Browsing by Author "El-Kadi, Samer Wassim"
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- Branched-Chain Amino Acid Metabolism in the Neonatal PigYonke, Joseph Allan (Virginia Tech, 2022-06-29)Branched-chain amino acids (BCAA) are a group of essential amino acids consisting of leucine, isoleucine, and valine. Leucine, in particular, has signaling functions affecting protein and energy metabolism. Plasma leucine concentration is positively correlated with obesity and associated metabolic disorders. We set out to test the hypothesis that metabolic dysfunction from high fat diets precedes dysfunctional BCAA metabolism. First, BCAA were supplemented to neonatal pigs for 4 weeks to evaluate whether the anabolic signaling function of leucine could increase muscle growth when fed for a longer duration than in previous studies. Neither normal pigs nor low birth weight pigs, which have naturally impaired muscle growth, grew better in response to BCAA supplementation, despite low birth weight pigs expressing less of the leucine sensing protein Sestrin2 in skeletal muscle. Furthermore, high plasma BCAA concentrations caused by the experimental diets had no effect on adiposity, liver fat accumulation, or expression of genes related to fatty acid synthesis, mitochondrial biogenesis, or energy expenditure in the pigs' livers. Having produced strong evidence that long term BCAA supplementation neither improves lean growth nor causes abnormal fat metabolism, we then tested whether fat supplementation changes BCAA metabolism. Pigs were fed milk replacer formula with either low energy (Control), or high energy from long-chain fatty acids (LCFA) or medium-chain fatty acids (MCFA) for 22 days. Although high fat diets did not increase plasma BCAA concentrations, the MCFA diet in particular caused metabolic changes which could lead to fatty liver disease and decreased oxidative BCAA disposal. Expression of fatty acid synthesizing genes were increased in the livers of pigs fed MCFA formula compared to Control and LCFA formula. Oxidation of α-ketoisocaproic acid was decreased in liver homogenate of pigs fed MCFA and LCFA formulas compared to Control. Additionally, hepatic oxidation of α-ketoisovalerate was decreased, and plasma concentration of α-ketoisovalerate was consequently increased, in pigs fed MCFA formula compared to Control, with LCFA formula causing intermediate results. In future research, it would be valuable to feed high MCFA formula for a longer period of time to determine whether nonalcoholic fatty liver disease will develop, and whether plasma BCAA concentrations will increase due to decreased oxidation. Overall, these studies concluded that long term BCAA supplementation does not increase muscle growth in neonatal pigs, but there is also no indication that they cause obesity or dysfunctional fat metabolism. On the other hand, high fat diets cause impairments in BCAA catabolism which may precede elevated plasma BCAA concentrations.
- Chronic Dietary Supplementation of Branched-Chain Amino Acids Does Not Attenuate Muscle Torque Loss in a Mouse Model of Duchenne Muscular DystrophySperringer, Justin Edward (Virginia Tech, 2019-09-12)Duchenne Muscular Dystrophy (DMD) is an X-linked recessive, progressive muscle-wasting disease characterized by mutations in the dystrophin gene. Duchenne muscular dystrophy is the most common and most severe form of inherited muscle diseases, with an incidence of 1 in 3,500 male births1,2. Mutations in the dystrophin gene result in non-functional dystrophin or the complete absence of the protein dystrophin, resulting in necrosis and fibrosis in the muscle, loss of ambulation, cardiomyopathies, inadequate or failure of respiratory function, and decreased lifespan. Although there has been little research for effective nutritional strategies, dietary intervention may be effective as an adjuvant treatment. In this study, wild type (WT) and mdx animals were provided either a control or elevated branched chain amino acid (BCAA) diet nocturnally for 25 weeks to determine if the elevated BCAAs would attenuate muscle torque loss. Twenty-five weeks of chronic, elevated BCAA supplementation had no impact on muscle function measures. Interestingly, mdx and WT animals had the same torque responses in the low stimulation frequencies (1 Hz – 30 Hz) compared to higher stimulation frequencies. Tetanus was reached at a much lower stimulation frequency in mdx animals compared to WT animals (100 Hz vs +150 Hz). The mdx mouse consistently had more cage activity in the light cycle X- and Y-planes. Interestingly, animals on the BCAA diet increased X-, Y-, and Z-plane activity in the dark cycles at four weeks while animals on the control diet more Z-plane activity at 25 weeks, although not significant. All three BCAAs were elevated in the plasma at 25 weeks, although only Leu was significantly elevated. The BCAAs had no effect on. The diaphragm and skeletal muscle masses were larger in mdx animals, and WT animals had a significantly larger epididymal fat pad. The active state of BCKDC determined by phosphorylation of the E1α enzyme was greater in WT animals in white skeletal muscle, but not red skeletal muscle. Protein synthesis effectors of the mTORC1 signaling pathway and autophagy markers were similar among groups. Wild type animals had increased mTORC1 effectors and animals on the BCAA diet had decreased autophagy markers, although not significant. Although BCAAs did not affect muscle function, fibrosis, or protein synthesis effectors, this study illustrates the functionality of mdx muscles over time. It would be interesting to see how the different muscle fiber types are affected by DMD, noting the differences between the diaphragm, heart, red muscle, and white muscle fibrosis markers. Although there was no increase in mTORC1 effectors with an elevated BCAA diet, it would be interesting to determine muscle protein synthesis, myofibrillar protein synthesis, and total protein turnover in the mdx mouse with an elevated BCAA diet, although the dietary intervention started when mice arrived at 4 weeks of age, earlier intervention may be beneficial early in the disease process.
