant analysis of principal components and principal component analysis cluster the accessions in latitudinal groups across country borders in Finland, Norway and Sweden. FST statistics indicate strong differentiation between accessions from southern Fennoscandia and accessions from central or northern Fennoscandia, and less differentiation between central and northern accessions. These findings are discussed in the context of contrasting historical records on intense within-country south to north seed movement. Our results suggest that although seeds were traded long distances, long-term cultivation has instead been of locally available, possibly better adapted, genotypes.

Use of gene profiling to evaluate the effects of a feed additive on immune function in periparturient dairy cattle.

Objectives were to investigate mechanisms by which a nutritional supplement alters immunity in dairy cattle. Our hypothesis was that feeding this product to dairy cattle altered neutrophil gene expression. Eight periparturient Jersey cattle were randomly assigned to one of two treatments: control and treated. Control animals were fed a dry cow ration for 1 month prior to calving. The treated cows were fed the same ration supplemented with OmniGen-AF. Following calving, blood samples were taken and neutrophils were prepared after which RNA was extracted. Gene expression in neutrophils of treated versus control-fed animals was then assessed using bovine-total leukocyte (BOTL-5) arrays. Eighteen genes were differentially regulated in the experimental group and of these, twice as many were up-regulated as down-regulated. Patterns of changes indicated that the additive might alter neutrophil apoptosis, signaling and sensitivity. Two of the regulated genes [interleukin-1beta converting enzyme (ICE) and interleukin-4 receptor (IL-4R)] were investigated in more detail using quantitative reverse transcriptase-polymerase chain reaction (QRT-PCR). Each was found to be elevated by the feeding of experimental product. Increased expression of ICE indicates potential for enhanced neutrophil expression of interleukin-1beta (IL-1beta), a cytokine which plays roles in the inflammatory response and which stimulates adaptive immunity following innate immune activation. Altered expression of IL-4R indicates potential for changes in neutrophil apoptosis. The experiment identified mechanisms by which the additive altered neutrophil gene expression. While many nutrients support the immune system, we have shown that a non-traditional nutritional approach may also have utility in modulating immune function.

Amino acid transport System L in muscle cells: biochemical properties and its relation to protein synthesis.

Objectives were to characterize mechanisms and biochemical properties of transport systems responsible for the uptake of branched-chain amino acids (BCAAs) in muscle cells. Rat omega myoblasts (RMo) were grown to confluency and allowed to differentiate prior to conduct of transport assays. Myotubes concentrated cycloleucine (cLeu) in a sodium (Na)-free medium. The Na gradient-independent transporter possessed high affinity (Km = 0.12 mM) and high capacity (Vmax = 6.4 nmol cLeu/mg protein per min). Cycloleucine transport was strongly inhibited by nonpolar neutral amino acids but not by alpha-aminoisobutyric acid or lysine. Myotubes possessed a Na gradient-independent trans-exchange mechanism. Hence, myotubes possess a System L-like transporter. In the second part of the study we determined that various inhibitors (KCN, oligomycin, iodoacetamide and cycloheximide) increased leucine transport. Their actions were not mediated by reductions in ATP concentration but were instead associated with changes in protein synthesis. Hence, regulation of muscle protein synthesis may also influence transporter activity.

Protein kinase C isoforms in muscle cells and their regulation by phorbol ester and calpain.

