![[Feedback]](/icons/banner/feedbackACT.gif){kind=icon}
![[For Subscribers]](/icons/banner/subscriberACT.gif){kind=icon}
![[Archive]](/icons/banner/archiveACT.gif){kind=icon}
![[Search]](/icons/banner/searchACT.gif){kind=icon}
![[Contents]](/icons/banner/tocACT.gif){kind=icon}
|
|
|
|
|
||
U.S. Department of Agriculture/Agriculture Research Service, Children’s Nutrition Research Center, Department of Pediatrics, Baylor College of Medicine, Houston, TX 77030 and * Department of Cellular and Molecular Physiology, Pennsylvania State University College of Medicine, Hershey, PA 17033
3To whom correspondence should be addressed. E-mail: tdavis{at}bcm.tmc.edu.
 |
ABSTRACT |
|---|
 TOP ABSTRACT MATERIALS AND METHODSRESULTSDISCUSSIONLITERATURE CITED |
|---|
KEY WORDS: • pigs • dietary protein • S6K1 • 4E-binding protein 1 • protein synthesis
The American Academy of Pediatrics recommends that preterm infants should be provided nutrients to support growth rates similar to the normal in utero growth rates (1). Despite improvements in the nutritional management of very-low-birth-weight (VLBW)4 infants over the past decade (2–4), most are discharged weighing less than the tenth percentile of intrauterine growth standards (5). Many remain small into adulthood, and some exhibit adverse, long-term developmental outcomes including learning impairment and reduced work capacity (6–9). Consumption of a high-protein diet [4.3 vs. 2.25 g/(kg · d)] can increase length and weight gain of VLBW infants (10–13). A recent study demonstrated that whole-body protein accretion and synthesis rates during the first days of life in VLBW infants are increased by parenteral feeding of 2.65 vs. 0.85 g amino acids/(kg · d) (14). However, the influence of a high-protein diet on rates of protein synthesis in specific tissues and the mechanisms that regulate the response have not been examined.
Due to the invasiveness of the techniques required to evaluate the effects of amino acids on tissue protein synthesis rates, neonatal pigs have been used as a research model for neonatal infants (15–17). Our laboratory showed that the feeding of sow’s milk stimulates protein synthesis in all tissues in neonatal pigs (16), and that this response can be reproduced with the i.v. infusion of insulin and amino acids (18–20). More specifically, muscle protein synthesis is responsive to both insulin and amino acids, whereas protein synthesis in liver and other visceral tissues is responsive only to amino acids (21). In fact, utilizing a hyperinsulinemic-euglycemic-euaminoacidemic clamp, it was demonstrated that muscle protein synthesis dose dependently increased as both insulin and amino acids levels increase (22), and that a plateau in protein synthesis in skeletal muscle was not achieved even at the highest infusion rate of amino acids (23). The intracellular mechanisms by which insulin and amino acids regulate protein synthesis were also investigated. Hyperphosphorylation of the eukaryotic initiation factor (eIF) 4E repressor protein, 4E binding protein 1 (4E-BP1), and 70-kDa ribosomal protein S6 kinase 1 (S6K1), which regulate the binding of mRNA to the 40S ribosomal complex, were positively correlated with both insulin and the amino acid infusion level (23). Because the effects of amino acids were linear, it would appear that higher levels of amino acids could further enhance protein synthesis and translation initiation factor activation. Although the research evaluated the acute effects of amino acids infused i.v. on protein synthesis and translation initiation factor activation, the effects of dietary protein level were not examined previously.
For the present study, the effects of prolonged high dietary protein intake above the NRC requirement (24) were determined. Piglets were weaned from the sow and fed artificial milk diets that contained marginal, adequate, or high levels of protein. Tissue protein synthesis rates and translation initiation factor activation were measured in either the food-deprived or fed state. The results show that feeding artificial milk diets stimulates protein synthesis in skeletal muscle and liver of neonatal pigs by modulating the translation initiation factors that regulate mRNA binding to the ribosomal complex. However, provision of a high-protein diet that exceeds the protein requirement does not further enhance protein synthesis or translation initiation factor activation.
|
MATERIALS AND METHODS |
|---|
|
TOPABSTRACTMATERIALS AND METHODSRESULTSDISCUSSIONLITERATURE CITED |
|---|
|
|
Hormone and substrate assays. The concentration of blood glucose was analyzed using a YSI 2300 STAT Plus (Yellow Springs Instruments). Plasma total BCAA were analyzed by rapid enzymatic kinetic assay (25). Plasma radioimmunoreactive insulin concentrations were measured using a porcine insulin RIA kit (Linco) that used porcine insulin antibody and human insulin standards. Plasma urea nitrogen (PUN; Biotron Diagnostics, Hemet, CA, Blood Urea Nitrogen #80-e), creatinine (CREA; Cayman Chemicals, Creatinine Assay Kit #500701), and ammonia (Sigma, Ammonia Assay Kit #AA0100) were measured using commercially available kits.