- Consumption of Endophyte Infected Fescue During Gestation in Beef CowsOliver, Katherine Rene (Virginia Tech, 2016-07-11)Tall fescue is a widely grown, cool season grass prevalent in the eastern United States that is known for its resistance to abiotic and biotic stresses. A main reason for tall fescue's resistance to these stresses is attributed to the presence of a fungal endophyte. Unfortunately, this endophyte also adversely affects cattle production. Cows consuming the ergot alkaloids produced by these endophytes can exhibit decreased feed intake, growth performance, organ vasoconstriction, and increased rectal temperature. This work is interested in examining how endophyte toxin exposure impacts pregnancy in cattle. Reduced blood flow to the fetus and inadequate maternal nutrition contributes to intra uterine growth restriction (IUGR), and this work proposed that fescue endophyte toxicity affects the gestating cow and fetus. Three studies were completed. In experiment 1, gestating cows grazed high or low endophyte fescue pastures during late gestation to determine if exposure to ergot alkaloids in utero results in IUGR and if calves from these pregnancies have altered growth performance. Creep feeding was evaluated as a mitigation strategy for impaired calf growth due to fescue toxicity, and feedlot performance was evaluated to determine if consuming fescue during gestation and creep feeding would affect feedlot performance. Calf BW was different (P < 0.01) by treatment x time. Birth weights of calves were similar , prior to creep feeding calves exposed to high endophyte fescue were lower, and post-supplementation creep fed calves had increased BW. Days on feed and dressing percentage were decreased in the supplemented group, and marbling score was decreased for both the supplemented and unsupplemented groups following the completion of the feedlot phase (P < 0.05). The second study was setup similar to study one, however cows were exposed to fescue pastures from d 170 of gestation until calving. Calf birth weights did not differ, but weights were increased in the supplemented group post creep feeding (P < 0.05). Average daily gains (ADG) of supplemented calves were greater during the supplementation period (P < 0.01). In the third study, indwelling vaginal temperature probes were used to evaluate differences in body temperature of cows fed fescue seed with high or low levels of ergot alkaloids during early gestation, and in varying environmental conditions. In the winter trial, body temperature was measured hourly from days 0-14 of gestation. In the summer trial, body temperature was measured hourly from days 0-32 of gestation. Body temperatures were different (P < 0.01) between treatments during both trials.
- Determining the Underlying Factors of Fresh Ham Color VariationElgin, Jennifer May (Virginia Tech, 2019-07-10)Consumers associate meat color with quality. In some cases, especially in fresh and cured hams, the surface of a ham, whole, boneless or sectioned and formed displays a color gradient, which is unsightly and generally is considered of lower quality and must be discounted or processed different where color is less critical to the ultimate value of the resulting product. This disparity in color uniformity across fresh and cured products is sometimes known as two-toning and is most often found in the semimembranosus (SM) and associated muscles of fresh hams and is exacerbated with curing. The underlying color of fresh meat may be a function of postmortem metabolism or the underlying characteristics of those muscles involved. Therefore, the objective of this study is to determine the changes in underlying muscle type and postmortem metabolism in those muscles responsible for fresh ham color variation. Semimembranosus (SM) muscles of 15 mixed bred pigs were collected at 30 min and 1440 min postmortem, and muscle color was determined and muscles were collected and snap frozen for various energy metabolism analyses. Differences in color (L*, a* and b*) were noted across the face of the muscle by zone and time (P < 0.0001) but no differences were detected in pH and lactate, glucose, glucose-6-phosphate, and glycogen metabolisms. Glycolytic potential was also measured on a lactate basis and showed no differences across zone (P = 0.0746) but increased over time (P < 0.006). Lactate and pH were plotted and showed a linear relationship linear relationship (R2 = 0.928337) at 30 min (P < 0.0001) and at 1440 min (R2 = 0.161412; P < 0.0015). Muscle type characteristics showed no difference between zones and time. Buffering capacity showed a significant difference at pH 6 (P < 0.0359) and with time across all pH measured (P < 0.0001). These data suggest inherent differences, such as location and function, in the semimembranosus muscle may be more critical in developing fresh color than aberrations in postmortem metabolism.
- Dietary supplementation of saccharin-based artificial sweeteners and capsicum oleoresin as a strategy to mitigate the negative consequences of heat stress on pig growth performance and intestinal physiologyBiggs, Morgan Eleanore (Virginia Tech, 2017-06-28)Pigs exposed to elevated ambient temperatures exhibit reduced average daily gain, alterations in muscle and fat deposition during growth phases, and decreased overall health. Negative aspects of gastrointestinal (GI) function, integrity and permeability also result from hyperthermia. Saccharin-based artificial sweeteners have the potential to ameliorate the negative effects of heat stress (HS) by increasing GI GLP-2 production while capsicum oleoresin has been shown to reduce inflammatory response. Study objectives were to examine the effects of two artificial sweeteners, neohesperidin dihydrochalcone and saccharin, in combination with capsicum oleoresin (TakTik Sweet Heat , Pancosma, SA) on growth performance of pigs. Forty-eight pigs (12 weeks of age, 47.3 ± 7.6kg) were assigned to six treatments: thermal neutral conditions (21°C) fed ad libitum with (TN+) or without supplement (TN-), heat stress (35°C) fed ad libitum with (HS+) or without supplement (HS-), and thermal neutral conditions pair-fed to HS intake with (PFTN+) or without supplement (PFTN-). Dietary supplementation began 2 days prior to the 3-day environmental treatment period. Body weight was recorded on day -1 and day 3 relative to start of environmental treatment. Body temperature (BT) and respiration rate (RR) were measured thrice daily and feed intake (FI) were recorded daily. Blood samples were collected on day -1 and day 3 to determine metabolite profiles and immune response. Following sacrifice, sections of duodenum, ileum, and colon were collected for histology. Pigs in HS conditions had increased average BT and RR (~2.7-fold) compared to TN and PFTN groups (P<0.01). Irrespective of day, HS+ animals had increased respiration rates when compared to HS- animals (P<0.04). Heat stress decreased FI compared to TN groups. Regardless of treatment, supplement increased feed efficiency by 0.12kg (P<0.04). Circulating glucose concentration tended to decrease in HS+ and PFTN+ pigs compared to non-supplemented treatments (P=0.1). Circulating insulin increased in HS compared to PFTN (P<0.04), but did not differ from TN. Lymphocyte count tended to be decrease in HS compared to PFTN and TN (P=0.052). Monocyte count increased during HS compared to PFTN (P=0.01), but HS and TN did not differ. Supplement increased basophil count (P<0.03), irrespective of treatment. Ileal villus height tended to decrease during HS and PFTN compared to TN (P<0.08). Duodenal and colon villus height nor duodenal, ileal and colon crypt cell depth did not differ between groups. Overall, TakTik Sweet Heat supplementation increases pig feed efficiency and may improve immune response.
