Reducing gut effects from Cryptosporidium parvum infection in dairy calves through prophylactic glucagon-like peptide 2 therapy or feeding of an artificial sweetener.

Glucagon-like peptide 2 (GLP-2) therapy was shown previously to reduce inflammation-related gut damage from coccidiosis in dairy calves, and feeding of artificial sweetener stimulates GLP-2 secretion from intestinal L cells. The purpose of this study was to determine whether GLP-2 treatment or artificial sweetener feeding beginning 1 wk before an experimental inoculation with the coccidian parasite Cryptosporidium parvum can reduce infection-related intestinal damage in Holstein bull calves. Newborn calves were assigned to 1 of 4 treatment groups of 6 calves each, including noninfected control calves injected s.c. every 12 h with control buffer (CON), infected control calves injected s.c. every 12 h with control buffer (INF), infected calves injected s.c. every 12 h with 50 µg/kg of body weight of GLP-2 (GLP2), and infected calves injected s.c. every 12 h with control buffer and supplemented in the diet with Sucram (Pancosma, Geneva, Switzerland) at 400 mg/kg of dry matter of milk replacer (SUC). Treatments were initiated on d 1, and calves in INF, GLP2, and SUC were orally dosed on d 8 with 12,500 C. parvum oocysts. Fecal scores were recorded daily, plasma was collected on d 1, 8, 12, 15, and 18 to evaluate markers of inflammation, and fecal samples were collected on d 1, 8, and every other day thereafter to determine the presence of oocysts. Calves were euthanized on d 18 for collection of intestinal tissues and histological and gene expression analyses. Relative to CON, calves in INF exhibited an increase in diarrhea severity, increased plasma serum amyloid A concentration on d 15 and 18, reduced intestinal villus height, increased villus apoptosis and crypt cell proliferation, and increased intestinal mRNA expression of MARVELD2 and GPX2. However, calves in SUC and GLP2 had reduced diarrhea severity and fecal C. parvum oocyst shedding, reduced plasma serum amyloid A concentration on d 15 and 18, and, depending on the intestinal segment, increased villus height, reduced crypt cell proliferation, and reduced mRNA expression of MARVELD2, GPX2, and other tight junction proteins relative to INF. Lastly, GLP2 and SUC exhibited increased intestinal mass-to-length ratio and decreased length-to-empty body weight ratio relative to INF. Our findings suggest that GLP-2 and Sucram treatments administered before a low-level C. parvum exposure may contribute to fewer effects on intestinal integrity, morphology, and inflammation in response to infection, and shorter, denser intestines.

Glucagon-like peptide 2 and its beneficial effects on gut function and health in production animals.

Numerous endocrine cell subtypes exist within the intestinal mucosa and produce peptides contributing to the regulation of critical physiological processes including appetite, energy metabolism, gut function, and gut health. The mechanisms of action and the extent of the physiological effects of these enteric peptides are only beginning to be uncovered. One peptide in particular, glucagon-like peptide 2 (GLP-2) produced by enteroendocrine L cells, has been fairly well characterized in rodent and swine models in terms of its ability to improve nutrient absorption and healing of the gut after injury. In fact, a long-acting form of GLP-2 recently has been approved for the management and treatment of human conditions like inflammatory bowel disease and short bowel syndrome. However, novel functions of GLP-2 within the gut continue to be demonstrated, including its beneficial effects on intestinal barrier function and reducing intestinal inflammation. As knowledge continues to grow about GLP-2's effects on the gut and its mechanisms of release, the potential to use GLP-2 to improve gut function and health of food animals becomes increasingly more apparent. Thus, the purpose of this review is to summarize: (1) the current understanding of GLP-2's functions and mechanisms of action within the gut; (2) novel applications of GLP-2 (or stimulators of its release) to improve general health and production performance of food animals; and (3) recent findings, using dairy calves as a model, that suggest the therapeutic potential of GLP-2 to reduce the pathogenesis of intestinal protozoan infections.

COMPARATIVE GUT PHYSIOLOGY SYMPOSIUM: Comparative physiology of glucagon-like peptide-2: Implications and applications for production and health of ruminants.