Objectives were to identify the PKC isoforms in cultured muscle cells, to examine roles of Ca(2+)-dependent proteinases (calpains) in processing of various muscle PKC isozymes and to obtain a mechanistic description of the processing of PKCs by examining the temporal relationships between phorbol ester-dependent translocation of muscle PKCs and calpains between cytosolic and membrane compartments. Using six isoform (alpha, beta, gamma, delta, epsilon, zeta)-specific polyclonal antibodies, PKC alpha, delta and zeta were detected in rat skeletal muscle and in L8 myoblasts and myotubes. PKC alpha and zeta were primarily localized in the cytosolic fraction of L8 myotubes whereas PKC delta was more abundant in the membrane fraction. Phorbol ester (TPA) caused rapid depletion of myotube PKC alpha and PKC alpha and PKC delta isoforms from the cytosolic compartment and rapid appearance of these isoforms in the membrane fraction. However, long-term exposure of myotubes to TPA eventually caused down-regulation of PKCs in the membrane compartment. Down-regulation of PKCs in the membrane fraction was partially blocked by calpain inhibitor II. However, the rapid TPA-dependent cytosolic depletion of PKCs was unaffected by calpain inhibitor. This suggests that calpains may be responsible for membrane-associated down-regulation of PKCs but not for cytosolic depletion. In the final study we assessed the effects of phorbol ester on compartmentation of m-calpain with PKCs in muscle cells. Like the PKCs, TPA caused rapid association of m-calpain with the membrane fraction followed by down-regulation. This demonstrates that phorbol esters cause translocation of both PKCs and calpains to membranes where processing of PKCs may occur via the limited proteolysis exerted by calpains.

Identification of glycogen phosphorylase and creatine kinase as calpain substrates in skeletal muscle.

The goal of this study was to identify calpain substrates in muscle cells. Our hypothesis was that the yeast two-hybrid method could be used to identify novel calpain substrates. To accomplish this, native mu- and m-calpains, as well as a variety of calpain DNA fragments, were expressed in yeast cells and used to screen for binding proteins in a human skeletal muscle cDNA library. Calpain constructs that were used in the screening process included native mu- and m-calpains, a dominant negative (DN) m-calpain (i.e. active site modified), N-terminal truncated DN m-calpain (i.e. autolyzed DN-m-calpain) and, finally, an N- and C-terminal truncated m-calpain (i.e. autolyzed DN-m-calpain lacking a calcium-binding domain). Yeast cells were transformed using yeast two-hybrid expression vectors containing the different calpain constructs as "baits". Beta-galactosidase activity was assayed as an index of interaction between calpain and its potential target proteins. From this analysis, four clones (Ca2+-ATPase, novel nebulin-related protein (N-RAP), creatine kinase and glycogen phosphorylase) were recovered. Two of these, creatine kinase and glycogen phosphorylase, were selected for further study. In in-vitro assays, calpain was able to partially digest both proteins, suggesting that both creatine kinase and glycogen phosphorylase are natural calpain substrates.

Evidence for the participation of the proteasome and calpain in early phases of muscle cell differentiation.

Objectives were to investigate the role of the proteasome and m-calpain to muscle cell differentiation. Accordingly, we investigated the effects of lactacystin, a proteasome inhibitor, and calpain inhibitor-II (CI-II) on L8 muscle cell differentiation and assessed concentrations of proteasomal and calpain subunit mRNAs during differentiation. L8 myoblasts were induced to differentiate by culturing in mitogen-depleted medium. To assess the importance of the proteasome and calpain to differentiation, we examined effects of lactacystin and CI-II on creatine kinase (CK) activity. In the absence of inhibitor, CK activity was detectable within 48 h of mitogen depletion and myotubes were formed. Addition of lactacystin or CI-II to cultures drastically reduced CK activity and prevented formation of myotubes. Hence, proteasome and calpain are both necessary for differentiation. In order to identify which proteasomal subunits were regulated during differentiation, we examined the concentrations of two 20S core subunits (C8 and C9) and three 22S ATPases (MSS1, S4 and TBP1) during differentiation. Concentrations of m-calpain and beta-tubulin mRNAs were also assessed. Differentiation was associated with slight increases (ca. 30%) in concentrations of mRNAs encoding the proteasomal 20S core subunits (C8 and C9) and with large increases (approximately 2-fold) in mRNAs encoding the regulatory subunit ATPases. m-calpain mRNA concentration also increased two-fold following mitogen depletion. beta-Tubulin mRNA concentration remained unchanged early in the differentiation process and thereafter declined. Of interest, changes in proteasomal and m-calpain mRNAs occurred within 6-24 h of mitogen depletion (i.e., at least 24-36 h prior to detectable changes in creatine kinase activity). These results indicate that changes in expression of proteasome and calpains subunits occur early in the differentiation process. These changes may be required for the normal course of differentiation to proceed. Differentiation is associated with larger changes in proteasomal ATPase mRNAs than in 20S core particle mRNAs indicating that either turnover rates of the 22S ATPase subunits are more rapid in differentiating cells than of the 20S core particles or that functions of the regulatory subunits become more important during muscle cell differentiation.