- Diminished satellite cell fusion and S6K1 expression in myotubes derived from skeletal muscle of low birth weight neonatal pigsChen, Ying; Zhu, Haibo; McCauley, Sydney R.; Zhao, Lidan; Johnson, Sally E.; Rhoads, Robert P.; El-Kadi, Samer Wassim (The American Physiological Society, 2017-02-09)Low birth weight (LBWT) is consistently associated with impaired postnatal muscle growth in mammals. Satellite cell (SC)‐mediated myonuclear incorporation precedes protein accumulation in the early stages of postnatal muscle development and growth. The objective of this study was to investigate proliferation and differentiation of SCs and the regulation of protein synthesis signaling in response to insulin‐like growth factor (IGF)‐I stimulation in SC‐derived myotubes of LBWT neonatal pigs. SCs isolated from Longissimus dorsi muscle of LBWT and NBWT pigs (3‐d‐old, n = 8) were cultured and induced to proliferate and differentiate to myotubes in vitro. On day 3 of differentiation, myotubes were fasted in serum‐free media for 3 h and treated with human recombinant R³‐insulin‐like growth factor‐I (rh IGF‐I) at 0, 25, and 50 ng × mL⁻¹ for 30 min. There was no difference in proliferation rates of SCs from LBWT and NBWT pigs. However, LBWT SC fusion was 15% lower (P ≤ 0.05) without a difference in MyoD or myogenin mRNA expression in comparison with NBWT pigs, suggesting SCs are not intrinsically different between the two groups. IGF‐Ι stimulation at physiological concentrations activated downstream effectors of mTOR similarly in myotubes from LBWT and NBWT pigs. However, abundance of ribosomal protein S6 kinase 1(S6K1) was lower in myotubes of LBWT compared to their NBWT siblings (P ≤ 0.05). These results indicate that the modest reduction in SC fusion and S6K1 expression are not the major contributors to the impaired postnatal muscle growth of LBWT pigs.
- Downregulated Translation Initiation Signaling Predisposes Low-Birth-Weight Neonatal Pigs to Slower Rates of Muscle Protein SynthesisChen, Y.; McCauley, Sydney R.; Johnson, Sally E.; Rhoads, Robert P.; El-Kadi, Samer Wassim (Frontiers, 2017-07-11)Low-birth-weight (LBWT) neonates experience restricted muscle growth in their perinatal life. Our aim was to investigate the mechanisms that contribute to slower skeletal muscle growth of LBWT neonatal pigs. Twenty-four 1-day old male LBWT (816 ± 55 g) and normal-birth-weight (NBWT; 1,642 ± 55 g) littermates (n = 12) were euthanized to collect blood and longissimus dorsi (LD) muscle subsamples. Plasma glucose, insulin, and insulin-like growth factor-I (IGF-I) were lower in LBWT compared with NBWT pigs. Muscle IGF-I mRNA expression were lower in LBWT than NBWT pigs. However, IGF-I receptor mRNA and protein abundance was greater in LD of LBWT pigs. Abundance of myostatin and its receptors, and abundance and phosphorylation of smad3 were lower in LBWT LD by comparison with NBWT LD. Abundance of eukaryotic initiation factor (eIF) 4E binding protein 1 and mitogen-activated protein kinase-interacting kinases was lower in muscle of LBWT pigs compared with NBWT siblings, while eIF4E abundance and phosphorylation did not differ between the two groups. Furthermore, phosphorylation of ribosomal protein S6 kinase 1 (S6K1) was less in LBWT muscle, possibly due to lower eIF3e abundance. In addition, abundance and phosphorylation of eIF4G was reduced in LBWT pigs by comparison with NBWT littermates, suggesting translation initiation complex formation is compromised in muscle of LBWT pigs. In conclusion, diminished S6K1 activation and translation initiation signaling are likely the major contributors to impaired muscle growth in LBWT neonatal pigs. The upregulated IGF-I R expression and downregulated myostatin signaling seem to be compensatory responses for the reduction in protein synthesis signaling.
- Effect of amino acids and vitamin D3 on performance and biological responses in poultryWen, Jinlei (Virginia Tech, 2018-06-08)As productive performance is improved by breed selection, amino acid requirements may change to support this higher performance in poultry. The first objective of this dissertation was to update the valine and tryptophan requirement of small-framed laying hens and the lysine requirement of young broilers using empirical dose-response methods. The tryptophan requirement was estimated as 155.8 mg/d for egg mass, 153.2 mg/d for egg production and 140.4 mg/d for feed conversion ratio using a linear broken line model. For valine, the requirement was highest for egg mass, 597.3 mg/d, followed by egg production, 591.9 mg/d and feed conversion ratio (FCR), 500.5 mg/d. The lysine requirement of young chicks was estimated by conducting four short term experiments from 1 to 3, 3 to 5, 5 to 8 and 8 to 11 days of age, respectively. The lysine requirement from 1 to 3, 3 to 5 and 5 to 8 days of age were not able to be estimated as no dose response was observed on growth performance most likely due to an overestimation of the lysine requirement. Digestible lysine requirement from 8 to 11 days of age was 1.057%, 1.050% and 1.016% based on body weight gain, FCR and pectoralis major weight using a linear broken line model, respectively. In addition to determining amino acid requirements, research was conducted to develop a new bacterial protein meal for use in laying hens diets. The data suggested that diets containing 7.5% of the bacterial protein meal was able to at least maintain egg production in laying hens, but 15% bacterial protein meal resulted in reduced performance. The second objective of this dissertation was to investigate the effects of various concentrations of dietary vitamin D3 on pullet and laying hen performance, eggshell quality and bone health in laying hens. Pullets/hens were randomly assigned to five dietary treatments containing vitamin D3 from 1,681 to 68,348 IU/kg diet from day of hatch until 68 weeks of age. These data suggested that dietary vitamin D3 fed at 68,348 IU/kg resulted in reduced egg production, but vitamin D from 8,348 to 35,014 IU/kg diet maintained egg production, increased egg vitamin D content in a dose dependent manner, and generally increased both eggshell quality and pullet and hen bone mineral status.
