N. C. Steele

Nutritional Modulation of Somatotropic Axis-cytokine Relationships in Cattle: A Brief Review

The objective of this review is to summarize data on the interrelationships that exist between nutrition, the endocrine system and their modulation of plasma tumor necrosis factor-alpha responses to endotoxin in cattle. During stress, intake of nutrients often is compromised and a percentage of available nutrients are diverted away from growth processes to stabilize other physiological processes of a higher survival priority. Management practices that minimize the magnitude and duration of disease stress will aid in speeding the return to homeostatic equilibrium. However, the shift away from growth during stress is almost inevitable as a mechanism to survive. Some degree of control and management of the metabolic cost of disease stress involves understanding the integration of nutritional, endocrine and immune signals by cells and working with the natural homeostatic processes. Endocrine hormones and immune system cytokine signals participate in redirecting nutrient use during disease stress. In an intricate interplay, hormones and cytokines regulate, modify and modulate each others production and tissue interactions to alter metabolic priorities. Levels of dietary protein and energy intake affect patterns of hormones and cytokines in the blood after endotoxin challenge and further modulate the biological actions of many of these regulatory effectors. In vivo, administration of growth hormone to young calves has significant effects to decrease the many specific physiological responses to endotoxemia. Many aspects of nutrition can attenuate or facilitate this effect.

Isolation and quantitation of metallothionein isoforms using reversed-phase high-performance liquid chromatography

Reversed-phase high-performance liquid chromatography (RP-HPLC) was used to isolate, characterize and quantitate liver and kidney metallothionein (MT) isoforms from a variety of animal species. The isoMTs were eluted from a radially compressed C18 column with a neutral sodium phosphate buffer and detected by UV absorbance (214 nm). Rabbit liver and horse kidney MTs were each found to be comprised of seven distinct isoforms. Pig liver and kidney MTs each exhibited three predominant isoforms, two of which were found to be subspecies of the MT-2 isoform. Rat liver MT was characterized by a less complex isoform pattern with only two major isoforms corresponding to the MT-1 and MT-2 isoMTs. Similarly, avian liver MT exhibited a distinct isoform pattern characterized by a low degree of complexity with the MT-2 isoform much more abundant than the MT-1 species. It was possible to apply the RP-HPLC separation to the resolution of individual MT isoforms from complex mixtures such as heat-treated cytosol. A standard curve was constructed using purified turkey hen liver MT-2 which demonstrated excellent linear correlation between integrated peak area and the quantity of MT injected onto the column. Recovery of MT from RP-HPLC was estimated to exceed 90%. Liver tissue from chicks injected on consecutive days with a dose of zinc was assayed for both MT-1 and MT-2 isoforms using the RP-HPLC analysis of cytosol samples. The MT-2 isoform was found to be preferentially expressed in response to zinc induction.

Effect of early feed restriction in male broiler chicks on plasma metabolic hormones during feed restriction and accelerated growth

1. Plasma GH was greater (P less than 0.05) on day 12 in ad libitum-fed birds compared to restricted chicks. Conversely, maximum GH levels were found to occur in the nutrient restricted chicks during the period of accelerated growth (day 42). 2. A significant decline in circulating insulin concentrations with advancing age was evident in both ad libitum-fed and restricted chicks. 3. Feed restriction significantly suppressed circulating T3 in restricted chicks, with concentrations returning to control levels upon refeeding. 4. A significant increase in T4 with advancing age was evident in both treatment groups, with T4 being significantly greater in controls compared to restricted chicks at 54 days of age.