Involvement of protein kinase-C, calpains, and calpastatin in prostaglandin F2 alpha-induced oxytocin secretion from the bovine corpus luteum.

An experiment was conducted to determine whether prostaglandin F2 alpha (PGF2 alpha)-induced secretion of oxytocin (OT) by the bovine corpus luteum (CL) was associated with changes in the activities of protein kinase-C (PKC), calpains, and calpastatin. On day 8 of the estrous cycle (estrus = day 0), beef heifers were restrained and given a 500-micrograms iv injection of cloprostenol, a PGF2 alpha analog. Corpora lutea were surgically removed from beef heifers 0, 2, 7.5, or 30 min (n = 4 animals/time period) after cloprostenol injection. Blood samples were collected before injection and at frequent intervals after injection. Distribution of PKC activity in cytosol and membrane fractions and activities of microcalpain, millicalpain, and calpastatin were determined for all CL. OT was measured in plasma and tissue by RIA. Relative to mean plasma levels of OT at time zero (85 +/- 7 pg/ml), peak plasma levels occurred between 1.5-10 min (270 +/- 36 pg/ml) for all animals. The mean luteal concentration of OT was greater at 0, 2, and 7.5 min (145 +/- 27, 232 +/- 82 and 269 +/- 115 ng/g, respectively) than at 30 min (93 +/- 33), but differences in tissue OT over time were not significant (P > 0.05). PKC activities (percentage over nonactivated control values) in the membrane or cytosolic fractions did not differ significantly among the times of CL removal; however, membrane PKC activity was positively correlated with the plasma OT level at the time of CL removal (r = 0.82; P < 0.0025). Luteal millicalpain activity was approximately twice that of microcalpain at each time point (P < 0.001), although the activities of the individual calpains over time after PGF2 alpha injection did not change. Calpastatin activity was significantly higher at 30 min (515 +/- 28 U/g tissue) than at 0, 2, or 7.5 min (373 +/- 26, 423 +/- 26, and 426 +/- 24 U/g tissue, respectively). PKC activity in the membrane appears to be positively correlated with OT secretion from the bovine CL, and increased calpastatin activity after PGF2 alpha injection may inhibit calpains present in the CL, thereby maintaining an active pool of PKC.

Effects of dexamethasone on muscle protein homeostasis and on calpain and calpastatin activities and gene expression in rabbits.

The objectives were to investigate the mechanisms by which glucocorticoids control proteolysis in muscle cells and the relationship between the calpain:calpastatin system and proteolysis in muscle. Female rabbits were treated with 1 mg dexamethasone (Dex)/kg body weight per day for 0, 1, 2 or 4 days after which animals were killed and muscle samples taken for analyses. Dex reduced urinary N tau-methylhistidine (NMH) 48% (day 4 versus day 1 of Dex treatment) and muscle NMH concentrations by 49% (day 1) to 40% (day 2) respectively, suggesting that protein degradation was reduced. To investigate whether the changes in apparent proteolysis were related to calpains, we examined the effects of Dex on muscle calpain and calpastatin activities. These were unaffected by Dex. This implies that Dex-dependent changes in degradation are not mediated by changes in muscle calpain or calpastatin activities. We studied the effects of Dex on calpain and calpastatin gene expression as a means of clarifying the relationships between proteinase gene expression and proteinase activities. mu-Calpain mRNA concentration was unaffected by Dex but m-calpain mRNA and calpastatin mRNA concentrations were reduced by 42-55% and 40% respectively. Dex had a similar effect on beta-actin mRNA. Although calpain and calpastatin genes behaved as house-keeping genes, changes in their expression mimicked apparent changes in proteolysis. The observation that calpain and calpastatin activities were unchanged indicates that additional regulation of the calpain:calpastatin system exists at other sites in muscle cells. To determine whether Dex-dependent changes in proteolysis were mediated indirectly, we assayed the effects of Dex on plasma thyroid hormone concentrations.(ABSTRACT TRUNCATED AT 250 WORDS)