- The Effects of Dietary Fructose and Fat on the Reproductive Parameters of Prepubertal and Pregnant GiltsMcCracken, Victoria Lorraine (Virginia Tech, 2015-04-21)Body adiposity is generally considered the most pertinent factor in puberty attainment; however, recent data suggests that pre-pubertal reproductive tract development may be altered by dietary sugar consumption. Two experiments were conducted to delineate the direct effects of fructose on the maturation of the pre-pubertal reproductive tract and fertility. At three weeks of age, forty gilts were placed on one of five dietary treatments (n=8) containing 15% fat (FAT), 35% fructose (FRU), both fat and fructose (HFHF), or two different controls: one standard industry (IND) diet meant to result in optimal lean growth and a second diet to account for the reduced lysine (LYS) intake in the treatment diets. Body weights did not differ amongst any of the five treatments on the day of sacrifice (P=0.32). As a percentage of BW, total reproductive tracts were heavier in fructose fed gilts (1.3±0.1 v. 0.8±0.1%; P=0.01) compared to non-fructose gilts. In the second experiment, starting at 130d of age, gilts were checked twice daily for puberty attainment. Gilts that attained puberty were artificially inseminated (AI) on their third estrous cycle. On gestational day 38±3, pregnant gilts were harvested for reproductive tract collection. Fewer fructose fed (FRU and HFHF) pigs became pregnant than non-fructose fed (IND, LYS, and FAT) gilts (25% v. 75% respectively; P=0.03). All HFHF gilts failed to become pregnant. Placental weights were greater in LYS fetuses than FAT fetuses (79.07 ± 6.55g v. 47.26 ± 6.45g, respectively, P= 0.04). Taken together, these results demonstrate that fructose consumption increases reproductive tract size, but that reproductive capabilities are reduced.
- Effects of early weaning on finishing feed efficiency, marbling development and retail product quality of beef steersSmith, Jason Kaleb (Virginia Tech, 2015-09-03)Beef cattle producers continuously search for nutritional management options that provide flexibility to production scenarios. Due to its positive effects on maternal productivity, early weaning is one such alternative strategy that has received considerable interest. To better understand the effects of early weaning on calf productivity, an extensive literature review and three experiments were conducted to evaluate the effects of early weaning on finishing feed efficiency, marbling development and retail product quality of beef steers. In experiment I, which included 90 Angus-sired steers from four calving seasons, early weaning followed by a short ad libitum concentrate-feeding and pasture-backgrounding phase reduced finishing residual feed intake (RFI) by 7 % (P < 0.0001) and increased carcass marbling score (MS) by 10 % (P < 0.01) when compared to conventionally weaned (CW) contemporaries. Similar effects were observed in experiment II, which included 28 Angus and Simmental-sired steers, as early weaning reduced RFI (P < 0.01) and increased carcass MS (P < 0.01). Lung mass of early weaned (EW) steers was greater than their CW contemporaries (P < 0.05), and was inversely related to RFI (R2 = 0.17; P < 0.05). Finishing treatments in this experiment included a high corn ration and an alternative low corn ration that iso-calorically replaced 50 % of the DM from corn with dried corn gluten feed. Iso-caloric replacement of corn reduced lung mass (P < 0.01), and when combined with the observed increase of EW steers suggests that lung development is affected by dietary energy type at various stages of growth. In experiment III, objective analyses of ribeye steaks obtained from steers included in experiment two revealed that early weaning increased cross-sectional muscle fiber area by 28 % (P < 0.001) and tended to increase (P = 0.08) Warner-Bratzler shear force by 36 %. Nonetheless, these effects were not great enough to alter un-trained consumer perception of texture (P ≥ 0.65), juiciness (P ≥ 0.55), flavor (P ≥ 0.25) or overall acceptability (P ≥ 0.34). Collectively, these results indicate that early weaning enhances finishing feed efficiency and carcass marbling without affecting un-trained consumer sensory perception.
- Effects of Phosphorus Supplementation on Grazing Beef CattleNeil, Scott Joseph (Virginia Tech, 2015-11-04)Phosphorus (P) losses due to overfeeding of the mineral to livestock can contribute to surface water degradation. The objective of this study was to examine the impact to supplementing various levels of mineral P to grazing beef cattle. A producer survey and a research trial were conducted to examine the effects of supplementing mineral phosphorus (P) to grazing cattle. In the first study, mineral tags, producer surveys, and fecal, forage, and soil samples were collected from beef cattle operations in Virginia's Chesapeake Bay watershed. Samples (n=166) were collected from 120 producers in 11 counties. Soil test P results were based on Virginia Cooperative Extension soil test guidelines characterized as low (12 %), medium (37 %), high (35 %), and very high (16 %). Pasture grab samples contained 0.34 ± 0.12% P and forage P concentration increased (P < 0.01) across soil P categories going from low to very high. Fecal total phosphorus (TP) was lowly correlated (R2 = 0.18, P < 0.01) to forage P concentration. Mineral supplements were categorized as nil (<1.0% P), low (1.0-<3.0% P), medium (3.0-<6.0% P), and high (>6.0% P). Fecal TP and inorganic phosphorus (Pi) concentration increased (P < 0.01) with mineral P levels. Fecal TP and Pi were lower (P < 0.01) when nil and low P mineral were supplemented as compared to medium and high P mineral. Soluble P (defined as fecal Pi/fecal TP*100) also increased (P < 0.01) with increasing mineral P content going from nil to high. All farms surveyed required little or no P supplementation in regard to beef cattle P requirements. The majority (82%) of producers were receptive to modifying mineral P supplementation practices based on forage P levels. A 56-d study was also conducted with eight yearling Hereford steers (261±30 kg) grazing cool-season grass fall re-growth to determine the effects of varying levels of P supplementation on fecal P excretion. Treatments consisted of dicalcium phosphate supplemented at 0 (D1), 10.0 (D2), 20.0 (D3), or 30.0 (D4) g/d in a randomized 4x4 replicated Latin square design. These treatments provided an additional 0, 1.9, 3.7, and 5.6 g/d of P respectively. Two esophageally cannulated steers were used to collect forage samples for nutrient analysis. Forage P content was analyzed from hand collected samples. Forage P concentrations averaged 0.49% of dry matter (DM) across all periods. Chromic oxide (Cr2O3) was administered twice daily via gelatin capsule at 0630 and 1830 to serve as an external marker for determination of fecal dry matter excretion (DME). Indigestible NDF (iNDF) was used as an internal marker to determine dry matter intake (DMI). Due to the high forage P content, average P intake was in excess of National Research Council (NRC) requirements for all diets (D1 = 281%; D2 = 297%; D3 = 323%; D4 =348%). Orthogonal contrasts were performed to assess the relationship between treatment and P excretion. A linear response (P < 0.01) in daily inorganic P (Pi) excretion (0.054, 0.052, 0.062 and 0.063 g/kg of BW ± 0.003 for D1, D2, D3 and D4, respectively) was observed across treatments. Daily total P (TP) excretion increased linearly (P < 0.01) across treatments (0.080, 0.079, 0.092 and 0.093 g/kg of BW ± 0.003 for D1, D2, D3 and D4, respectively. When forage P content is sufficient to meet the requirement of grazing cattle, increasing P supplementation results in greater P excretion without additional benefits to growth or nutrient digestibility.