Glucagon-like peptide-2 (GLP-2) is a 33-amino acid peptide derived from proteolytic cleavage of proglucagon by prohormone convertase 1/3 in enteroendocrine L cells. Studies conducted in humans, in rodent models, and in vitro indicate that GLP-2 is secreted in response to the presence of molecules in the intestinal lumen, including fatty acids, carbohydrates, amino acids, and bile acids, which are detected by luminal chemosensors. The physiological actions of GLP-2 are mediated by its G protein-coupled receptor expressed primarily in the intestinal tract on enteric neurons, enteroendocrine cells, and myofibroblasts. The biological activity of GLP-2 is further regulated by dipeptidyl peptidase IV, which rapidly cleaves the N-terminus of GLP-2 that is responsible for GLP-2 receptor activation. Within the gut, GLP-2 increases nutrient absorption, crypt cell proliferation, and mesenteric blood flow and decreases gut permeability and motility, epithelial cell apoptosis, and inflammation. Outside the gut, GLP-2 reduces bone resorption, can suppress appetite, and is cytoprotective in the lung. Thus, GLP-2 has been studied intensively as a therapeutic to improve intestinal function of humans during parenteral nutrition and following small bowel resection and, more recently, as a treatment for osteoporosis and obesity-related disorders and to reduce cellular damage associated with inflammation of the gut and lungs. Recent studies demonstrate that many biological actions and properties of GLP-2 in ruminants are similar to those in nonruminants, including the potential to reduce intestinal nitro-oxidative stress in calves caused by parasitic diseases such as coccidiosis. Because of its beneficial impacts on nutrient absorption, gut healing, and normal gut development, GLP-2 therapy offers significant opportunities to improve calf health and production efficiency. However, GLP-2 therapies require an extended time course to achieve desired physiological responses, as well as daily administration because of the hormone's short half-life. Thus, practical means of administration and alternative strategies to enhance basal GLP-2 secretion (e.g., through specific feed additives), which are more likely to achieve consumer acceptance, are needed. Opportunities to address these challenges are discussed.

Short communication: Glucagon-like peptide-2 and coccidiosis alter tight junction gene expression in the gastrointestinal tract of dairy calves.

Tight junction (TJ) proteins are integral factors involved in gut barrier function, and therapy with glucagon-like peptide-2 (GLP-2) enhances gut integrity. Our aim was to assess effects of GLP-2 treatment on mRNA expression of 8 TJ complex proteins in the intestine of dairy calves not infected or infected with Eimeria bovis at 11±3d of age. Mucosal epithelium from jejunum, ileum, and cecum was collected at slaughter from Holstein bull calves assigned to 4 groups: noninfected, buffer-treated (n=5); noninfected, GLP-2 treated (n=4); E. bovis-infected, buffer-treated (n=5); and E. bovis-infected, GLP-2-treated (n=4). Infected calves were orally dosed with 100,000 to 200,000 sporulated E. bovis oocysts on d 0; GLP-2-treated calves received 50 µg of GLP-2/kg of body weight subcutaneously twice daily for 10d beginning on d 18; and buffer-treated calves received an equal injection volume of 0.01 M Na bicarbonate buffer. All calves were killed on d 28. The mRNA expression of coxsackie and adenovirus receptor (CXADR), claudins 1, 2, and 4 (CLDN1, CLDN2, and CLDN4), F11 receptor (F11R), junction adhesion molecule 2 (JAM2), occludin (OCLN), and tight junction protein ZO-1 (TJP1) was determined by real-time quantitative PCR. In jejunum and ileum, an interaction of E. bovis infection and GLP-2 treatment on gene expression was noted. In jejunum of noninfected calves, GLP-2 increased CXADR, CLDN2, OCLN, and TJP1 mRNA expression but had no effect on mRNA expression in infected calves. Treatment with GLP-2 also increased tight junction protein ZO-1 protein expression in jejunum of noninfected calves as determined by immunohistochemistry. In ileum, E. bovis decreased expression of JAM2, OCLN, and TJP1 in buffer-treated calves, and GLP-2 increased TJP1 expression in infected calves. In cecum, E. bovis infection reduced expression of CXADR, CLDN4, F11R, and OCLN, and GLP-2 therapy increased expression of CLDN4, F11R, OCLN, and TJP1. Results are consistent with studies in nonruminants showing decreased expression of TJ complex proteins in the intestinal tract during pathogen-induced diarrhea and increased TJ protein expression in intestinal tissues in response to GLP-2 treatment. In conclusion, E. bovis reduces gene expression of TJ proteins primarily in cecum of calves 28d postinfection, and GLP-2 increases expression of selected TJ genes in intestinal tissues. Use of GLP-2 to improve gut barrier function in ruminants during pathogen-induced diarrhea warrants additional study.

Glucagon-like peptide 2 therapy reduces negative effects of diarrhea on calf gut.