Tissue protein synthesis in vivo.
Tissue protein synthesis was measured in vivo using a modification of the flooding-dose technique (26). At 60 min, pigs were injected via the jugular vein catheter with 10 mL/kg BW of a flooding dose of phenylalanine (Amersham), which provided 1.5 mmol phenylalanine/kg BW and 1 mCi of L-[4-3H]phenylalanine/kg BW. Samples of whole blood were taken 5, 15, and 30 min after the injection of [3H]phenylalanine for measurement of the specific radioactivity of the extracellular free pool of phenylalanine. Immediately after the 30-min blood sample was taken, pigs were given a lethal injection of sodium pentobarbital (50 mg/kg body weight). Fractional rates of protein synthesis (Ks; percentage of protein mass synthesized in a day) for each tissue were calculated as:
 |
where Sb (dpm/min) is the specific radioactivity of the protein-bound phenylalanine, Sa (dpm/min) is the specific radioactivity of the tissue-free phenylalanine at the time of tissue collection and the linear regression of the blood specific radioactivity of the pig at 5, 15, and 30 min against time, the constant T equals 1,440 min/d, and t is the time of labeling in minutes of the specific tissue. We demonstrated previously that the specific radioactivity of the tissue-free phenylalanine after a flooding dose of phenylalanine is in equilibrium with the aminoacyl-tRNA specific radioactivity; hence, the tissue-free phenylalanine reflects the specific radioactivity of the tissue precursor pool (27).
Protein immunoblot analysis. Homogenized muscle and liver proteins separated on polyacrylamide gels were electrophoretically transferred to a polyvinylidene difluoride transfer membrane (BioRad) as previously described (28). The membrane was incubated with primary antibody overnight followed by a 1-h incubation with secondary antibody. The membrane was then washed with Tris-buffered saline-Tween 20 solution. Blots were developed using an enhanced chemiluminescence Western blotting kit (ECL-plus, Amersham), visualized using a GeneGenome bioimaging system, and analyzed using GeneTools software (Syngene).
Measurement of protein kinase B (PKB/Akt) phosphorylation. To determine PKB phosphorylation on Ser473, samples were subjected to SDS-PAGE, and the phosphorylation of PKB on Ser473 was determined by protein immunoblot analysis with an antibody that recognizes the protein only when it is phosphorylated on that residue or a separate antibody that recognizes total PKB. Antibodies were obtained from Cell Signaling. Data are expressed as arbitrary units (AU).
Measurement of S6K1 phosphorylation. Muscle homogenates were combined with an equal volume of SDS sample buffer, and the diluted samples were subjected to electrophoresis on a 7.5% polyacrylamide gel. The samples were then subjected to protein immunoblot analysis by using a rabbit anti-rat S6K polyclonal antibody (Santa Cruz Biotechnology), as previously described (29). Data are reported as the percentage of phosphorylation.
Examination of 4E-BP1 phosphorylation on Thr70. Aliquots of muscle homogenates (supernatants) were centrifuged at 10,000 x g for 10 min at 4°C and then frozen at –80°C until analyzed. The supernatants were diluted with SDS sample buffer and then subjected to protein immunoblot analysis, as described previously (28,29). The membranes were incubated with a polyclonal antibody that specifically recognizes phosphorylation of 4E-BP1 at Thr70 (Cell Signaling). Data are reported as the percentage of phosphorylation.
Statistics. Treatments were arranged as a 2 x 3 factorial, with 2 feeding states (food-deprived and fed) and 3 levels of protein intake (21, 33, and 45%). Data were analyzed using the General Linear Models procedure of SAS (SAS Institute). The statistical model included the main effects and interaction of feeding state and protein level. Linear and quadratic polynomial contrasts were used to determine the effects of increasing dietary protein in the food-deprived or fed state. Differences were considered significant at P < 0.05. Data are presented as means ± SEM.
|
RESULTS |
|---|
|
TOPABSTRACTMATERIALS AND METHODSRESULTSDISCUSSIONLITERATURE CITED |
|---|
|
Plasma insulin, BCAA, and glucose. Plasma insulin concentrations were higher in fed pigs than in food-deprived pigs at 60 and 90 min (P < 0.001) (Table 4). Plasma insulin levels in the 45% protein groups were lower than in the 21 or 33% protein groups, resulting in the main effect of dietary protein at 60 min (P < 0.01). Also at 60 min, there was a significant feeding state x protein interaction (P < 0.01). In the food-deprived state, plasma insulin levels decreased linearly (P < 0.01) as protein intake increased; in the fed state, plasma insulin levels were lowest for pigs fed high protein.
|
Plasma glucose concentrations were higher in fed pigs than in food-deprived pigs at 60 and 90 min (P < 0.001). In addition, plasma glucose tended to decrease as protein intake increased at 60 min (P = 0.07).