- Evaluation of amino acid transport and protein metabolism in the mammary gland of dairy cattleYoder, Peter Samuel (Virginia Tech, 2019-05-28)Improving our understanding of milk protein production regulation and AA transport is important for successfully formulating diets for AA and improving N efficiency. The objectives were to study protein synthesis regulation and AA transport using in vitro and in vivo models. In the first experiment, the objective was to evaluate the ability of five distinct AA profiles and balancing Lys to Met ratio to 3:1 to stimulate protein translation. No single AA profile uniquely stimulated phosphorylation of translational machinery related proteins suggesting identification of a single optimal AA profile as unlikely. In the second experiment, an in vitro method using three different AA isotopes was developed to trace AA movement. The method assesses bi-directional transport of multiple AA simultaneously enabling evaluation of unidirectional uptake kinetics. This method was used to evaluate AA concentrations representing 16, 100, 186, and 271% of cow plasma AA concentrations. Amino acid uptake was not saturable within the in vivo range for eleven AA. Arginine, Val, and Pro exhibited saturation with the Michaelis-Menten km being 95, 49, and 65% of in vivo concentrations. Results suggest that AA transport is generally non-saturable and that high bi-directional transport exists which enables a mechanism for mitigating AA shortages. In experiment 3, the objective was to evaluate milk protein production and regulation from infusing Met, Lys, and His (MKH) or Ile and Leu (IL). The two EAA groups independently and additively increased milk protein yield. This finding contradicts the single limiting AA theory that a single nutrient will limit milk protein yield. Changes in udder AA extraction and blood flow from supplemental EAA reveal flexible delivery mechanisms. The phosphorylation state of proteins associated with the mTOR pathway was impacted by both EAA treatments. Changes in the udder proteome suggest negative feedback on mTOR pathway activation when milk protein yield was increased by the EAA groups separately but when supplemented together, negative feedback was lessened. Results indicate that multiple EAA can stimulate milk protein production, the ability of AA transport to match intracellular needs, and that the single limiting AA theory or existence of a unique optimal AA profile is likely irrelevant in dairy cows.
- Examining the Influence of Muscle Fiber Type on Protein Turnover Signaling in Growing PigsSeymour, Kacie Tinnesz (Virginia Tech, 2020-05-28)Postnatal skeletal muscle growth occurs through myonuclear accretion and high protein turnover rate. While fiber type composition of the muscle could affect protein turnover rate, less is known about how fiber type influences the regulation of protein synthesis and degradation signaling pathways. Thus, the hypothesis of this work was that variation in fiber type composition will differentially affect the regulation of signaling pathways related to protein turnover in skeletal muscle hypertrophy in growing pigs. Downregulated protein synthesis signaling and reduced expression of type II MyHC isoforms have been reported in skeletal muscles of low birth weight (LBWT) neonatal pigs. Therefore, we sought to determine whether these changes are sustained until weaning and would explain the reduction in LBWT pig growth compared to their normal birth weight (NBWT) sibling at weaning. Another objective was to determine whether the regulation of protein turnover signaling pathways are correlated to fiber type differences in skeletal muscles. Our data suggest that the longissimus dorsi (LD, glycolytic) muscle of LBWT pigs experienced compensatory growth while the soleus (oxidative) remained proportionally smaller. Growth of the LD was accompanied by upregulation of translation initiation. Additionally, there was no difference in expression of MyHC isoforms between NBWT and LBWT pigs. These data suggest the rapid growth of the LD of LBWT pigs may be attributed to an upregulation of protein synthesis signaling and occurred only in glycolytic muscles. A caveat in LBWT pig model is that the reduction in type II MyHC at birth is not the only factor that could influence muscle growth, and that other factors may have confounded our results. This is why we aimed to use β-adrenergic agonist as a means to induce a shift fiber type in muscles to a more glycolytic phenotype. Our objective was to determine the influence of the β-adrenergic agonist Ractopamine (RAC) induced slow-to-fast fiber type transformation on the regulation of protein synthesis and degradation pathways. Although supplementation improved translational capacity, enhanced S6K1 phosphorylation, and reduced the abundance of calcium-dependent proteases, RAC feeding had no effect on body or muscle weights. These results suggest that a fiber type transformation without other physiological influences does not alter protein turnover signaling in favor of hypertrophy in growing pigs.
