Amino Acids

, Volume 37, Issue 2, pp 297–308 | Cite as

Acute and long-term effects of resistance exercise with or without protein ingestion on muscle hypertrophy and gene expression

  • Juha J. HulmiEmail author
  • Vuokko Kovanen
  • Harri Selänne
  • William J. Kraemer
  • Keijo Häkkinen
  • Antti A. Mero
Original Article


The effects of timed ingestion of high-quality protein before and after resistance exercise are not well known. In this study, young men were randomized to protein (n = 11), placebo (n = 10) and control (n = 10) groups. Muscle cross-sectional area by MRI and muscle forces were analyzed before and after 21 weeks of either heavy resistance training (RT) or control period. Muscle biopsies were taken before, and 1 and 48 h after 5 × 10 repetition leg press exercise (RE) as well as 21 weeks after RT. Protein (15 g of whey both before and after exercise) or non-energetic placebo were provided to subjects in the context of both single RE bout (acute responses) as well as each RE workout twice a week throughout the 21-week-RT. Protein intake increased (P ≤ 0.05) RT-induced muscle cross-sectional area enlargement and cell-cycle kinase cdk2 mRNA expression in the vastus lateralis muscle suggesting higher proliferating cell activation response with protein supplementation. Moreover, protein intake seemed to prevent 1 h post-RE decrease in myostatin and myogenin mRNA expression but did not affect activin receptor IIb, p21, FLRG, MAFbx or MyoD expression. In conclusion, protein intake close to resistance exercise workout may alter mRNA expression in a manner advantageous for muscle hypertrophy.


cdk2 Myostatin Activin receptor IIb Skeletal muscle Whey 



The authors thank Hanna Salmijärvi, Marja Katajavuori, Liisa Kiviluoto, Marko Haverinen, Mikko Pietikäinen, Hermanni Oksanen, Tuomas Kaasalainen, Tuovi Nykänen, Risto Puurtinen, and Aila Ollikainen for their help in data collection and analysis. We also thank the very dedicated group of subjects who made this project possible. The Finnish Ministry of Education and the Ellen and Artturi Nyyssönen Foundation (Juha Hulmi personal grant) supported this research.