Effects of dexamethasone on protein degradation and protease gene expression in rat L8 myotube cultures.

We examined the effects of a synthetic glucocorticoid (dexamethasone; Dex) on protoeolysis and on protease messenger RNA (mRNA) concentrations in rat L8 skeletal myotube cultures. Protein degradation was measured as release of radioactive trichloroacetic acid-soluble material from intracellular proteins pre-labelled with [3H]tyrosine. Dex (1 microM) stimulated protein degradation (P < 0.01). This effect was entirely blocked by the glucocorticoid antagonist, RU38486 (mifepristone; P < 0.01). Hence, actions of Dex on muscle protein degradation are mediated via intracellular glucocorticoid receptors. Molecular mechanisms by which glucocorticoids stimulate protein degradation in skeletal muscle are not known. Here, we investigated the regulation of protease (cathepsin B, cathepsin D, proteasome C2 subunit and m-calpain) mRNA concentrations by Dex in cultured L8 muscle cells. Cathepsin B mRNA concentration was enhanced 3.3-fold by Dex. This effect was blocked by RU38486. RU38486 alone did not affect cathepsin B mRNA concentration or mRNAs of other proteases. Concentrations of cathepsin D and m-calpain mRNAs were also increased by Dex. These effects were also abolished by RU38486. Proteasome C2 mRNA was unaffected by Dex and Dex reduced alpha-tubulin mRNA. Thus, glucocorticoids specifically regulate the concentrations of mRNAs encoding some proteases in muscle cells. The regulation of protease mRNA concentration is mediated via interaction between Dex with glucocorticoid receptors and is independent of the actions of Dex on mRNA encoding house-keeping proteins. These changes may underlie glucocorticoid-dependent control of proteolysis in muscle.

Effects of cimaterol on muscle protein metabolism and its actions in hypothyroid and hyperthyroid rats.

Objectives were to examine mechanisms underlying anabolic actions of cimaterol in skeletal muscle and to evaluate cimaterol's actions in hypothyroid and hyperthyroid rats. In the first study growing rats were fed either a control diet or a diet containing cimaterol for 10 days. In a second study sham-thyroidectomized and thyroidectomized (Tx) rats were assigned to one of 5 treatments: control (sham-Tx), Tx, Tx supplemented with cimaterol, Tx injected with triiodothyronine (T3), and Tx rats injected with T3 and supplemented with cimaterol. Effects of treatments on growth, muscle weights and urinary NT-methylhistidine (NMH) excretion were evaluated in both trials. Muscle was also collected for determinations of DNA, RNA, protein and activities of several proteolytic enzymes. Cimaterol caused muscle hypertrophy and increased urinary NMH excretion. Hence, anabolic actions of cimaterol may result from an increase in myofibrillar protein synthesis which exceeds changes in myofibrillar protein degradation. Urinary NMH excretion was reduced by thyroidectomy and increased in hyperthyroid rats and both hypothyroidism and hyperthyroidism were characterized by myopathy. Cimaterol increased muscle weights in hypothyroid but not in hyperthyroid rats. Therefore, cimaterol's anabolic properties are thyroid hormone-independent and antagonized by excess thyroid hormone. Anabolic actions of cimaterol in hypothyroid rat muscle were attributed to an action on protein synthesis because urinary NMH excretion was not affected by cimaterol but muscle RNA concentration was increased. Activities of cathepsins B, D and L and neutral proteinase were dose-related to thyroid status, however, were unrelated to cimaterol-dependent perturbations in NMH excretion. Control of muscle protein balance by thyroid hormones may involve regulation of these enzymes; however, control of muscle protein degradation by cimaterol is likely directed towards other proteolytic mechanisms or to mechanisms which alter susceptibility of myofibrillar proteins to degradation.