Plasma lactate, urea nitrogen, CREA, and ammonia. Plasma lactate and CREA were not affected by feeding state or dietary protein level (P = 0.26) (Table 5). However, PUN and ammonia concentrations increased as protein intake increased (P < 0.001 and P < 0.02, respectively).
|
|
|
|
DISCUSSION |
|---|
|
TOPABSTRACTMATERIALS AND METHODSRESULTSDISCUSSIONLITERATURE CITED |
|---|
One objective of the study was to confirm previous findings that feeding stimulates an increase in protein synthesis (15,16) and the activation of translation initiation factors (29,31). The effect of feeding on protein synthesis in neonates is attributed to the increase in circulating insulin and amino acid concentrations in plasma (20,21) and is associated with the activation of translation initiation factors along the insulin-signaling pathway (30). Earlier studies utilized colostrum or mature sow’s milk as the enteral source of nutrients (15,16). In this study, liquid diets containing whey protein, dried skim milk, and lactose as the predominant ingredients were fed. As expected, we observed an increase in plasma insulin and amino acids concentrations, and a subsequent increase in protein synthesis and translation initiation factor activation in response to feeding the liquid diets. The relative increases in the fractional rates of protein synthesis observed in our study were similar to those reported by Burrin et al. (15) and Davis et al. (16).
The effect of feeding on protein synthesis is likely mediated through the insulin- and nutrient-signaling pathways via an mammalian target of rapamycin (mTOR)-dependent process (31). More specifically, the phosphorylation of PKB, S6K1, and 4E-PB1 increases in muscle with feeding mature sow’s milk (29–31). PKB is a serine/threonine kinase that is activated by insulin and plays a major role in linking PI3K activation to protein synthesis (32). Downstream of PKB and mTOR lie both S6K1 and 4E-BP1. Once S6K1 becomes phosphorylated on multiple serine and threonine residues, it becomes activated, and then phosphorylates ribosomal protein S6, which may lead to an increase in translation of mRNAs containing a terminal oligopyrimidine sequence adjacent to the m7GTP cap structure at the 5' end of the message (33,34). The phosphorylation of 4E-BP1 causes this repressor protein to dissociate from eIF4E, allowing eIF4E to bind with eIF4G to eventually form the eIF4F complex. This eIF4F complex mediates binding of mRNA to the 40S ribosomal subunit in the initiation of mRNA translation (35). The increase in phosphorylation of PKB in response to the feeding of our liquid formula diets is likely the result of the rise in circulating insulin concentrations, whereas the increases in phosphorylation of the mTOR-dependent initiation factors, S6K1 and 4E-BP1, are most likely attributed to the rise in both insulin and amino acid concentrations (36–38).
In rats, it was also shown that feeding stimulates protein synthesis and 4E-BP1 phosphorylation in skeletal muscle and liver (39,40). In fact, Yoshizawa et al. (39) demonstrated by feeding a 20% protein vs. a protein-free diet to rats that the feeding-induced stimulation of protein synthesis and phosphorylation of 4E-BP1 in skeletal muscle and liver is protein dependent. However, they did not rule out the possibility that an increase in plasma insulin is required for the stimulation of protein synthesis by the dietary protein. Unfortunately, graded levels of dietary protein were not investigated.
Using insulin-glucose-amino acid clamp techniques developed in our laboratory, we showed previously that muscle protein synthesis is regulated by the infusion of both insulin and amino acids (21). Interestingly, once plasma insulin concentrations reached 
70 pmol/L, no additional increase in muscle protein synthesis occurred when amino acids were clamped at either the food-deprived or fed level (22). In contrast to muscle, protein synthesis in liver, kidney, and pancreas were not affected by increasing insulin concentrations, but by increasing amino acid concentrations. In our previous infusion studies, amino acids were infused at a rate to achieve levels similar to that of piglets fed mature sow’s milk. This corresponds to 1000 µmol/L of BCAA. At this highest level of infusion, fractional rates of protein synthesis in skeletal muscle were still increasing (23). Our goal with the current feeding study was to further increase the concentration of BCAA with our high-protein diet to determine whether protein synthesis continued to increase. Although we raised BCAA levels to 1350 µmol/L in the piglets fed the 45% protein diet, we did not consistently observe increases in protein synthesis rates in any tissues on d 7. Thus, it is likely that maximal rates of protein synthesis in these tissues were reached at lower protein intakes.