Damage to the intestinal epithelium reduces nutrient absorption and animal growth, and can have negative long-term health effects on livestock. Because the intestinotropic hormone glucagon-like peptide 2 (GLP-2) has been shown to contribute to gut integrity, reduce inflammation, and improve nutrient absorption, the present study was designed to determine whether administration of GLP-2 to calves with coccidiosis in the first month of life affects intestinal growth and mediates negative effects of the proinflammatory response. Holstein bull calves (n=19) were assigned to 4 treatment groups of 4 to 5 calves each: (1) infected with Eimeria bovis, GLP-2 treated; (2) noninfected, GLP-2 treated; (3) infected with E. bovis, buffer treated; and (4) noninfected, buffer treated. Infected calves received 100,000 to 200,000 sporulated E. bovis oocysts suspended in milk replacer on d 0 of the study. On d 18, calves in the GLP-2 groups received a subcutaneous injection of 50 µg of bovine GLP-2/kg of body weight twice daily for 10 d, and calves in the buffer-treated groups received an equivalent volume of sodium bicarbonate buffer only. On d 28, calves were slaughtered 2h after injection of 5-bromo-2'-deoxyuridine (BrdU). Intestinal tissues were measured and villus height, crypt depth, and BrdU immunostaining were evaluated in segments of the small intestine. Nitrotyrosine immunostaining, a measure of nitro-oxidative damage, was evaluated in the ileum and cecum. No GLP-2 treatment by E. bovis infection interaction was observed for any parameter measured, with the exception of nitrotyrosine immunostaining in the cecum. Large intestinal weight was greater in infected than noninfected calves and with GLP-2 treatment relative to buffer treatment. Calves that received GLP-2 also had greater small intestinal weight but no difference in cell proliferation, as assessed by BrdU labeling, relative to buffer-treated calves. No treatment effects were detected for villus height, crypt depth, or villus height:crypt depth ratio in segments of the small intestine. Protein tyrosine nitration was over 3-fold greater in the ileum and cecum of infected calves relative to noninfected calves, and GLP-2 therapy reduced tyrosine nitration in infected calves by 47% in the ileum and 69% in the cecum relative to buffer-treated calves. Treatment with GLP-2 promotes intestinal growth in neonatal calves and reduces the detrimental effects of nitro-oxidative stress in the ileocecum of calves with coccidiosis.

Bovine mammary stem cells: cell biology meets production agriculture.

Mammary stem cells (MaSC) provide for net growth, renewal and turnover of mammary epithelial cells, and are therefore potential targets for strategies to increase production efficiency. Appropriate regulation of MaSC can potentially benefit milk yield, persistency, dry period management and tissue repair. Accordingly, we and others have attempted to characterize and alter the function of bovine MaSC. In this review, we provide an overview of current knowledge of MaSC gained from studies using mouse and human model systems and present research on bovine MaSC within that context. Recent data indicate that MaSC retain labeled DNA for extended periods because of their selective segregation of template DNA strands during mitosis. Relying on this long-term retention of bromodeoxyuridine-labeled DNA, we identified putative bovine MaSC. These label-retaining epithelial cells (LREC) are in low abundance within mammary epithelium (<1%). They are predominantly estrogen receptor (ER)-negative and localized in a basal or suprabasal layer of the epithelium throughout the gland. Thus, the response of MaSC to estrogen, the major mitogen in mammary gland, is likely mediated by paracrine factors released by cells that are ER-positive. This is consistent with considerable evidence for cross-talk within and between epithelial cells and surrounding stromal cells. Excision of classes of cells by laser microdissection and subsequent microarray analysis will hopefully provide markers for MaSC and insights into their regulation. Preliminary analyses of gene expression in laser-microdissected LREC and non-LREC are consistent with the concept that LREC represent populations of stem cells and progenitor cells that differ with regard to their properties and location within the epithelial layer. We have attempted to modulate the MaSC number by infusing a solution of xanthosine through the teat canal and into the ductal network of the mammary glands of prepubertal heifers. This treatment increased the number of putative stem cells, as evidenced by an increase in the percentage of LREC and increased telomerase activity within the tissue. The exciting possibility that stem cell expansion can influence milk production is currently under investigation.

Characterization of glucagon-like peptide 2 pathway member expression in bovine gastrointestinal tract.