- Glucose Metabolism in Low Birth Weight Neonatal PigsMcCauley, Sydney Russelle (Virginia Tech, 2019-02-04)The neonatal period in mammals is characterized by high growth rates and is dominated by skeletal muscle hypertrophy. Low birth weight (LBWT) neonates experience restricted growth and development of skeletal muscle, leading to metabolic perturbations later in life. The overall hypothesis of this dissertation was that in utero disturbances in glucose metabolism and increased energy requirements predisposes LBWT neonatal pigs to metabolic disturbances after birth. We sought to increase growth of skeletal muscle and improve glucose production through increasing dietary energy and to determine the changes in glucose catabolism and metabolic flexibility in different skeletal muscle fiber types in LBWT neonates. Piglets were considered normal birth weight (NBWT) and LBWT when birth weight was within 0.5 SD and below 2 SD of the litter average, respectively. Increasing dietary energy increased lean deposition in the longissimus dorsi (LD) in both NBWT and LBWT neonates. Although glucose rate of appearance was greater in LBWT compared to their NBWT sibling, glucose concentrations were reduced in LBWT compared to NBWT pigs, regardless of diet fed. Postprandial glucose concentrations were lower in LBWT compared to NBWT pigs, regardless of diet fed, although rate of appearance did not differ between them. This would suggest that glucose is being absorbed in the peripheral tissues to be utilized. However, expression of enzymes related to glycolysis were downregulated in both the soleus and LD of LBWT compared to NBWT neonatal pigs. In addition, expression of enzymes related to the catabolism of glucose in the serine biosynthetic pathway were decreased in both the soleus and LD muscles of LBWT compared to NBWT neonatal pigs. Expression of the pentose phosphate pathway was slightly increased in LBWT compared to NBWT siblings in both muscle types. Increased expression of pyruvate dehydrogenase 4 was exhibited in both the soleus and LD of LBWT pigs compared to NBWT siblings. This would indicate a switch in fuel utilization to more fatty acid oxidation. By contrast, CO2 production from the oxidation of palmitate was reduced in LBWT compared with NBWT pigs along with reduced oxidation of glucose and pyruvate. In conclusion, lipid supplementation increased growth at the expense of fat deposition in the liver of NBWT and LBWT pigs. However, supplementing with fat did not increase glucose production due to the contribution of glycerol remaining constant. Hypoglycemia cannot be attributed to greater catabolism in skeletal muscle due to decreased expression of glycolytic genes and the addition of fatty acids did not spare glucose oxidation in skeletal muscle of LBWT pigs.
- Harvest practices cause aberrations in pork qualityDaniels, Rachel Page (Virginia Tech, 2019-08-16)Consumers are often confused and reluctant to purchase products from fresh retail meat counters that exhibit significant color variations. This hesitation to purchase results in annual profit loss to the pork industry. Processes to minimize color variation remain a major focus of the industry and many meat science programs across the globe. Previously, we found inherent muscle characteristics contribute to variations in pork quality but these characteristics fail to explain the high frequency of two-toning and other pork quality defects routinely occurring in many hog processing facilities. Therefore, we hypothesized harvesting practices, such as scald alter color across muscles of the ham. Scald time was initially investigated using 32 carcasses subjected to either a 4 (n=16) or 8 (n=16) min scald time. Samples were collected before or after scalding and at 24 hrs. A 50% reduction in scald time resulted in (p < 0.0001) lighter muscle color (L*) early postmortem, although the 8 min scald treatment was lighter (p < 0.005) at 24 hrs. Although differences in pH (p < 0.0001) and color were noted, ultimate carcass temperature was not affected. To that end, we moved to validate our hypothesis in an industrial plant setting. Carcasses (n=200) were assigned treatments of 6.5 or 7.5 min scald times, and SM muscle samples were collected at 24 hrs. Surprisingly, the shorter scald time resulted in (p < 0.05) a lighter color, contradicting our first study. To explore this color issue further, we uncoupled scald from the dehairing process. To achieve this goal, carcasses (n=24) were assigned to either an 8 or 16 min time to dehair, with or without scalding. Protracted time to dehair resulted in higher ultimate pH (p < 0.005) and less color variation across the muscle (p < 0.05). Though a color gradient remained, the variation across the muscle was reduced by increasing time to dehair. These data show time to dehair affects pork quality development and suggest that delaying time to physical manipulation of the carcass may improve pork color, thus increasing consumer acceptance.
- In ovo and feed application of probiotics or synbiotics and response of broiler chicks to post-hatch necrotic enteritisWhite, Mallory Beth (Virginia Tech, 2021-06-04)Immediately post-hatch, broiler chicks are exposed to microbes that begin colonizing the gut, including environmental pathogens. One of the costliest enteric diseases in broiler production is necrotic enteritis (NE), caused by the ubiquitous opportunistic bacteria Clostridium perfringens (CP). With the worldwide reduction in antibiotic growth-promoters (AGPs), there is increased interest in natural alternatives to reduce disease and improve broiler health. The overall objective of the studies described herein was to apply probiotics or synbiotics to birds by in ovo application or orally before they leave the hatchery, then evaluate bird performance and various intestinal responses. Data were analyzed in JMP with LS Means to separate means with significance assigned at P ≤ 0.05 and trends at 0.05 < P ≤ 0.10. The first 21-day (D) study used 480 male Cobb 500 broilers randomly divided into one of four treatments using a 2x2 factorial design: a no-additive control (CTRL), a one-time oral application of synbiotic at the hatchery fed a basal diet (HS), an oral application of water at the hatchery with dietary synbiotics (DS), and