Interestingly, growth rates from d 2 to 7 were higher in piglets fed the adequate and high-protein diets compared with piglets fed the marginal diet. Because protein synthesis rates on d 7 did not parallel growth rates, it is likely that the higher dietary protein intakes may have increased protein synthesis before d 7, resulting in the overall higher growth rates. Alternatively, it is possible that degradation rates were lower in piglets fed adequate and high protein compared with those fed low protein, leading to the overall increase in growth. Indeed, i.v. amino acid infusion was reported to suppress whole-body proteolysis in newborn humans (41), although not in adults (42). However, we cannot discount the possibility that the differences in growth rates were due to differences in lipid rather than protein accretion.
O’Connor et al. (23,43) demonstrated that phosphorylation of S6K1 and 4E-BP1 was highly correlated with the level of infusion of both insulin and amino acids in skeletal muscle and liver. As was observed with protein synthesis, phosphorylation of these proteins increased in a linear manner up to the highest infusion level of amino acids. Amino acids exert their effects via mTOR, a signaling protein that phosphorylates S6K1 and 4E-BP1 (36–38). In our current feeding study, we did not observe consistent increases in the phosphorylation of S6K1 or 4E-BP1 with increasing protein intake; however, these signaling proteins were already highly phosphorylated at the lowest protein intake. In fact, the high phosphorylation of S6K1 and 4E-BP1, even at the lowest protein intake, may help explain why high levels of dietary protein did not further increase protein synthesis in skeletal muscle and liver.
The diets used in this study were designed to provide isocaloric intakes; therefore, as protein content increased, the amount of lactose decreased. This necessary change in the diet composition of 2 macronutrients complicates the interpretation of the data. For example, the lower lactose level likely influenced the plasma insulin concentrations of these piglets. This change in insulin level may have affected the phosphorylation of PKB, in which lactose intake and insulin concentration decreased as protein intake increased, and therefore phosphorylation of PKB decreased. The reduction in PKB phosphorylation, in both the food-deprived and fed states, may have blunted any effect of increasing amino acid level on S6K1 and 4E-BP1 phosphorylation and protein synthesis rates. Because muscle protein synthesis in neonatal pigs is exquisitely sensitive to very low concentrations of insulin (22), it seems probable that the decrease in muscle protein synthesis in the food-deprived state as protein level increased (and lactose level decreased) was due to the decrease in circulating insulin level. Feeding studies designed to evaluate the effects of dietary carbohydrate level as a stimulus of insulin secretion and the subsequent effects on translation initiation factor activation and protein synthesis should clarify the effects just described.
As indicated previously, there were no additional increases in protein synthesis or translation initiation factor activation with the provision of the high-protein diet. The 21, 33, and 45% diets used in our study provided 12.6, 19.8, and 27 g protein/(kg BW · d), respectively. Based on the NRC (24) nutrient requirements for swine, a 2-kg piglet requires on the order of 16.9 g protein/(kg BW · d) to maximize growth rate. Although less information is available for human infants compared with piglets, the current recommended protein intake for a low-birth-weight infant (BW < 2500 g) consuming 120 kcal/d is 4 g/(kg BW · d) (1,44). Thus, the protein requirement for piglets is roughly 323% greater than that of low-birth-weight infants. Therefore, the protein intakes achieved in our study would be comparable to 3.0, 4.7, and 6.4 g/(kg BW · d) for low-birth-weight infants.
Protein intakes > 5.0 g/(kg BW · d) have been associated with neurodevelopmental deficiencies in low-birth-weight infants (44). Goldman et al. (45–47) observed that low-birth-weigh infants weighing <2000 g BW fed 6.0–7.2 g protein/(kg BW · d) had more fever, lethargy, and frequency of strabismus than infants fed between 3.0 and 3.6 g/(kg BW · d). Low-birth-weight infants weighing <1300 g given the higher protein intake also had significantly increased incidence of low IQ scores. Since the time this work was reported, little research has been published investigating higher protein intakes in infants due to ethical considerations. Using the neonatal pig model, we were able to feed very high-protein diets with less concern for the possible developmental problems that are associated with protein toxicity. However, it is unlikely that our diets would have led to protein toxicity in the piglets given their extremely high potential for protein deposition up to 7 d of age. Nevertheless, measures of protein metabolism were evaluated. Although liver weight, kidney weight, PUN, and plasma ammonia all increased with protein intake, these effects were reported previously (48–50). The plasma ammonia concentrations observed in our feeding study were well below a hyperammonemia value of 270 µmol/L determined by Brunton et al. (51). In addition, brain weight, plasma lactate, and plasma CREA were not affected by protein intake, nor were there any observed morphological abnormalities in any tissues. In sum, our high-protein diet did not elicit any measurable degree of toxicity in the piglets.