Glucagon-like peptide 2 (GLP-2), secreted by enteroendocrine cells, has several physiological effects on the intestine of monogastric species, including promotion of growth of intestinal epithelium, reduction of epithelial cell apoptosis, and enhancement of intestinal blood flow, nutrient absorption, and epithelial barrier function. The regulatory functions of GLP-2 in the ruminant gastrointestinal tract (GIT) have not been well studied. The objectives of this investigation were to characterize the mRNA expression of 4 members of the GLP-2 pathway throughout the bovine GIT, including (1) proglucagon (GCG), the parent peptide from which GLP-2 is derived through cleavage by prohormone convertase; (2) prohormone convertase (PCSK1); (3) GLP-2 receptor (GLP2R); and (4) dipeptidyl peptidase IV (DPP4), the enzyme that inactivates GLP-2. Gene expression was evaluated in rumen, reticulum, omasum, abomasum, duodenum, jejunum, ileum, cecum, and rectum collected at slaughter from prepubertal heifers, mature cows in early, mid, and late lactation, and nonlactating cows (n=3 per stage) by a gene expression profiling assay. In addition, mRNA expression of 14 genes involved in nutrient transport, enzyme activity, blood flow, apoptosis, and proliferation were evaluated in the 9 GIT tissues for their association with GCG and GLP2R mRNA expression. Immunohistochemistry was used to localize GLP2R protein in tissues of the lower GIT. Results indicated that mRNA expression of GCG, PCSK1, GLP2R, and DPP4 varies across the 9 GIT tissues, with greatest expression in small and large intestines, and generally nondetectable levels in forestomachs. Expression of DPP4 and GLP2R mRNA varied by developmental stage or lactational state in intestinal tissues. Expression of GCG or GLP2R mRNA was correlated with molecular markers of proliferation, apoptosis, blood flow, enzyme activity, and urea transport, depending on the tissue examined, which suggests a potential for involvement of GLP-2 in these physiological processes in the ruminant GIT. The GLP2R protein was expressed in intestinal crypts of the bovine GIT, which is consistent with the distribution in monogastric species. Our findings support a functional role of the GLP-2 pathway in bovine GIT and the potential for use of GLP-2 as a therapy to improve intestinal function and nutrient absorption in ruminants.

Technical note: A rapid method for 5-bromo-2'-deoxyuridine (BrdU) immunostaining in bovine mammary cryosections that retains RNA quality.

A rapid method of 5-bromo-2'-deoxyuridine (BrdU) immunostaining was developed in cryosections of bovine mammary tissue while preserving RNA quality of the stained section. A thymidine analog that is incorporated into DNA of proliferating cells, BrdU serves as a proliferation marker. Immunostaining of BrdU-labeled cells within a histological section requires heat, enzymatic or chemical-mediated antigen retrieval to open double-stranded DNA, and exposure to the BrdU antigen. Although these established treatments permit staining, they preclude use of cells within the tissue section for further gene expression experiments. Additionally, long antibody incubations and washing steps lead to extensive RNA degradation and elution. A protocol was developed for immunolocalization of BrdU-labeled cells in cryosections of bovine mammary tissue, which does not require harsh DNA denaturation and preserved RNA integrity and quantity. This protocol used an initial acetone:polyethylene glycol 300 [9:1 (vol/vol)] fixation (2 min) followed by staining with methyl green (0.5% aqueous; 2 min) to stabilize macromolecules, antigen retrieval with deionized formamide (70% in nuclease-free phosphate buffered saline; 4 min incubation), antibody incubation in the presence of RNase inhibitors (5 min), and minimal washing to facilitate recovery of RNA from cells from the stained sections. Applicability of this protocol to other nuclear antigens was evaluated by testing its suitability for staining estrogen receptor alpha and Ki-67 antigen. In both cases, use of the protocol provided good immunostaining and tissue morphology. The RNA quality of estrogen receptor alpha- and Ki-67-stained sections was not evaluated. Quality of the isolated RNA from BrdU-stained sections was evaluated by micro-fluidic electrophoresis and its utility was confirmed using quantitative reverse transcription-PCR. Staining intensity obtained with this labeling protocol was similar to that obtained using conventional immunohistochemistry protocols. When coupled with laser microdissection and RNA or cDNA amplification, this immunostaining protocol provided a means for future transcriptome analysis of BrdU-labeled cells within a complex tissue.

Genome-wide linkage analysis to identify chromosomal regions affecting phenotypic traits in the chicken. III. Skeletal integrity.

Two unique chicken F(2) populations generated from a broiler breeder male line and 2 genetically distinct inbred (>99%) chicken lines (Leghorn and Fayoumi) were used for whole genome QTL analysis. Twelve phenotypic skeletal integrity traits (6 absolute and 6 relative traits) were measured or calculated, including bone mineral content, bone mineral density, tibia length, shank length, shank weight, and shank length:shank weight. All traits were also expressed as a percentage of BW at 8 wk of age. Birds were genotyped for 269 microsatellite markers across the entire genome. The QTL affecting bone traits in chickens were detected by the QTL express program. Significance levels were obtained using the permutation test. For the 12 traits, a total of 56 significant QTL were detected at the 5% chromosome-wise significance level, of which 14 and 10 were significant at the 5% genome-wise level for the broiler-Leghorn cross and broiler-Fayoumi cross, respectively. Phenotypic variation for each trait explained by all detected QTL across the genome ranged from 12.0 to 35.6% in the broiler-Leghorn cross and 2.9 to 31.3% in the broiler-Fayoumi cross. Different QTL profiles identified between the 2 related F(2) crosses for most traits suggested that genetic background is an important factor for QTL analysis. Study of associations of biological candidate genes with skeletal integrity traits in chickens will reveal new knowledge of understanding biological process of skeletal homeostasis. The results of the current study have identified markers for bone strength traits, which may be used to genetically improve skeletal integrity in chickens by MAS, and to identify the causal genes for these traits.