Effects of serum and insulin-like growth factor I on protein degradation and protease gene expression in rat L8 myotubes.

We examined the effects of horse and fetal bovine sera and insulin-like growth factor I (IGF-I) on proteolysis and protease gene expression in rat L8 skeletal myotube cultures. Protein degradation was measured as release of radioactive trichloroacetic acid (TCA)-soluble materials from intracellular proteins prelabeled with [3H]tyrosine. Horse serum and fetal bovine serum inhibited (P < .05) protein degradation by 19.7 and 8.1%, respectively. The IGF-I at 200 ng/mL inhibited protein degradation by 14% (P < .01) over a 6-h measurement period. To study the regulation of proteolysis by IGF-I, we evaluated its effects on protease mRNA and alpha-tubulin mRNA concentrations by Northern blot analysis. Proteases under investigation included cathepsins B and D, proteasome C2 subunit, and m-calpain. The IGF-I had no effect (P > .05) on cathepsin B and D gene expression but slightly increased (P < .05) m-calpain and alpha-tubulin mRNA concentrations. Proteasome mRNA concentration was reduced (P < .05) by IGF-I treatment. The changes in proteasome mRNA levels paralleled the IGF-I-dependent alterations in proteolysis. These observations suggest that effects of IGF-I on muscle protein degradation may be mediated by the specific down-regulation of proteasomal subunit mRNAs.

Internationalization of the animal science undergraduate curriculum: a survey of its current status, barriers to its implementation and its value.

The goal of this project was to identify the current level at which internationalization has been adopted as a theme in the North American animal science curriculum and to identify its value and the barriers to its implementation. We surveyed animal, dairy, and poultry science departments across Canada and the United States. One hundred twenty-four surveys were mailed and 60% were returned. Associations between aspects of internationalization and student outcomes (admission to veterinary and graduate schools and starting salaries) were examined. Although administrators strongly believed internationalization had value, implementation was limited. The most common practices included international content in core animal science classes, advising, international internships, and participation of faculty in international scholarly activities. Few departments have incorporated internationalization into their mission statements or developed a specific international-themed class, scholarships devoted to international activities, or roles for international students. Few departments reported participation of students in international programs. Barriers included finances and limited commitment from higher administration. Student outcomes were positively associated with faculty size, percentage of international faculty, the ratio of international students to the total student population, international content in core animal science classes, a specific international-themed class, availability of international internships, and exchange of class material internationally via the Internet. Departments that did not offer international opportunities had a negative association (r = -0.79) with starting salary, but these relationships may not be causal. Alternatively, progressive departments may attract and retain exceptional students. The analysis indicated an awareness of the value of international programs, positive impacts in student outcomes, and financial barriers to implementation.

Effects of age and castration on activities of calpains and calpastatin in sheep skeletal muscle.