Research from our laboratory has clearly demonstrated that feeding stimulates muscle protein synthesis in neonatal pigs, and this response can be reproduced with the infusion of insulin and amino acids. The stimulation of muscle protein synthesis is mediated by the activation of signaling proteins and translation initiation factors including S6K1 and 4E-BP1. The current study was an attempt to apply the knowledge generated by our previous research by feeding high levels of protein and measuring biomarkers of translation initiation and protein synthesis. The results of the study confirm that feeding stimulates protein synthesis by modulating the activation of translation initiation factors that regulate the binding of mRNA to the ribosomal complex. Furthermore, this study demonstrates that the provision of a high-protein diet does not further enhance protein synthesis or translation initiation factor activation, although we cannot discount the possibility that the reduction in plasma insulin levels due to the lower lactose intake may have blunted the effect of increasing amino acid concentrations on protein synthesis. Overall, the results of this study suggest that the recommended level of protein intake for neonatal pigs to maximize growth rate also provides an adequate supply of protein to maximize fractional rates of protein synthesis in skeletal muscle and liver.
|
ACKNOWLEDGMENTS |
|---|
|
FOOTNOTES |
|---|
2 Funded in part by the National Institutes of Health grants R01-AR-44474 (T.A.D.), DK-15658 (L.S.J.), and DK-13499 (L.S.J.) and by the USDA/ARS under Cooperative Agreement no. 58–6250–6–001 (T.A.D.).
4 Abbreviations used: BW, body weight; CREA, creatinine; eIF4E, eukaryotic initiation factor 4E; 4E-BP1, eIF4E-binding protein 1; mTOR, mammalian target of rapamycin; PI3K, phosphoinositide 3-kinase; PKB, protein kinase B (Akt); PUN, plasma urea nitrogen; S6K1, 70-kDa ribosomal protein S6 kinase 1; VLBW, very low birth weight.
Manuscript received 20 January 2005. Initial review completed 12 February 2005. Revision accepted 28 March 2005.
|
LITERATURE CITED |
|---|
|
TOPABSTRACTMATERIALS AND METHODSRESULTSDISCUSSIONLITERATURE CITED |
|---|
1. American Academy of Pediatrics (2003) Nutritional needs of preterm infants. Kleinman, R. E. eds. Pediatric Nutrition Handbook 4th ed. 2003:23-54 American Academy of Pediatrics Elk Grove Village, IL. .
2. Hack, M., Horbar, J. D., Malloy, M. H., Tyson, J. E., Wright, E. & Wright, L. (1991) Very low birth weight outcomes of the National Institute of Child Health and Human Development Neonatal Network. Pediatrics 87:587-597.
3. Hack, M., Wright, L. L., Shankaran, S., Tyson, J. E., Horbar, J. D., Bauer, C. R. & Younes, N. (1995) Very-low-birth-weight outcomes of the National Institute of Child Health and Human Development Neonatal Network, November 1989 to October 1990. Am. J. Obstet. Gynecol. 172:457-464.[Medline]
4. Lemons, J. A., Bauer, C. R., Oh, W., Korones, S. B., Papile, L. A., Stoll, B. J., Verter, J., Temprosa, M., Wright, L. L., Ehrenkranz, R. A., Fanaroff, A. A., Stark, A., Carlo, W., Tyson, J. E., Donovan, E. F., Shankaran, S. & Stevenson, D. K. (2001) Very low birth weight outcomes of the National Institute of Child health and human development neonatal research network, January 1995 through December 1996. NICHD Neonatal Research Network. Pediatrics 107:E1.
5. Ehrenkranz, R. A., Younes, N., Lemons, J. A., Fanaroff, A. A., Donovan, E. F., Wright, L. L., Katsikiotis, V., Tyson, J. E., Oh, W., Shankaran, S., Bauer, C. R., Korones, S. B., Stoll, B. J., Stevenson, D. K. & Papile, L. A. (1999) Longitudinal growth of hospitalized very low birth weight infants. Pediatrics 104:280-289.