Genome-wide linkage analysis to identify chromosomal regions affecting phenotypic traits in the chicken. IV. Metabolic traits.

The current study is a comprehensive genome analysis to detect QTL affecting metabolic traits in chickens. Two unique F(2) crosses generated from a commercial broiler male line and 2 genetically distinct inbred lines (Leghorn and Fayoumi) were used in the present study. The plasma glucagon, insulin, lactate, glucose, tri-iodothyronine, thyroxine, insulin-like growth factor I, and insulin-like growth factor II concentrations at 8 wk were measured in the 2 F(2) crosses. Birds were genotyped for 269 microsatellite markers across the entire genome. The program QTL Express was used for QTL detection. Significance levels were obtained using the permutation test. For the 10 traits, a total of 6 and 9 significant QTL were detected at a 1% chromosome-wise significance level, of which 1 and 6 were significant at the 5% genome-wise level for the broiler-Leghorn cross and broiler-Fayoumi cross, respectively. Most QTL for metabolic traits in the present study were detected in Gga 2, 6, 8, 9, 13, and Z for the broiler-Leghorn cross and Gga 1, 2, 4, 7, 8, 13, 17, and E47 for the broiler-Fayoumi cross. Phenotypic variation for each trait explained by all QTL across genome ranged from 2.73 to 14.08% in the broiler-Leghorn cross and from 6.93 to 21.15% in the broiler-Fayoumi cross. Several positional candidate genes within the QTL region for metabolic traits at the 1% chromosome-wise significance level are biologically associated with the regulation of metabolic pathways of insulin, triiodothyronine, and thyroxine.

Genome-wide linkage analysis to identify chromosomal regions affecting phenotypic traits in the chicken. II. Body composition.

Two informative chicken F(2) populations based on crosses between a broiler breeder male line and dams from genetically distinct, highly inbred (>99%) chicken lines, the Leghorn G-B2 and Fayoumi M15.2, have been used for genome-wide linkage and QTL analysis. Phenotypic data on 12 body composition traits (breast muscle weight, breast muscle weight percentage, abdominal fat weight, abdominal fat weight percentage, heart weight, heart weight percentage, liver weight, liver weight percentage, spleen weight, spleen weight percentage, and drumstick weight, and drumstick weight percentage) were collected. Birds were genotyped for 269 microsatellite markers across the genome. The QTL Express program was used to detect QTL for body composition traits. Significant levels were obtained using the permutation test. For the twelve traits, a total of 61 (Gga 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 12, 15, 18, 24, and Z) and 45 (Gga 1, 2, 3, 4, 6, 7, 8, 9, 10, 12, 15, 17, and E46) significant QTL were detected at the 5% chromosome-wise significance level, of which 19 and 11 were significant at the 5% genome-wise level for the broiler-Leghorn cross and broiler-Fayoumi cross, respectively. Phenotypic variation for each trait explained by all QTL across the genome ranged from 3.22 to 33.31% in the broiler-Leghorn cross and 4.83 to 47.12% in broiler-Fayoumi cross. Distinct QTL profiles between the 2 crosses were observed for most traits. Cryptic alleles were detected for each trait. Potential candidate genes within the QTL region for body composition traits at the 1% chromosome-wise significance level were identified from databases for future association study. The results of the current study will increase the knowledge of genetic markers associated with body composition traits and aid the process of identifying causative genes. Knowledge of beneficial genetic variation can be incorporated in breeding programs to enhance genetic improvement through marker-assisted selection in chickens.

Genome-wide linkage analysis to identify chromosomal regions affecting phenotypic traits in the chicken. I. Growth and average daily gain.

A genome scan was used to detect chromosomal regions and QTL that control quantitative traits of economic importance in chickens. Two unique F(2) crosses generated from a commercial broiler male line and 2 genetically distinct inbred lines (Leghorn and Fayoumi) were used to identify QTL affecting BW and daily average gain traits in chickens. Body weight at 2, 4, 6, and 8 wk was measured in the 2 F(2) crosses. Birds were genotyped for 269 microsatellite markers across the entire genome. Linkage distance among microsatellite markers was estimated by the CRIMAP program. The program QTL Express was used for QTL detection. Significance levels were obtained using the permutation test. For the 8 traits, a total of 18 and 13 significant QTL were detected at a 1% chromosome-wise significance level, of which 17 and 10 were significant at the 5% genome-wise level for the broiler-Leghorn cross and broiler-Fayoumi cross, respectively. Highly correlated growth traits showed similar QTL profiles within each cross but different QTL profiles between the 2 crosses. Most QTL for growth traits in the current study were detected in Gga 1, 2, 4, 7, and 14 for the broiler-Leghorn cross and Gga 1, 2, 4, 5, 8, and 13 for the broiler-Fayoumi cross. Potential candidate genes within the QTL region for growth traits at 1% chromosome-wise significance level were discussed. The results in the current study lay the foundations for fine mapping these traits in the advanced intercross lines and provide a start point for identification causative genes responsible for growth traits in chickens.