The objectives of this experiment were to assess effects of animal age and castration on activities of calpain I, calpain II, and calpastatin in sheep skeletal muscle. Ten newborn male lambs (2.9 kg), six weaned wethers (23.2 kg), six weaned rams (22.2 kg), six market wethers (55.4 kg), and six market rams (60.2 kg) were slaughtered and samples of biceps femoris were taken for assay of calpain I (micromolar calcium-dependent proteinase), calpain II (millimolar calcium-dependent proteinase), and calpastatin. Preweaning weight gain was similar for rams and wethers; however, postweaning ram growth exceeded (P less than .05) that of wethers. Ram biceps femoris weights at market were greater than those of wethers (P less than .05). Irrespective of age or gender, activity of calpain II was two- to threefold greater than that of calpain I. Muscle calpastatin activity was severa fold higher than calpain I and II activities. Activities of calpains and calpastatin declined (P less than .05) between birth and weaning. A portion of these losses were due to a dilution effect caused by accumulation of muscle proteins. Neonatal attenuation of calpain activities may underlie age-related attenuation of fractional rates of muscle protein degradation. Although ram muscle growth exceeded that of wethers, no differences (P greater than .05) in activities of muscle calpains or calpastatin were detected between these two groups at weaning or at market weight. Hence, castration did not influence lamb muscle growth by altering muscle calpain or calpastatin activities.

Effects of cimaterol on rabbit growth and myofibrillar protein degradation and on calcium-dependent proteinase and calpastatin activities in skeletal muscle.

The objectives of this study were to examine effects of a beta-adrenergic agonist (cimaterol) on growth and muscle development in rabbits and to examine cimaterol's effects on myofibrillar protein degradation (MPD) and on activities of several proteolytic enzymes including the calcium-dependent proteinases (CDP). Twelve New Zealand White rabbits were assigned to either control diets or to diets containing cimaterol for 35 d, after which they were killed and effects on performance and tissue weight gains were determined. Urine was collected from d 21 through 28 from each rabbit for assessment of N tau-methylhistidine (NMH) excretion. Cimaterol increased rates of gain, efficiency of gain and skeletal muscle weights. Enhancement in muscle weight was associated with an increase in total DNA and with a reduction in NMH. Cimaterol did not affect activities of cathepsin B, cathepsin D or neutral serine proteinase, but it reduced activities of the millimolar and micromolar forms of the CDP by 58 and 57%, respectively, and it reduced activity of the inhibitor of the CDP (calpastatin) by 52%. Cimaterol-dependent myofibrillar protein accretion was likely mediated, at least in part, by a reduction in MPD. The change in MPD was associated with a reduction in muscle CDP activities. Cimaterol-dependent muscle hypertrophy therefore may involve changes in calcium-dependent proteolysis of myofibrillar proteins. The significance of the effects of cimaterol on calpastatin activity is not known.

Effect of cimaterol on sheep adipose tissue lipid metabolism.

Effects of dietary cimaterol (5 mg/kg) on adipose tissue metabolism of wether lambs were studied. Lipogenesis, lipolysis, fatty acid composition and adipocyte size and number were measured. Cimaterol feeding increased lipogenesis; however, this effect was not statistically significant. Insulin (1,000 microU/ml) stimulated lipogenesis of adipose tissue from control sheep. However, this elevated rate was abolished by in vitro cimaterol. Insulin had no stimulatory effect on lipogenesis in cimaterol-fed sheep. Lipolysis was depressed by cimaterol feeding. However, 10(-4) M cimaterol stimulated lipolysis in the adipose tissue from both control and cimaterol-fed sheep. Insulin inhibited stimulated lipolysis in adipose tissue from control sheep but had no effect on the stimulated lipolysis in cimaterol-fed sheep. Mean adipocyte diameter was smaller (from 74 to 70 microns) and adipocyte size distribution also was changed in the cimaterol-fed sheep. Adipocyte number per gram of tissue was not affected by cimaterol. There was a significant increase in percentage of unsaturated fatty acids in adipose tissue from cimaterol-fed sheep. These results indicate that lipogenic and lipolytic responses to insulin and cimaterol in sheep adipose tissue were altered by cimaterol feeding. The carcass fat content decrease in cimaterol-fed sheep may be attributed to the reduction in adipocyte size.