6. Peralta-Carcelen, M., Jackson, D. S., Goran, M. I., Royal, S. A., Mayo, M. S. & Nelson, K. G. (2000) Growth of adolescents who were born at extremely low birth weight without major disability. J. Pediatr. 136:633-640.[Medline]
7. Ford, G. W., Doyle, L. W., Davis, N. M. & Callanan, C. (2000) Very low birth weight and growth into adolescence. Arch. Pediatr. Adolesc. Med. 154:778-784.
8. Saigal, S., Stoskopf, B. L., Streiner, D. L. & Burrows, E. (2001) Physical growth and current health status of infants who were of extremely low birth weight and controls at adolescence. Pediatrics 108:407-415.
9. Lundgren, E. M., Cnattingius, S., Jonsson, B. & Tuvemo, T. (2001) Intellectual and psychological performance in males born small for gestational age with and without catch-up growth. Pediatr. Res. 50:91-96.[Medline]
10. Kashyap, S., Forsyth, M., Zucker, C., Ramakrishnan, R., Dell, R. B. & Heird, W. C. (1986) Effects of varying protein and energy intakes on growth and metabolic response in low birth weight infants. J. Pediatr. 108:955-963.[Medline]
11. Kashyap, S., Okamoto, E., Kanaya, S., Zucker, C., Abildskov, K., Dell, R. B. & Heird, W. C. (1987) Protein quality in feeding low birth weight infants: a comparison of whey-predominant versus casein-predominant formulas. Pediatrics 79:748-755.
12. Kashyap, S., Schulze, K. F., Forsyth, M., Zucker, C., Dell, R. B., Ramakrishnan, R. & Heird, W. C. (1988) Growth, nutrient retention, and metabolic response in low birth weight infants fed varying intakes of protein and energy. J. Pediatr. 113:713-721.[Medline]
13. Kashyap, S., Schulze, K. F., Ramakrishnan, R., Dell, R. B. & Heird, W. C. (1994) Evaluation of a mathematical model for predicting the relationship between protein and energy intakes of low-birth-weight infants and the rate and composition of weight gain. Pediatr. Res. 35:704-712.[Medline]
14. Thureen, P. J., Melara, D., Fennessey, P. V. & Hay, W. W., Jr (2003) Effect of low versus high intravenous amino acid intake on very low birth weight infants in the early neonatal period. Pediatr. Res. 53:24-32.[Medline]
15. Burrin, D. G., Shulman, R. J., Reeds, P. J., Davis, T. A. & Gravitt, K. R. (1992) Porcine colostrum and milk stimulate visceral organ and skeletal muscle protein synthesis in neonatal piglets. J. Nutr. 122:1205-1213.
16. Davis, T. A., Burrin, D. G., Fiorotto, M. L. & Nguyen, H. V. (1996) Protein synthesis in skeletal muscle and jejunum is more responsive to feeding in 7- than in 26-day-old pigs. Am. J. Physiol. 270:E802-E809.
17. House, J. D., Pencharz, P. B. & Ball, R. O. (1997) Tyrosine kinetics and requirements during total parenteral nutrition in the neonatal piglet: the effect of glycyl-L-tyrosine supplementation. Pediatr. Res. 41:575-583.[Medline]
18. Wray-Cahen, D., Beckett, P. R., Nguyen, H. V. & Davis, T. A. (1997) Insulin-stimulated amino acid utilization during glucose and amino acid clamps decreases with development. Am. J. Physiol. 273:E305-E314.[Medline]
19. Wray-Cahen, D., Nguyen, H. V., Burrin, D. G., Beckett, P. R., Fiorotto, M. L., Reeds, P. J., Wester, T. J. & Davis, T. A. (1998) Response of skeletal muscle protein synthesis to insulin in suckling pigs decreases with development. Am. J. Physiol. 275:E602-E609.
20. Davis, T. A., Burrin, D. G., Fiorotto, M. L., Reeds, P. J. & Jahoor, F. (1998) Roles of insulin and amino acids in the regulation of protein synthesis in the neonate. J. Nutr. 128:347S-350S.
21. Davis, T. A., Fiorotto, M. L., Burrin, D. G., Reeds, P. J., Nguyen, H. V., Beckett, P. R., Vann, R. C. & O’Connor, P. M. (2002) Stimulation of protein synthesis by both insulin and amino acids is unique to skeletal muscle in neonatal pigs. Am. J. Physiol. 282:E880-E890.