  1. Adams GR, Caiozzo VJ, Haddad F, Baldwin KM (2002) Cellular and molecular responses to increased skeletal muscle loading after irradiation. Am J Physiol Cell Physiol 283:C1182–C1195PubMedGoogle Scholar
  2. Andersen LL, Tufekovic G, Zebis MK, Crameri RM, Verlaan G, Kjaer M, Suetta C, Magnusson P, Aagaard P (2005) The effect of resistance training combined with timed ingestion of protein on muscle fiber size and muscle strength. Metabolism 54:151–156PubMedCrossRefGoogle Scholar
  3. Berthet C, Aleem E, Coppola V, Tessarollo L, Kaldis P (2003) Cdk2 knockout mice are viable. Curr Biol 13:1775–1785PubMedCrossRefGoogle Scholar
  4. Bodine SC, Latres E, Baumhueter S, Lai VK, Nunez L, Clarke BA, Poueymirou WT, Panaro FJ, Na E, Dharmarajan K, Pan ZQ, Valenzuela DM, DeChiara TM, Stitt TN, Yancopoulos GD, Glass DJ (2001) Identification of ubiquitin ligases required for skeletal muscle atrophy. Science 294:1704–1708PubMedCrossRefGoogle Scholar
  5. Boirie Y, Dangin M, Gachon P, Vasson MP, Maubois JL, Beaufrere B (1997) Slow and fast dietary proteins differently modulate postprandial protein accretion. Proc Natl Acad Sci USA 94:14930–14935PubMedCrossRefGoogle Scholar
  6. Borsheim E, Tipton KD, Wolf SE, Wolfe RR (2002) Essential amino acids and muscle protein recovery from resistance exercise. Am J Physiol Endocrinol Metab 283:E648–E657PubMedGoogle Scholar
  7. Burke DG, Chilibeck PD, Davidson KS, Candow DG, Farthing J, Smith-Palmer T (2001) The effect of whey protein supplementation with and without creatine monohydrate combined with resistance training on lean tissue mass and muscle strength. Int J Sport Nutr Exerc Metab 11:349–364PubMedGoogle Scholar
  8. Candow DG, Burke NC, Smith-Palmer T, Burke DG (2006a) Effect of whey and soy protein supplementation combined with resistance training in young adults. Int J Sport Nutr Exerc Metab 16:233–244PubMedGoogle Scholar
  9. Candow DG, Chilibeck PD, Facci M, Abeysekara S, Zello GA (2006b) Protein supplementation before and after resistance training in older men. Eur J Appl Physiol 97:548–556PubMedCrossRefGoogle Scholar
  10. Charge SB, Rudnicki MA (2004) Cellular and molecular regulation of muscle regeneration. Physiol Rev 84:209–238PubMedCrossRefGoogle Scholar
  11. Cribb PJ, Hayes A (2006) Effects of supplement timing and resistance exercise on skeletal muscle hypertrophy. Med Sci Sports Exerc 38:1918–1925PubMedCrossRefGoogle Scholar
  12. Cribb PJ, Williams AD, Stathis CG, Carey MF, Hayes A (2007) Effects of whey isolate, creatine, and resistance training on muscle hypertrophy. Med Sci Sports Exerc 39:298–307PubMedCrossRefGoogle Scholar
  13. Durnin JV, Womersley J (1974) Body fat assessed from total body density and its estimation from skinfold thickness: measurements on 481 men and women aged from 16 to 72 years. Br J Nutr 32:77–97PubMedCrossRefGoogle Scholar
  14. Esmarck B, Andersen JL, Olsen S, Richter EA, Mizuno M, Kjaer M (2001) Timing of postexercise protein intake is important for muscle hypertrophy with resistance training in elderly humans. J Physiol 535:301–311PubMedCrossRefGoogle Scholar
  15. Gibala MJ, MacDougall JD, Tarnopolsky MA, Stauber WT, Elorriaga A (1995) Changes in human skeletal muscle ultrastructure and force production after acute resistance exercise. J Appl Physiol 78:702–708PubMedGoogle Scholar
  16. Guernec A, Chevalier B, Duclos MJ (2004) Nutrient supply enhances both IGF-I and MSTN mRNA levels in chicken skeletal muscle. Domest Anim Endocrinol 26:143–154PubMedCrossRefGoogle Scholar
  17. Ha E, Zemel MB (2003) Functional properties of whey, whey components, and essential amino acids: mechanisms underlying health benefits for active people (review). J Nutr Biochem 14:251–258PubMedCrossRefGoogle Scholar
  18. Halevy O, Nadel Y, Barak M, Rozenboim I, Sklan D (2003) Early posthatch feeding stimulates satellite cell proliferation and skeletal muscle growth in turkey poults. J Nutr 133:1376–1382PubMedGoogle Scholar