a hatchery synbiotic plus dietary synbiotic (HSDS). Performance was measured on day-of-hatch (DOH), D3, D7, D14, and D21. mRNA abundance of various intestinal markers was measured at D7 and D21, including tight junction proteins ZO-1, ZO-2, and CLD-1; nutrient transporters SGLT1 and PepT1; and immune response markers TLR2, TLR4, and IL-10. HS lowered feed intake (FI) and feed conversion ratio (FCR) without lowering body weight (BW) from D14-21. There was greater abundance of PepT1 mRNA (P ≤ 0.1) and IL-10 mRNA (P ≤ 0.05) on D21 in HSDS. Second, a 21-day pilot study with 480 male and female Cobb 500 broilers was conducted to determine the optimum in ovo dosage level of a probiotic or synbiotic (PROB or SYNB) applied at embryonic day 18 (E18) with subsequent NE challenge using seven treatments: in ovo application of sterile water (CTRL), low (PROB-L or SYNB-L: 1x105 CFU), medium (PROB-M or SYNB-M: 1x106 CFU), or high (PROB-H or SYNB-H: 1x107 CFU) probiotic or synbiotic doses dissolved in sterile water. Performance measurements were taken on DOH, D4, D8, D14 and D21. On D8, NE lesion scores were not impacted by treatment. D8 ileal samples were taken for mRNA abundance of TLR4, IL-10, IL-1β, AvBD8, AvBD10, and AvBD13. SYNB-H had higher abundance of AvBD10 mRNA compared to CTRL (P ≤ 0.1), and higher IL-1β mRNA compared to SYNB-L (P ≤ 0.05). PROB-H and SYNB-H had better performance than the low and medium doses, but were not better than the CTRL. The high doses were chosen for subsequent studies. Third, a longer 42-day study using 1,630 Ross 708 male and female broilers was conducted consisting of the following six treatments. A negative control (NC): sterile water in ovo fed basal corn/soybean meal mash diet without NE challenge; antibiotic growth-promoter (AGP+): sterile water in ovo fed basal diet with virginiamycin (0.5 kg/MT) as an AGP with NE challenge; NC+: same as NC plus NE challenge; SI+: synbiotic in ovo fed the basal diet and NE challenged; SD+: sterile water in ovo fed basal diet supplemented with synbiotic (0.5 kg/MT feed) and NE challenged; and SID+: synbiotic in ovo fed basal diet with synbiotic (0.5 kg/MT feed) with NE challenge. Cumulatively, SID+ had lower FI and FCR than NC+, but no change in BW or BWG. The combination treatment (SID+) often had an additive effect compared to SD+ or SI+ alone on mRNA abundance and D7 cecal fatty acid profiles. SD+ and SID+ also had higher D42 lean:fat ratios compared to NC+. Last, a 42-day study was conducted using 1,630 male and female Ross 708 broilers and the in ovo application of probiotics and subsequent NE challenge with five treatments. NC: sterile water in ovo, fed basal corn/soybean meal mash diet without NE challenge; AGP+: sterile water in ovo, fed basal diet with virginiamycin (0.5 kg/MT of feed) as AGP with NE challenge; NC+: NC treatment, with NE challenge; PI+: probiotic in ovo, fed basal diet, with NE challenge; PD+: sterile water in ovo, fed basal diet supplemented with probiotic (1.3 kg/MT of feed), with NE challenge. The use of probiotics in this study had little effect on performance, lean:fat ratios, and cecal fatty acid profiles, but PD+ increased mRNA abundance of D14 TLR2, D14 TNF-α, and D42 LEAP2 in cecal tonsils compared to controls. PI+ increased mRNA abundance of D7 and D42 MUC2, D7 LEAP2, and D42 TNF-α in the ileum. PI+ increased mRNA abundance in the cecal tonsils of D7 TLR2 and D42 TNF-α. These studies yielded interesting results about probiotics and synbiotics during a NE challenge by evaluating performance, intestinal immune responses, and fatty acid profiles in the ceca of broilers. In conclusion, the probiotic in this study did not improve broiler health during a NE challenge, but synbiotic use in ovo and continuation in the feed showed improvement over in ovo or dietary application alone. Synbiotic improved FCR over a challenged control, and altered mRNA abundance in the small intestine.
- Intermittent Bolus Compared with Continuous Feeding Enhances Insulin and Amino Acid Signaling to Translation Initiation in Skeletal Muscle of Neonatal PigsSuryawan, Agus; El-Kadi, Samer Wassim; Nguyen, Hanh V.; Fiorotto, Marta L.; Davis, Teresa A. (Oxford University Press, 2021-09-01)Background: Nutrition administered as intermittent bolus feeds rather than continuously promotes greater protein synthesis rates in skeletal muscle and enhances lean growth in a neonatal piglet model. The molecular mechanisms responsible remain unclear. Objectives: We aimed to identify the insulin- and/or amino acid-signaling components involved in the enhanced stimulation of skeletal muscle by intermittent bolus compared to continuous feeding in neonatal pigs born at term. Methods: Term piglets (2-3 days old) were fed equal amounts of sow milk replacer [12.8 g protein and 155 kcal/(kg body weight · d)] by orogastric tube as intermittent bolus meals every 4 hours (INT) or by continuous infusion (CTS). After 21 days, gastrocnemius muscle samples were collected from CTS, INT-0 (before a meal), and INT-60 (60 minutes after a meal) groups (n = 6/group). Insulin- and amino acid-signaling components relevant to mechanistic target of rapamycin complex (mTORC) 1 activation and protein translation were measured. Results: Phosphorylation of the insulin receptor, IRS-1, PDK1, mTORC2, pan-Akt, Akt1, Akt2, and TSC2 was 106% to 273% higher in the skeletal muscle of INT-60 piglets than in INT-0 and CTS piglets (P < 0.05), but phosphorylation of PTEN, PP2A, Akt3, ERK1/2, and AMPK did not differ among groups, nor did abundances of PHLPP, SHIP2, and Ubl4A. The association of GATOR2 with Sestrin1/2, but not CASTOR1, was 51% to 52% lower in INT-60 piglets than in INT-0 and CTS piglets (P < 0.05), but the abundances of SLC7A5/LAT1, SLC38A2/SNAT2, SLC38A9, Lamtor1/2, and V-ATPase did not differ. Associations of mTOR with RagA, RagC, and Rheb and phosphorylation of S6K1 and 4EBP1, but not eIF2α and eEF2, were 101% to 176% higher in INT-60 piglets than in INT-0 and CTS piglets (P < 0.05). Conclusions: The enhanced rates of muscle protein synthesis and growth with intermittent bolus compared to continuous feeding in a neonatal piglet model can be explained by enhanced activation of both the insulin- and amino acid-signaling pathways that regulate translation initiation.