22. O’Connor, P. M., Bush, J. A., Suryawan, A., Nguyen, H. V. & Davis, T. A. (2003) Insulin and amino acids independently stimulate skeletal muscle protein synthesis in neonatal pigs. Am. J. Physiol. 284:E110-E119.
23. O’Connor, P. M., Kimball, S. R., Suryawan, A., Bush, J. A., Nguyen, H. V., Jefferson, L. S. & Davis, T. A. (2003) Regulation of translation initiation by insulin and amino acids in skeletal muscle of neonatal pigs. Am. J. Physiol. 285:E40-E53.
24. NRC (1998) Nutrient Requirements of Swine 10th rev. ed. 1998 National Academy Press Washington, DC.
25. Beckett, P. R., Hardin, D. S., Davis, T. A., Nguyen, H. V., Wray-Cahen, D. & Copeland, K. C. (1996) Spectrophometric assay for measuring branched-chain amino acid concentrations: application for measuring the sensitivity of protein metabolism to insulin. Anal. Biochem. 240:48-53.[Medline]
26. Garlick, P. J., McNurlan, M. A. & Preedy, V. R. (1980) A rapid and convenient technique for measuring the rate of protein synthesis in tissues by injection of [3H]phenylalanine. Biochem. J. 192:719-723.[Medline]
27. Davis, T. A., Fiorotto, M. L., Nguyen, H. V. & Burrin, D. G. (1999) Aminoacyl-tRNA and tissue free amino acid pools are equilibrated after a flooding dose of phenylalanine. Am. J. Physiol. 277:E103-E109.
28. Kimball, S. R., Karinch, A. M., Feldhoff, R. C., Mellor, H. & Jefferson, L. S. (1994) Purification and characterization of eukaryotic translational initiation factor eIF-2B from liver. Biochim. Biophys. Acta 1201:473-481.[Medline]
29. Davis, T. A., Nguyen, H. V., Suryawan, A., Bush, J. A., Jefferson, L. S. & Kimball, S. R. (2000) Developmental changes in the feeding-induced stimulation of translation initiation in muscle of neonatal pigs. Am. J. Physiol. 279:E1226-E1234.
30. Kimball, S. R., Farrell, P. A., Nguyen, H. V., Jefferson, L. S. & Davis, T. A. (2002) Developmental decline in components of signal transduction pathways regulating protein synthesis in pig muscle. Am. J. Physiol. 282:E585-E592.
31. Kimball, S. R., Jefferson, L. S., Nguyen, H. V., Suryawan, A., Bush, J. A. & Davis, T. A. (2000) Feeding stimulates protein synthesis in muscle and liver of neonatal pigs through an mTOR-dependent process. Am. J. Physiol. 279:E1080-E1087.
32. Scheid, M. P. & Woodgett, J. R. (2003) Unravelling the activation mechanisms of protein kinase B/Akt. FEBS Lett. 546:108-112.[Medline]
33. Fumagalli, S. & Thomas, G. (2000) S6 phosphorylation and signal transduction. Sonenberg, N. Hershey, J.W.B. Mathews, M. B. eds. Translational Control of Gene Expression 2000:695-717 Cold Spring Harbor Laboratory Press Cold Spring Harbor, NY. .
34. Meyuhas, O. & Hornstein, E. (2000) Translational control of TOP mRNAs. Sonenberg, N. Hershey, J.W.B. Mathews, M. B. eds. Translational Control of Gene Expression 2000:671-693 Cold Spring Harbor Laboratory Press Cold Spring Harbor, NY. .
35. Gingras, A. C., Raught, B. & Sonenberg, N. (1999) eIF4 initiation factors: effectors of mRNA recruitment to ribosomes and regulators of translation. Annu. Rev. Biochem. 68:913-963.[Medline]
36. Hara, K., Yonezawa, K., Weng, Q. P., Kozlowski, M. T., Belham, C. & Avruch, J. (1998) Amino acid sufficiency and mTOR regulate p70 S6 kinase and eIF-4E BP1 through a common effector mechanism. J. Biol. Chem. 273:14484-14494.
37. Patti, M. E., Brambilla, E., Luzi, L., Landaker, E. J. & Kahn, C. R. (1998) Bidirectional modulation of insulin action by amino acids. J. Clin. Investig. 101:1519-1529.[Medline]
38. Wang, X., Campbell, L. E., Miller, C. M. & Proud, C. G. (1998) Amino acid availability regulates p70 S6 kinase and multiple translation factors. Biochem. J. 334:261-267.
39. Yoshizawa, F., Kimball, S. R., Vary, T. C. & Jefferson, L. S. (1998) Effect of dietary protein on translation initiation in rat skeletal muscle and liver. Am. J. Physiol. 275:E814-E820.