Long-term recombinant porcine somatotropin (PST) treatment mitigates the responses to subchronic lipopolysaccharide in swine.

The effect of multiple lipopolysaccharide (LPS) challenges in swine undergoing long-term treatment with porcine somatotropin (PST) was determined. Changes in aspartate serine transaminase (AST) occurred only at 24h following the first LPS challenge dose (P<0.05), while PST treatment moderated any change from occurring. Nonesterified free fatty acid (NEFA) levels were elevated in PST treated animals for the first 3 days following daily LPS treatment (P<0.05), while LPS treatment alone had no effect on plasma NEFA levels. Plasma urea nitrogen (PUN) levels were unchanged by LPS following the initial LPS challenge, but were decreased following the second challenge dose (P=0.014). These changes were long lasting, with a return to normal PUN levels not evident until Day 6. The PST treatment mitigated changes in PUN (P<0.05) when LPS was administered. Haptoglobin plasma levels, along with lipid peroxide production were not affected by LPS challenge or PST administration. LPS challenge reduced the levels of immunoreactive heat shock protein 70 (HSP70) throughout the entire challenge period (P<0.001). PST-LPS animals had normal levels of this protein. The results of the present study demonstrate that long-term PST treatment mitigates the adverse effects of subchronic LPS administration.

Dietary conjugated linoleic acid alters fatty acid composition of pig skeletal muscle and fat.

The dietary dose responsiveness of conjugated linoleic acid (CLA) addition relative to the fatty acid profile of edible lean tissue was examined in grower pigs treated with or without porcine somatotropin (pST). Gilts and barrows were fed CLA at 0, 0.25, 0.5, 1.0, or 2.0% of diet by weight from 20 to 55 kg BW. Additional pigs were administered (pST) at 0 or 100 microg x kg BW x d(-1) and fed either 0.5 or 2.0% CLA. Animals were fed diets containing 18% CP, 1.2% lysine, and 3.5 Mcal of DE/kg at 110% of ad libitum intake. The fatty acid profile in latissimus dorsi and dorsal s.c. adipose tissue samples was determined by gas chromatography. Dietary CLA replacement of corn oil increased the percentage of total fatty acids as stearic acid, whereas the percentages as oleic and linolenic acids were reduced in lattisimus muscle. Treatment with CLA + pST increased the percentages of linoleic and arachidonic acids while reducing the percentages of palmitic and oleic acids in lattisimus muscle. Dietary CLA increased the percentages of palmitic and stearic acids in s.c. adipose tissue while reducing the percentages of oleic, linoleic, linolenic, and arachidonic acids. The percentage of palmitic acid was reduced in s.c. adipose tissue, whereas linoleic acid was increased with CLA + pST. No synergistic effect was detected between CLA and pST for reducing carcass lipid content in grower pigs. However, pST increased the percentage of polyunsaturated fatty acids in lattisimus muscle and s.c. adipose tissue while reducing the percentages of saturated fatty acids in swine fed CLA.

Challenge differentially affects cytokine production and metabolic status of growing and finishing swine.

Growing (35 kg body weight) and finishing (85 kg body weight) swine challenged with endotoxin (Escherichia coli O55:B5) at a dose of either 2 or 20 microg/kg produced tumor necrosis factor (TNF)alpha in a dose-response relationship as measured by bioassay. Peak TNFalpha plasma levels were observed 1-2 hr post-challenge, returning to basal values 4 hr post-challenge. However, both an enzyme-linked immunosorbent assay specific for swine TNFalpha and total human TNFalpha demonstrated no dose-response relationship; peak plasma levels of immunoreactive TNFalpha were also observed 1-2 hr post-challenge. Maximal plasma interleukin-6 levels occurred 1-2 hr post-challenge and remained elevated through 8 hr post-challenge; there was no effect of lipopolysaccharide dose or metabolic status. Although the metabolic status of the animals also affected glucose levels, with growing animals exhibiting greater sensitivity compared with finishing animals, endotoxin-induced decreases in blood glucose levels were primarily dose-dependent. In contrast, changes in plasma urea nitrogen and free fatty acid (FFA) levels were strictly related to the metabolic status. Urea nitrogen levels were unchanged in growing swine, whereas they were increased in finishing swine and remained elevated 24 hr post-challenge. FFA levels in growing and finishing swine increased 3-6 hr post-challenge. FFA levels returned to basal values for finishing swine 24 hr post challenge, but in growing swine remained elevated 24 hr post-challenge. Plasma aspartate transaminase levels were increased through 24 hr post-challenge; animals given a dose of 20 microg/kg exhibited the greatest increase. Similarly, swine challenged with a dose of 20 microg/kg also exhibited the greatest increase in levels of conjugated bilirubin; there was no effect on unconjugated (free) bilirubin. These results demonstrate that endotoxin challenge of swine result in a pattern of changes that are dependent on both the dose of endotoxin used and the metabolic status of the animal examined.