Effects of growth hormone administration on vitamin D metabolism and vitamin D receptors in the pig

Twelve 36-kg pigs were given either 100 micrograms/kg porcine pituitary growth hormone (pGH) or placebo injections daily for 33 days. Serum was obtained weekly for analysis of minerals and vitamin D metabolites. On day 34, the pigs were sacrificed and renal and duodenal tissue were obtained for analysis of vitamin D receptor content (VDR). Animals treated with pGH grew faster and had a higher rate of bone accretion than did control animals. Serum concentrations of 1,25-dihydroxyvitamin D (1,25-(OH)2D) were significantly higher in pGH-treated pigs than in control pigs at all time points following initiation of treatment, with the greatest difference observed at day 28 (42.4 +/- 4.9 pg/ml in controls vs. 65.4 +/- 4.7 pg/ml in pGH-treated pigs). Serum 24,25-dihydroxy-vitamin D tended to be lower in pGH-treated pigs than in control pigs, being significantly lower on day 21 of the experiment (3.22 +/- .52 vs. 6.73 +/- 1.22 ng/ml, respectively). Serum concentrations of 25-hydroxyvitamin D and calcium were unaffected by pGH treatment. Kidneys of control pigs contained significantly more unoccupied vitamin D receptors than did kidneys from pGH-treated pigs (73.3 +/- 4.3 vs. 58.3 +/- 4.1 fmoles/mg protein). Duodenal tissue unoccupied vitamin D receptor content was similar in both pGH-treated (245 +/- 17.9 fmoles/mg protein) and control (263 +/- 21.8 fmoles/mg protein) pigs. Duodenal occupied vitamin D receptor concentration was similar in both pGH-treated (6.8 +/- .75 fmoles/mg protein) and control pigs (5.32 +/- .77 fmoles/mg protein).(ABSTRACT TRUNCATED AT 250 WORDS)

Characterization of the chicken muscle insulin receptor

Insulin receptors are present in chicken skeletal muscle. Crude membrane preparations demonstrated specific 125I-insulin binding. The nonspecific binding was high (36-55% of total binding) and slightly lower affinity receptors were found than are typically observed for crude membrane insulin binding in other chicken tissues. Affinity crosslinking of 125I-insulin to crude membranes revealed insulin receptor alpha-subunits of Mr 128K, intermediate between those of liver (134K) and brain (124K). When solubilized and partially purified on wheat germ agglutinin (WGA) affinity columns, chicken muscle insulin receptors exhibited typical high affinity binding, with approximately 10(-10) M unlabeled insulin producing 50% inhibition of the specific 125I-insulin binding. WGA purified chicken muscle insulin receptors also exhibited insulin-stimulated autophosphorylation of the beta-subunit, which appeared as phosphorylated bands of 92- and 81K. Both bands were immunoprecipitated by anti-receptor antiserum (B10). WGA purified membranes also demonstrated dose-dependent insulin-stimulated phosphorylation of the exogenous substrate poly(Glu,Tyr)4:1. However, unlike chicken liver, chicken muscle insulin receptor number and tyrosine kinase activity were unaltered by 48 hr of fasting or 48 hr of fasting and 24 hr of refeeding. Thus, despite the presence of insulin receptors in chicken muscle showing normal coupling to receptor tyrosine kinase activity, nutritional alterations modulate these parameters in a tissue-specific manner in chickens.

Regulation of Somatomedin Production, Release, and Mechanism of Action

Early studies revealed that a pituitary factor, growth hormone (GH), was in-volved in long bone growth and nitrogen retention and, without conclusive evidence, accepted theories suggested that GH interacted directly with target tissues to induce such adaptations in metabolism. The elegant studies by Salmon and Daughaday (1957) described a factor in normal serum that, when added to cartilage expiants, stimulated the incorporation of 35S04 into proteoglycans. Serum from hypophysectomized rats failed to stimulate sulfate incorporation; however, when such rats were treated with GH, serum stimulation of sulfate uptake was observed within 24 hr. In contrast, direct addition of GH to the expiant media, either in the -presence or absence of hypophysectomized rat serum, failed to stimulate sulfate incorporation. Based on the bioassay response, the serum component was named sulfation factor.