  19. Han B, Tong J, Zhu MJ, Ma C, Du M (2008) Insulin-like growth factor-1 (IGF-1) and leucine activate pig myogenic satellite cells through mammalian target of rapamycin (mTOR) pathway. Mol Reprod Dev 75:810–817PubMedCrossRefGoogle Scholar
  20. Hartman JW, Tang JE, Wilkinson SB, Tarnopolsky MA, Lawrence RL, Fullerton AV, Phillips SM (2007) Consumption of fat-free fluid milk after resistance exercise promotes greater lean mass accretion than does consumption of soy or carbohydrate in young, novice, male weightlifters. Am J Clin Nutr 86:373–381PubMedGoogle Scholar
  21. Hlaing M, Shen X, Dazin P, Bernstein HS (2002) The hypertrophic response in C2C12 myoblasts recruits the G1 cell cycle machinery. J Biol Chem 277:23794–23799PubMedCrossRefGoogle Scholar
  22. Hulmi JJ, Ahtiainen JP, Kaasalainen T, Pollanen E, Häkkinen K, Alen M, Selanne H, Kovanen V, Mero AA (2007) Postexercise myostatin and activin IIb mRNA levels: effects of strength training. Med Sci Sports Exerc 39:289–297PubMedCrossRefGoogle Scholar
  23. Hulmi JJ, Kovanen V, Lisko I, Selanne H, Mero AA (2008) The effects of whey protein on myostatin and cell cycle-related gene expression responses to a single heavy resistance exercise bout in trained older men. Eur J Appl Physiol 102:205–213PubMedCrossRefGoogle Scholar
  24. Häkkinen K, Pakarinen A, Newton RU, Kraemer WJ (1998) Acute hormone responses to heavy resistance lower and upper extremity exercise in young versus old men. Eur J Appl Physiol Occup Physiol 77:312–319PubMedCrossRefGoogle Scholar
  25. Häkkinen K, Pakarinen A, Kraemer WJ, Häkkinen A, Valkeinen H, Alen M (2001) Selective muscle hypertrophy, changes in EMG and force, and serum hormones during strength training in older women. J Appl Physiol 91:569–580PubMedGoogle Scholar
  26. Jeanplong F, Bass JJ, Smith HK, Kirk SP, Kambadur R, Sharma M, Oldham JM (2003) Prolonged underfeeding of sheep increases myostatin and myogenic regulatory factor myf-5 in skeletal muscle while IGF-I and myogenin are repressed. J Endocrinol 176:425–437PubMedCrossRefGoogle Scholar
  27. Kerksick CM, Rasmussen CJ, Lancaster SL, Magu B, Smith P, Melton C, Greenwood M, Almada AL, Earnest CP, Kreider RB (2006) The effects of protein and amino acid supplementation on performance and training adaptations during ten weeks of resistance training. J Strength Cond Res 20:643–653PubMedCrossRefGoogle Scholar
  28. Kim JS, Petrella JK, Cross JM, Bamman MM (2007) Load-mediated downregulation of myostatin mRNA is not sufficient to promote myofiber hypertrophy in humans: a cluster analysis. J Appl Physiol 103:1488–1495PubMedCrossRefGoogle Scholar
  29. Kraemer WJ, Adams K, Cafarelli E, Dudley GA, Dooly C, Feigenbaum MS, Fleck SJ, Franklin B, Fry AC, Hoffman JR, Newton RU, Potteiger J, Stone MH, Ratamess NA, Triplett-McBride T, American College of Sports Medicine (2002) American college of sports medicine position stand. Med Sci Sports Exerc 34:364–380PubMedCrossRefGoogle Scholar
  30. Kuang S, Charge SB, Seale P, Huh M, Rudnicki MA (2006) Distinct roles for Pax7 and Pax3 in adult regenerative myogenesis. J Cell Biol 172:103–113PubMedCrossRefGoogle Scholar
  31. Lee SJ, McPherron AC (2001) Regulation of myostatin activity and muscle growth. Proc Natl Acad Sci USA 98:9306–9311PubMedCrossRefGoogle Scholar
  32. Liu W, Saint DA (2002) Validation of a quantitative method for real time PCR kinetics. Biochem Biophys Res Commun 294:347–353PubMedCrossRefGoogle Scholar
  33. Malumbres M, Ortega S, Barbacid M (2000) Genetic analysis of mammalian cyclin-dependent kinases and their inhibitors. Biol Chem 381:827–838PubMedCrossRefGoogle Scholar
  34. Mascher H, Tannerstedt J, Brink-Elfegoun T, Ekblom B, Gustafsson T, Blomstrand E (2008) Repeated resistance exercise training induces different changes in mRNA expression of MAFbx and MuRF-1 in human skeletal muscle. Am J Physiol Endocrinol Metab 294:E43–E51PubMedCrossRefGoogle Scholar
  35. McCroskery S, Thomas M, Maxwell L, Sharma M, Kambadur R (2003) Myostatin negatively regulates satellite cell activation and self-renewal. J Cell Biol 162:1135–1147PubMedCrossRefGoogle Scholar