40. Preedy, V. R. & Garlick, P. J. (1986) The response of muscle protein synthesis to nutrient intake in postabsorptive rats: the role of insulin and amino acids. Biosci. Rep. 6:177-183.[Medline]
41. Poindexter, B. B., Karn, C. A., Ahlrichs, J. A., Wang, J., Leitch, C. A., Liechty, E. A. & Denne, S. C. (1997) Amino acids suppress proteolysis independent of insulin throughout the neonatal period. Am. J. Physiol. 272:E592-E599.
42. Tessari, P., Inchiostro, S., Biolo, G., Trevisan, R., Fantin, G., Marescotti, M. C., Iori, E., Tiengo, A. & Crepaldi, G. (1987) Differential effects of hyperinsulinemia and hyperaminoacidemia on leucine-carbon metabolism in vivo. Evidence for distinct mechanisms in regulation of net amino acid deposition. J. Clin. Investig. 79:1062-1069.
43. O’Connor, P. M., Kimball, S. R., Suryawan, A., Bush, J. A., Nguyen, H. V., Jefferson, L. S. & Davis, T. A. (2004) Regulation of neonatal liver protein synthesis by insulin and amino acids in pigs. Am. J. Physiol. 286:E994-E1003.
44. Klein, C. J. (2002) Nutrient requirements for preterm infant formulas. J. Nutr. 132:1395S-1577S.
45. Goldman, H. I., Goldman, J., Kaufman, I. & Liebman, O. B. (1974) Late effects of early dietary protein intake on low-birth-weight infants. J. Pediatr. 85:764-769.[Medline]
46. Goldman, H. I., Freudenthal, R., Holland, B. & Karelitz, S. (1969) Clinical effects of two different levels of protein intake on low-birth-weight infants. J. Pediatr. 74:881-889.[Medline]
47. Goldman, H. I., Liebman, O. B., Freudenthal, R. & Reuben, R. (1971) Effects of early dietary protein intake on low-birth-weight infants: evaluation at 3 years of age. J. Pediatr. 78:126-129.[Medline]
48. Eggum, B. O. (1970) Blood urea measurement as a technique for assessing protein quality. Br. J. Nutr. 24:983-988.[Medline]
49. Chen, H. Y., Lewis, A. J., Miller, P. S. & Yen, J. T. (1999) The effect of excess protein on growth performance and protein metabolism of finishing barrows and gilts. J. Anim. Sci. 77:3238-3247.
50. Coma, J., Carrion, D. & Zimmerman, D. R. (1995) Use of plasma urea nitrogen as a rapid response criterion to determine the lysine requirement of pigs. J. Anim. Sci. 73:472-481.[Abstract]
51. Brunton, J. A., Bertolo, R. F., Pencharz, P. B. & Ball, R. O. (1999) Proline ameliorates arginine deficiency during enteral but not parenteral feeding in neonatal piglets. Am. J. Physiol. 277:E223-E231.
This article has been cited by other articles:
|
|
 |
|
  C. L. Yuan, N. Sharma, D. A. Gilge, W. C. Stanley, Y. Li, M. Hatzoglou, and S. F. Previs Preserved protein synthesis in the heart in response to acute fasting and chronic food restriction despite reductions in liver and skeletal muscle Am J Physiol Endocrinol Metab, July 1, 2008; 295(1): E216 - E222. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
A. S. Jeyapalan, R. A. Orellana, A. Suryawan, P. M. J. O'Connor, H. V. Nguyen, J. Escobar, J. W. Frank, and T. A. Davis Glucose stimulates protein synthesis in skeletal muscle of neonatal pigs through an AMPK- and mTOR-independent process Am J Physiol Endocrinol Metab, August 1, 2007; 293(2): E595 - E603. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 M. Z. Fan, L. I. Chiba, P. D. Matzat, X. Yang, Y. L. Yin, Y. Mine, and H. H. Stein Measuring synthesis rates of nitrogen-containing polymers by using stable isotope tracers J Anim Sci, April 1, 2006; 84(13_suppl): E79 - E. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
J. W. Frank, J. Escobar, A. Suryawan, H. V. Nguyen, S. R. Kimball, L. S. Jefferson, and T. A. Davis Dietary protein and lactose increase translation initiation factor activation and tissue protein synthesis in neonatal pigs Am J Physiol Endocrinol Metab, February 1, 2006; 290(2): E225 - E233. [Abstract] [Full Text] [PDF]
|
|
| |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
-form. St = feeding state: food-deprived or fed. Pr = protein level: 21, 33, or 45% protein.