Composition analysis of pork carcasses by dual-energy x-ray absorptiometry.

Dual-energy x-ray absorptiometry (DXA) was used as a noninvasive method to measure the composition of pig carcasses. A total of 181 half-carcasses (10 to 51 kg, from pigs slaughtered at approximately 30, 60, 90, and 120 kg) were scanned using a Lunar (Madison, WI) DPX-L densitometer. The DXA measurements of fat, lean, bone mineral, and total tissue mass were compared with chemical analysis for fat, water, protein, total ash, and scale weight. The mean value for total tissue mass by DXA was slightly less than the mean carcass weight (32.3 kg vs 33.6 kg, P > .05, R2 = .998). Although highly correlated (R2 = .81), the DXA measurement of the percentage of fat in the half-carcass was less (P < .001) than the chemical measurement (19.5 vs 24.9%). The DXA measurement of lean tissue mass (total mass less fat and bone mineral) was correlated with carcass protein (R2 = .97) and water (R2 = .99) content. The correlation (R2) between DXA bone mineral content and carcass ash content was only .68; however, DXA bone mineral content was more highly correlated with carcass weight (R2 = .93) than was carcass ash content (R2 = .70). When we used the DXA R value (ratio of the attenuation coefficients for fat and lean) to predict percentage of fat in the carcass, the mean value for predicted carcass fat was 25.9% (P > .05). Similarly, carcass protein and water content were predicted from DXA lean. Using DXA region of interest analysis, estimates of the fat content of the shoulder and ham regions were close to chemical values; however, DXA underestimated the fat content of the loin and side regions by 20 and 28%, respectively. When prediction equations were used to evaluate DXA measurements of the half-carcasses of 28 gilts and 37 boars slaughtered at approximately 120 kg, the half-carcasses of gilts contained more fat (33.9 vs 27.8%, P < .001), less protein (14.1 vs 16.1%, P < .001), and less water (45.9 vs 52.1%, P < .001) than those of boars. These results indicate that DXA could be a valuable research tool for measuring the composition of pig carcasses. On the basis of the results of this study, prediction equations were revised for the DXA estimation of fat, protein, and water content of the half-carcass: Fat (%) = 450 - (315 x DXA R value), Protein (g) = -145 + (.23 x DXA lean), and Water (g) = 150 + (.73 x DXA lean). Furthermore, it seems that separate prediction equations are needed for regional analysis.

Redefining body composition: nutrients hormones, and genes in meat production.

Growth rate and body composition of livestock can be optimized to meet consumer needs for a leaner product and to improve the efficiency of meat-animal production. Optimization strategies have traditionally focused on genetic selection and cost-effective ration formulation to achieve the genetic potential. Advances in understanding the mechanisms of growth and its control have led to additional opportunities for its manipulation. These include nutritional manipulation,the use of growth promotants, and, more recently, the ability to change the genetic potential through genetic engineering. Selection of appropriate candidate genes for manipulation depends on understanding the mechanisms underlying differentiation and growth of embryonic muscle cells. Recent advances in genetic engineering techniques, including gene therapy and germline transgenesis, will likely hasten the genetic progress toward a leaner carcass in domestic livestock. Such strategies may prove to be more beneficial then the controlled enhancement of somatotropin expression.

Effects of an endotoxin challenge on growth performance, carcass accretion rates, and serum hormone and metabolite concentrations in control pigs and those treated with recombinant porcine somatotropin.