Structural and metabolic integrity of sheep external intercostal muscle during extended culture.

The objectives of this study were to examine the structural and metabolic integrity of isolated sheep external intercostal muscle bundles following variable lengths of preincubation (0 to 192 h). Samples of intact external intercostal muscle (10 to 15 g), with tendons attached, were prepared from growing wethers and maintained at their resting lengths during preincubation for 0 to 192 h. Protein synthesis (PS), protein degradation (PD), acetate oxidation and ultrastructural integrity of muscle samples were examined at 0 to 192 h, 0 to 96 h, 0 to 48 h and 0 to 96 h following isolation, respectively. Additionally, the effects of variable fetal calf serum (FCS) concentrations (0 to 20%; w/v) on PS and PD and acetate oxidation were examined. Rate of PS increased as preincubation time increased to 192 h; however, most of this increase was due to the proliferation of fibroblasts on the surface of the muscle sample. Addition of cytosine arabinoside to the incubation media prevented the fibroblast-dependent increase in PS; however, it did not entirely prevent the preincubation time-dependent increase in PS. Rate of PD increased greatly upon preincubation. The nitrogen balance of incubated muscles was negative at all times examined. Acetate oxidation was maintained through 12 h of preincubation and thereafter declined. Relatively normal myofibrillar structure was maintained through 48 h of preincubation; however, loss of mitochondrial integrity and dissolution of Z-disks at 48 h and at 96 h of preincubation were evident. Isolated tissues were able to respond to FCS concentration in medium following 48 h of preincubation.

Effects of the callipyge phenotype on serum creatinine, total cholesterol, low-density lipoproteins, very-low-density lipoproteins, high-density lipoproteins, and triacylglycerol in growing lambs.

The goals of this study were to investigate the effects of the callipyge (CLPG) phenotype on serum creatinine and lipid profiles of growing lambs. Preliminary studies in our laboratories indicated that creatinine may have utility in distinguishing the CLPG phenotype and that expression of the CLPG gene altered concentrations of serum total cholesterol (TC). As a result, in this study, we examined the influence of the CLPG gene on concentrations of creatinine, TC, very-low-density lipoproteins (VLDL), low-density lipoproteins (LDL), high-density lipoproteins (HDL), and triacylglycerol (TG) at varying stages of maturity in lambs. Ten homozygous (c/c) Polypay ewes were crossed with Dorset rams heterozygous for the CLPG gene (C/c). From this cross, 20 lambs (13 females and 7 males) were born, of which 11 were homozygotic (c/c) and 9 were heterozygotic (C/c; CLPG) based on muscle weights and longissimus dorsi (LD) area at slaughter. Blood samples were taken at monthly intervals and serum lipid constituents were assayed. At 1 mo of age, no differences (P > .05) in plasma lipids were detectable between phenotypes. However, at 2 mo age, CLPG lambs had higher (P < .01) concentration of TG, TC, HDL, and VLDL compared to homozygotic (c/c) lambs. Triglycerides and VLDL were elevated (P < .05) in CLPG lambs at 3 mo of age. By slaughter, no differences (P > .05) in serum lipid constituents were detectable between genotypes. Hence, the increase in serum TC is due to elevated levels of HDL and VLDL. These observations indicate that creatinine may be used to distinguish CLPG lambs and that the CLPG gene alters serum lipid profiles during the postnatal period.

The effects of selenium deficiency on differentiation, degradation, and cell lysis of L8 rat skeletal muscle cells.