CHICKEN HEPATIC METABOLISM IN VITRO. PROTEIN AND ENERGY RELATIONS IN THE BROILER CHICKEN--VI. EFFECT OF DIETARY PROTEIN AND ENERGY RESTRICTIONS ON IN VITRO CARBOHYDRATE AND LIPID METABOLISM AND METABOLIC HORMONE PROFILES*

1. Ross male broiler chicks growing from 14 to 28 days of age were fed 14 and 20% protein diets (4 kcal day-1/body wt0.66) or 20 and 28% protein diets (2.8 kcal day-1/body wt0.66) in a 2 x 2 factorial arrangement to determine the effects of protein and energy intakes on in vitro lipogenesis (IVL) and net glucose production (NGP). Plasma concentrations of insulin, glucagon, thyroid hormones (T3 and T4) and somatomedin-C (Sm-C) were estimated by radioimmunoassay. 2. There was a significant (P less than 0.05) decrease in IVL in the chicks given the higher daily protein intake. 3. The higher protein intake increased (P less than 0.05) NGP while the lower energy intake decreased (P less than 0.05) NGP. 4. Insulin, both thyroid hormones and Sm-C were affected by dietary energy and protein intakes.

Effect of Recombinant Growth Hormone and Chromium Picolinate on Cytokine Production and Growth Performance in Swine

The effect of dietary chromium picolinate (CrP) and recombinant porcine growth hormone, somatotropin (rPST) administration on growth performance and cytokine production in Landrace-Poland China gilts was determined using a 2 by 2 treatment array. Treatments were: (1) control (basal diet), (2) CrP-supplemented diet (basal diet + 300 micrograms Cr3+/kg diet as CrP), (3) rPST (100 pg/kg body weight/day), and (4) rPST+CrP. CrP-supplemented diets were fed beginning at 20 kg body weight through 90 kg. Administration of rPST was begun at 60 kg weight and continued through 90 kg. All rPST treated pigs demonstrated improvements in growth performance versus controls. Pigs given CrP-supplemented diets showed no differences in growth performance. At 90 kg, pigs were challenged with endotoxin (lipopolysaccharide, 0.2 microgram/kg i.v.). Blood samples were collected at 0, 1, and 3 h postchallenge. Plasma IL-6 levels increased from 23 U/ml at time 0 to 1,927 U/ml at 3 h for control swine. Swine from the CrP treatment group had IL-6 levels of 8,130 U/ml at 3 h post-LPS. There were no differences in plasma IL-6 from pigs in the rPST and rPST+CrP treatment groups compared to the controls. Endotoxin challenge had no effect on either blood glucose levels or induction of TNF-alpha in any treatment group. PBMC from CrP-treated animals produced more IL-2 than peripheral blood mononuclear cells from all other groups.

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.

A Diabetic-Like Condition of Turkey Embryos Maintained in Shell-Less Culture

Abstract Serum insulin concentration and pancreatic insulin content were determined for turkey embryos incubated in ovo and in long-term shell-less culture (ex ovo). Insulin was undetectable (< 10 pg) in serum from 87% of the ex ovo embryos compared with their in ovo counterparts. This was evident at all incubation ages, although insulin was detectable in more of the ex ovo embryos on Day 24. Insulin increased in the embryos incubated in ovo from 122 (Day 15) to levels exceeding 2000 pg/ml at hatching. Total pancreatic insulin content was greater in the cultured embryos on Days 15, 17, and 22 compared with their in ovo counterparts. Serum glucose was significantly greater (P < 0.05) in the ex ovo embryos at all ages. In response to an infusion of l-arginine, serum insulin increased from 566 to 1256 pg/ml in the in ovo embryos, whereas no change was evident in the ex ovo embryos (233 vs 257 pg/ml). When embryos incubated in ovo were injected with insulin, a significant (P < 0.05) reduction of serum glucose was observed at 60 min after injection. Serum glucose concentrations remained elevated in the embryos incubated ex ovo despite the insulin injection. Liver glucose 6-phosphatase activity, assessed on Days 15 and 22 of incubation, was found to be significantly (P < 0.05) lower in the ex ovo embryos. Turkey embryos incubated in shell-less culture exhibited chronic hyperglycemia in concert with extremely low circulating levels of insulin. The pancreatic beta cells of these embryos were not responsive to arginine or elevated glucose. Taken together these findings suggest the occurrence of a diabeticlike condition in the ex ovo embryos. This defect in insulin secretion may, in part, be responsible for some of the developmental abnormalities characteristic of the turkey embryo cultured ex ovo.