Barrows were restrictively fed starting at 20 kg BW to determine the effects of endotoxin on growth performance of control and somatotropin-treated pigs. The following treatments were used: 1) daily i.m. vehicle injection until 55 kg BW; 2) daily i.m. injections of 100 micrograms of recombinant porcine somatotropin (pST)/kg BW, until 55 kg; 3) i.v. saline injections for 7 d consecutively starting at 60 kg BW; 4) i.v. injections of 1 microgram of bacterial lipopolysaccharide (LPS)/kg BW for 7 d starting at 60 kg BW; and 5) the combined LPS+pST treatment, with pST injections from 20 kg through the 7 d of LPS treatment. Pigs evaluated for LPS effects were fed to 60 kg anticipating a weight loss. Pigs were bled at 0800 and 1100 at 55 kg and on d 7 of LPS treatment. Rectal temperatures were taken on d 7. Treatment with pST increased ADG by 13 to 20% and improved feed:gain by 17 to 23% before LPS treatment. During the 7 d of LPS injections, ADG and feed:gain did not differ, although feed efficiency was impaired and variable. Rectal temperatures at 1100 were progressively increased: control < LPS < LPS-pST (P < .01). Protein accretion was improved 27% by pST treatment, and lipid accretion was decreased 45% before LPS. Lipid stores decreased (P < .01) after LPS treatment in the pST-treated pigs. Lipopolysaccharide treatment and(or) decreased feed intake reduced the hyperinsulinemia and hyperglycemia (P < .01) associated with pST treatment. These results indicate that LPS induced a simulated septicemia and that the effects were not negated by pST treatment. The observed hyperthermia was additive, possibly due to increased lean body mass induced by pST combined with the pyrogenic effect of LPS.

Effect of growth hormone or chromium picolinate on swine metabolism and inflammatory cytokine production after endotoxin challenge exposure.

OBJECTIVE: To determine whether recombinant porcine somatotropin (PST) or chromium picolinate (CrP) affected cytokine production and metabolism in swine after endotoxin challenge exposure. ANIMALS: 20 Poland China X Landrace pigs, 5/group. PROCEDURE: Pigs were given CrP-supplemented feed at body weight of 20 kg; PST treatment began at 60 kg, and both treatments continued through body weight of 90 kg. At 90 kg, pigs were challenge exposed with 20 micrograms of lipopolysaccharide (LPS)/kg of body weight. Blood samples were obtained at various times through 24 hours after LPS challenge exposure. RESULTS: In all pigs not given PST, glucose concentration decreased 2 to 4 hours after LPS. In PST-treated pigs, blood glucose concentration was decreased at 6 to 8 hours after LPS. Plasma insulin concentration paralleled changes in glucose concentration. Nonesterified fatty acid concentration was high 2 to 24 hours after LPS in pigs not given PST and at 6 to 24 h in PST-treated pigs. Plasma urea nitrogen concentration was high at 6 to 24 hours after LPS in pigs not given PST. The urea nitrogen values in PST-treated pigs were lower at all times. Serum aspartate transaminase activity was high 6 to 24 hours after LPS in pigs not given PST, whereas PST treatment prevented the increase in this enzyme activity. In untreated (PST) pigs, plasma bilirubin (total and direct) concentrations were high 4 to 8 hours after LPS and returned to normal at 24 hours. The PST- and CrP-treated pigs maintained normal plasma bilirubin concentrations. Interleukin 6 activity was unaffected by CrP and PST treatments. Treatment with CrP and PST decreased the tumor necrosis factor alpha response to LPS, compared with that in control pigs. CONCLUSIONS: PST, and to a lesser extent CrP, provide protection against the adverse metabolic effects of LPS-induced septic shock.

Beneficial effects of chromium on glucose and lipid variables in control and somatotropin-treated pigs are associated with increased tissue chromium and altered tissue copper, iron, and zinc.

Chromium (Cr) and somatotropin have been shown to increase lean body mass in pigs but by independent mechanisms. Somatotropin and Cr also affect blood glucose, lipids, and tissue trace metal concentrations. Twenty-four castrated male pigs were divided into four groups: 1) control basal diet; 2) basal diet + 300 micrograms of Cr/kg of diet as Cr picolinate; 3) basal diet + pituitary porcine somatotropin (ppST; 100 micrograms/kg live weight injected daily); and 4) basal diet + Cr + ppST. Pigs were fed the diets from 30 to 60 kg body weight and then killed. Supplemental Cr led to increased total Cr in kidney (1.1 vs 2.3 micrograms) and liver (5.9 vs 8.8 micrograms) but not in the heart independent of ppST treatment. Chromium concentrations in longissimus muscle were less than 1.5 ng/g in all samples, and any increases due to supplemental Cr were not detected. Somatotropin treatment led to decreased hepatic Cr, Cu, Fe, and Zn concentrations and increased total renal Cu, Fe, and Zn. These data demonstrate that supplemental Cr causes increased tissue Cr in the liver and kidney but not in the heart or muscle in control and somatotropin treated pigs. Somatotropin treatment caused decreased kidney and liver Cr concentrations that were offset by increased tissue weights. Somatotropin effects on tissue Cr, Cu, Zn, and Fe were variable and difficult to evaluate due in part to growth hormone-induced changes in organ weights.