We investigated the effects of selenium (Se) deficiency on differentiation, protein degradation, and cell lysis in cultured skeletal muscle cells, using L8 rat skeletal muscle cells cultured in serum-free (SF) medium to induce differentiation and to maintain myotubes. Creatine kinase activity was reduced (p < 0.05) by approximately 15% without Se supplementation for 96 h. Confluent myoblasts were treated with SF media with four different levels of vitamin E (0, 10, 35, and 100 microM) in the absence and presence of Se (0 and 0.25 microM, respectively). After 96 h, vitamin E at a high dose (100 microM) was effective in the prevention of the decrease of differentiation caused by Se deficiency (p < 0.05). Following differentiation, the effects of three Se concentrations (0, 0.25, and 2.5 microM) on degradation of proteins as assessed by release of 3H-labeled free amino acids secreted into the media were studied. Selenium supplementation did not affect (p > 0.05) total protein degradation. However, Se deficiency increased (p < 0.05) lactate dehydrogenase released from lyzed dead cells. The results indicate that Se is required to maintain an optimal rate of muscle cell differentiation and health of myotube cultures.

Effect of dietary supplementation on the antimicrobial activity of blood leukocytes isolated from Holstein heifers.

The purpose of this investigation was to evaluate the effect of an immunostimulating feed supplement (OmniGen-AF®) on the antimicrobial properties of blood leukocytes in dairy heifers in an attempt to prevent mastitis. Blood leukocytes from supplemented and unsupplemented controls were used. Phagocytic activity and reactive oxygen species (ROS) production were studied on d 0 (prior to feed supplementation) and on days 30 and 60 after supplementation. L-selectin and IL-8R mRNA expressions on blood leukocytes were evaluated on d 0 (prior to feed supplementation) and monthly thereafter for 15 mo. On d 30 after supplementation, neutrophils from treated heifers exhibited greater binding and internalization of Escherichia coli and greater ROS production compared with unsupplemented controls. L-selectin mRNA expression was increased in supplemented heifers vs. controls; however, IL-8R mRNA expression was not different. Results support the continued study of dietary supplementation as an additional management tool to enhance udder health in dairy heifers.

Organic and inorganic selenium: II. Transfer efficiency from ewes to lambs.

Adequate Se transfer from ewes to lambs is important to prevent Se-deficiency diseases. To evaluate how different chemical forms of Se administered at comparative dosages to mature ewes affect Se status of their lambs, 240 ewes were divided into 8 treatment groups (n = 30 each) and drenched weekly (at an amount equal to their summed daily intake) with no-Se (controls); at recommended amounts (4.9 mg of Se/wk) with inorganic Na-selenite, inorganic Na-selenate, or organic Se-yeast; or at supranutritional amounts (14.7 and 24.5 mg of Se/wk) with Na-selenite or Se-yeast for 1 yr. Weekly drenching of Se was effective at increasing (P < 0.002) Se concentrations in ewe colostrum and milk at 30 d of lactation and in improving (P < 0.001) the Se status of lambs (whole-blood and serum-Se concentrations at birth, and skeletal-muscle Se concentrations at 14 d of age). Selenium concentrations in lacteal secretions were greater in ewes drenched with Se-yeast (colostrum: 374, 436, and 982 ng/mL at 4.9, 14.7, and 24.5 mg of Se/wk, respectively; milk: 26, 39, 64 ng/mL) compared with ewes drenched with Na-selenite (colostrum: 204, 334, 428 ng/mL; milk: 16, 21, 24 ng/mL), and were also greater (P < 0.001) in their lambs. Selenium concentrations continued to increase (P < 0.001) in lamb whole blood (558 and 695 ng/mL at 14.7 and 24.5 mg of Se/wk, respectively), serum (126, 183 ng/mL), and skeletal muscle (991, 1,696 ng/mL) with supranutritional concentrations of Se-yeast, whereas Se concentrations did not differ in whole blood (304, 332 ng/mL), serum (77, 85 ng/mL), or skeletal muscle (442, 482 ng/mg) of lambs from ewes drenched with 14.7 or 24.5 mg of Se/wk of Na-selenite. We conclude that weekly oral drenching of ewes during gestation and lactation with organic Se-yeast results in a more efficient transfer of Se (over a wide range of supplementation rates) from ewe to lamb than does inorganic Na-selenite.