  36. McPherron AC, Lawler AM, Lee SJ (1997) Regulation of skeletal muscle mass in mice by a new TGF-beta superfamily member. Nature 387:83–90PubMedCrossRefGoogle Scholar
  37. Nagasawa T, Hirano J, Yoshizawa F, Nishizawa N (1998) Myofibrillar protein catabolism is rapidly suppressed following protein feeding. Biosci Biotechnol Biochem 62:1932–1937PubMedCrossRefGoogle Scholar
  38. Nakazato K, Hirose T, Song H (2006) Increased myostatin synthesis in rat gastrocnemius muscles under high-protein diet. Int J Sport Nutr Exerc Metab 16:153–165PubMedGoogle Scholar
  39. Ohkubo Y, Kishimoto T, Nakata T, Yasuda H, Endo T (1994) SV40 large T antigen reinduces the cell cycle in terminally differentiated myotubes through inducing Cdk2, Cdc2, and their partner cyclins. Exp Cell Res 214:270–278PubMedCrossRefGoogle Scholar
  40. Olsen S, Aagaard P, Kadi F, Tufekovic G, Verney J, Olesen JL, Suetta C, Kjaer M (2006) Creatine supplementation augments the increase in satellite cell and myonuclei number in human skeletal muscle induced by strength training. J Physiol 573:525–534PubMedCrossRefGoogle Scholar
  41. Paddon-Jones D, Sheffield-Moore M, Aarsland A, Wolfe RR, Ferrando AA (2005) Exogenous amino acids stimulate human muscle anabolism without interfering with the response to mixed meal ingestion. Am J Physiol Endocrinol Metab 288:E761–E767PubMedCrossRefGoogle Scholar
  42. Reeves ND, Maganaris CN, Narici MV (2004) Ultrasonographic assessment of human skeletal muscle size. Eur J Appl Physiol 91:116–118PubMedCrossRefGoogle Scholar
  43. Rios R, Carneiro I, Arce VM, Devesa J (2002) Myostatin is an inhibitor of myogenic differentiation. Am J Physiol Cell Physiol 282:C993–C999PubMedGoogle Scholar
  44. Sedliak M, Finni T, Cheng S, Haikarainen T, Häkkinen K (2007) Diurnal variation in maximal and submaximal strength, power and neural activation of leg extensors in men: multiple sampling across two consecutive days. Int J Sports Med 29(3):217–224PubMedCrossRefGoogle Scholar
  45. Thalacker-Mercer AE, Fleet JC, Craig BA, Carnell NS, Campbell WW (2007) Inadequate protein intake affects skeletal muscle transcript profiles in older humans. Am J Clin Nutr 85:1344–1352PubMedGoogle Scholar
  46. Tipton KD, Wolfe RR (2001) Exercise, protein metabolism, and muscle growth. Int J Sport Nutr Exerc Metab 11:109–132PubMedGoogle Scholar
  47. Tipton KD, Elliott TA, Cree MG, Aarsland AA, Sanford AP, Wolfe RR (2007) Stimulation of net muscle protein synthesis by whey protein ingestion before and after exercise. Am J Physiol Endocrinol Metab 292:E71–E76PubMedCrossRefGoogle Scholar
  48. Vissing K, Andersen JL, Schjerling P (2005) Are exercise-induced genes induced by exercise? FASEB J 19:94–96PubMedGoogle Scholar
  49. Wagner KR (2005) Muscle regeneration through myostatin inhibition. Curr Opin Rheumatol 17:720–724PubMedCrossRefGoogle Scholar
  50. Wernbom M, Augustsson J, Thomee R (2007) The influence of frequency, intensity, volume and mode of strength training on whole muscle cross-sectional area in humans. Sports Med 37:225–264PubMedCrossRefGoogle Scholar
  51. Wilkinson SB, Tarnopolsky MA, Macdonald MJ, Macdonald JR, Armstrong D, Phillips SM (2007) Consumption of fluid skim milk promotes greater muscle protein accretion after resistance exercise than does consumption of an isonitrogenous and isoenergetic soy-protein beverage. Am J Clin Nutr 85:1031–1040PubMedGoogle Scholar

Copyright information

© Springer-Verlag 2008

Authors and Affiliations

  • Juha J. Hulmi
    • 1
    Email author
  • Vuokko Kovanen
    • 2
    • 3
  • Harri Selänne
    • 4
  • William J. Kraemer
    • 5
  • Keijo Häkkinen
    • 1
  • Antti A. Mero
    • 1
  1. 1.Department of Biology of Physical ActivityUniversity of JyväskyläJyväskyläFinland
  2. 2.Department of Health SciencesUniversity of JyväskyläJyväskyläFinland
  3. 3.Finnish Centre for Interdisciplinary Gerontology (FCIG)University of JyväskyläJyväskyläFinland
  4. 4.LIKES Research CenterJyväskyläFinland
  5. 5.Human Performance Laboratory, Department of KinesiologyUniversity of ConnecticutStorrsUSA

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