- Intermittent Bolus Feeding Enhances Organ Growth More Than Continuous Feeding in a Neonatal Piglet ModelEl-Kadi, Samer Wassim; Boutry-Regard, Claire; Suryawan, Agus; Nguyen, Hanh V.; Kimball, Scot R.; Fiorotto, Marta L.; Davis, Teresa A. (2021-01)Background: Orogastric tube feeding is frequently prescribed for neonates who cannot ingest food normally. In a piglet model of the neonate, greater skeletal muscle growth is sustained by upregulation of translation initiation signaling when nutrition is delivered by intermittent bolus meals, rather than continuously. Objectives: The objective of this study was to determine the long-term effects of feeding frequency on organ growth and the mechanism by which feeding frequency modulates protein anabolism in these organs. Methods: Eighteen neonatal pigs were fed by gastrostomy tube the same amount of a sow milk replacer either by continuous infusion (CON) or on an intermittent bolus schedule (INT). After 21 d of feeding, the pigs were killed without interruption of feeding (CON; n = 6) or immediately before (INT-0; n = 6) or 60 min after (INT-60; n = 6) a meal, and fractional protein synthesis rates and activation indexes of signaling pathways that regulate translation initiation were measured in the heart, jejunum, ileum, kidneys, and liver. Results: Compared with continuous feeding, intermittent feeding stimulated the growth of the liver (+64%), jejunum (+48%), ileum (+40%), heart (+64%), and kidney (+56%). The increases in heart, kidney, jejunum, and ileum masses were proportional to whole body lean weight gain, but liver weight gain was greater in the INT-60 than the CON, and intermediate for the INT-0 group. For the liver and ileum, but not the heart, kidney, and jejunum, INT-60 compared with CON pigs had greater fractional protein synthesis rates (22% and 48%, respectively) and was accompanied by an increase in ribosomal protein S6 kinase 1 and eukaryotic initiation factor 4E binding protein 1 phosphorylation. Conclusions: These results suggest that intermittent bolus compared with continuous orogastric feeding enhances organ growth and that in the ileum and liver, intermittent feeding enhances protein synthesis by stimulating translation initiation.
- Intermittent bolus feeding promotes greater lean growth than continuous feeding in a neonatal piglet modelEl-Kadi, Samer Wassim; Boutry, Claire; Suryawan, Agus; Gazzaneo, Maria C.; Orellana, Renan A.; Srivastava, Neeraj; Nguyen, Hanh V.; Kimball, Scot R.; Fiorotto, Marta L.; Davis, Teresa A. (2018-10)Background: Orogastric tube feeding is indicated in neonates with an impaired ability to ingest food normally and can be administered with an intermittent bolus or continuous feeding schedule. Objectives: The objectives were to 1) compare the long-term effect of continuous with intermittent feeding on growth using the newborn pig as a model, 2) determine whether feeding frequency alters lean tissue and fat mass gain, and 3) identify the signaling mechanisms by which protein deposition is controlled in skeletal muscle in response to feeding frequency. Design: Neonatal pigs were fed the same amount of a balanced formula by orogastric tube either as an intermittent bolus meal every 4 h (INT) or as a continuous infusion (CON). Body composition was assessed at the start and end of the study by dual-energy X-ray absorptiometry, and hormone and substrate profiles, muscle mass, protein synthesis, and indexes of nutrient and insulin signaling were measured after 21 d. Results: Body weight, lean mass, spine length, and skeletal muscle mass were greater in the INT group than in the CON group. Skeletal muscle fractional protein synthesis rates were greater in the INT group after a meal than in the CON group and were associated with higher circulating branched-chain amino acid and insulin concentrations. Skeletal muscle protein kinase B (PKB) and ribosomal protein S6 kinase phosphorylation and eukaryotic initiation factor (eIF) 4E-eIF4G complex formation were higher, whereas eIF2 alpha phosphorylation was lower in the INT group than in the CON group, indicating enhanced activation of insulin and amino acid signaling to translation initiation. Conclusions: These results suggest that when neonates are fed the same amounts of nutrients as intermittent meals rather than continuously there is greater lean growth. This response can be ascribed, in part, to the pulsatile pattern of amino acids, insulin, or both induced by INT, which enables the responsiveness of anabolic pathways to feeding to be sustained chronically in skeletal muscle.
- Muscle Growth and Development in Intrauterine Growth Restricted PigsZhu, Haibo (Virginia Tech, 2015-03-16)Intrauterine growth restriction causes impaired growth and development of mammalian fetus, and leads to long-term negative effect on postnatal growth. Among domestic animals, pigs exhibit the most severe naturally occurring IUGR and reduced postnatal muscle growth. The objectives of this research project were to: 1) determine muscle stem cell characteristics in IUGR pigs; 2) determine how intrauterine growth restriction alters protein deposition in skeletal muscle; 3) investigate whether branched-chain amino acids (BCAA) are able to enhance protein synthesis in intrauterine growth restricted (IUGR) pig muscle. Newborn piglets were considered normal body weight (NBWT) or IUGR when birth weight was within ± 0.5 SD and -2 SD of litter average respectively. Muscle satellite cell numbers, believed to be the major nuclei source for postnatal muscle growth, were lower in newborn IUGR pigs which could result in reduced muscle hypertrophy potential. In addition, cultures derived from IUGR muscle satellite cells had a lower fusion percentage. Fewer satellite cells and impaired differentiation ability may contributor to impaired muscle growth in these pigs. Protein synthesis rate was significantly lower in IUGR pig hindquarter in the first hour after feeding, but BCAA supplementation had no effect on protein synthesis in IUGR pigs. Further, eukaryotic translation initiation factor 4E (eIF4E) expression is down regulated in IUGR pig muscle. These results suggest that impaired translation initiation may provide a plausible explanation for the lower protein synthesis rates observed in IUGR pigs. Overall, reduced muscle stem cell number and changes in their activity, as well as impaired translation initiation may be important explanations for compromised postnatal muscle growth in intrauterine growth restricted pigs.