Leucine or Protein Supplementation and Resistance Training
Condition: Skeletal Muscle
Study type: Interventional

Summary: 
      The purpose of this study was to determine the effects of L-leucine (LEU) or different
      protein supplements standardized to LEU (~3.0 g/serving) on changes in body composition,
      strength, and histological attributes in skeletal muscle and adipose tissue. Seventy-five
      untrained, college-aged males (mean±SE; age=21±1 yr, body mass=79.2±0.3 kg) were randomly
      assigned to an isocaloric, lipid-, and organoleptically-matched maltodextrin placebo (PLA,
      n=15), LEU (n=14), whey protein concentrate (WPC, n=17), whey protein hydrolysate (WPH,
      n=14), or soy protein concentrate (SPC, n=15) group. Participants performed whole-body
      resistance training three days per week for 12 weeks while consuming supplements twice daily.
      Skeletal muscle and subcutaneous (SQ) fat biopsies were obtained at baseline (T1) and ~72 h
      following the last day of training (T39). Tissue samples were analyzed for changes in type I
      and II fiber cross sectional area (CSA), non-fiber specific satellite cell count, and SQ
      adipocyte CSA. On average, all supplement groups including PLA exhibited similar training
      volumes and experienced statistically similar increases in total body skeletal muscle mass
      determined by dual x-ray absorptiometry (+2.2 kg; time p=0.024) and type I and II fiber CSA
      increases (+394 µm2 and +927 µm2; time p<0.001 and 0.024, respectively). Notably, all groups
      reported increasing Calorie intakes ~600-800 kcal/d from T1 to T39 (time p<0.001), and all
      groups consumed at least 1.1 g/kg/d of protein at T1 and 1.3 g/kg/d at T39. There was a
      training, but no supplementation, effect regarding the reduction in SQ adipocyte CSA (-210
      µm2; time p=0.001). Interestingly, satellite cell counts within the WPC (p<0.05) and WPH
      (p<0.05) groups were greater at T39 relative to T1. In summary, LEU or protein
      supplementation (standardized to LEU content) does not provide added benefit in increasing
      whole-body skeletal muscle mass or strength above PLA following 3 months of training in
      previously untrained college-aged males that increase Calorie intakes with resistance
      training and consume above the recommended daily intake of protein throughout training.
      However, whey protein supplementation increases skeletal muscle satellite cell number in this
      population, and this phenomena may promote more favorable training adaptations over more
      prolonged periods.
    

Detailed description: 
      OVERALL STUDY DESIGN The study design implemented was double-blinded and placebo-controlled.
      Likewise, guidelines established by the CONSORT Transparent Reporting of Trials established
      in 2010 (www.consort-statement.org) were followed. Participants were encouraged to refrain
      from rigorous physical activity for 4-5 days prior to baseline testing (T1). For T1,
      participants were instructed to report to the laboratory in a well-hydrated, 4-hour fasted
      state whereby they were subjected to the following assessments: a) urine specific gravity, b)
      height and body mass, c) body composition using dual-energy X-ray absorptiometry (DXA)
      (General Electric Lunar Prodigy enCORE, software version 10.50.086; Madison, WI, USA), d)
      vastus lateralis thickness using ultrasonography (General Electric LOGIQ S7 Expert; Chicago,
      IL, USA), e) venipuncture, f) percutaneous skeletal muscle biopsy collection from the vastus
      lateralis, and g) a percutaneous SQ fat biopsy from the gluteal region. Two to three days
      following T1, subjects reported back to the laboratory in a 4-hour fasted state for a second
      visit (T2) whereby maximal force production capacity was assessed using an isometric
      mid-thigh pull (IMTP) test, lower body strength was assessed using a three repetition maximum
      (3-RM) squat, and upper body strength was assessed using a 3-RM bench press. Additionally,
      during T2, subjects were familiarized with all lifts that were to be performed during the
      training intervention. Following T2, subjects engaged in 12 weeks of resistance training and
      supplementation. The last training bout (T38) consisted of IMTP as well as squat and bench
      press 3-RM re-assessments in a 4-hour fasted state. Seventy two hours following T38, subjects
      reported back to the laboratory in a 4-hour fasted state for post-testing (T39) which
      consisted of all body composition, and blood and biopsy collection procedures noted for T1.

      RESISTANCE TRAINING PROTOCOL For visits 3-37 (T3-T37), a daily undulating periodization
      training model was employed over the 12-week training period given that this model has been
      shown to be more beneficial in eliciting greater increases in strength and hypertrophy than
      traditional linear training models. Specifically, participants were instructed to perform
      free-weighted barbell squats, bench press, deadlifts, and bent-over-rows for 4 sets of 10
      repetitions (Monday or Tuesday), 6 sets of 4 repetitions (Wednesday or Thursday), and 5 sets
      of 6 repetitions (Friday or Sunday). Immediately following each completed set, a rating of
      perceived exertion score was acquired from each participant (scale: 1-10) in order to monitor
      and progress each participant accordingly while minimizing the potential risk of injury. The
      RPE scale was described to participants as the remaining number of repetitions that the
      participant would be able to complete while employing good technique (i.e., 1 = 9 remaining
      repetitions in reserve, 10 = 0 remaining repetitions in reserve). More information on
      relative training intensities and progression can be found in Table 1. Participants were
      instructed to attend all 36 resistance training sessions throughout the duration of the
      study, but those that missed more than 4 sessions were not included in the analysis due to
      lack of training compliance. All participants were supervised by laboratory personnel for
      each training session to ensure that proper lifting technique was executed, and training
      volumes for each session were recorded.

      SUPPLEMENTATION PROTOCOL As stated above, participants were assigned to ingest either a PLA,
      LEU, WPC, WPH, or SPC supplement throughout the training intervention. On training days
      (T3-T37), participants consumed an individually-packaged serving in ~500 ml of tap water
      immediately following each training session under direct observation of the study personnel.
      Additionally, participants were instructed to consume an individual serving within 30 min
      prior to bedtime. On non-training days, participants were instructed to consume an individual
      serving between a meal of their choosing and 30 min prior to bedtime. Supplements were
      separated into individual ready-made supplement-coded packets for daily consumption, and
      participants were given a 3-week supply. Study personnel collected and counted empty packets
      from each participant every 3 weeks before the next 3-week supply was distributed.
      Participants that did not consume >80% were not included in the analysis due to lack of
      compliance. Each supplement, except PLA, was formulated to provide ~3 g of leucine, per
      serving. Furthermore, each supplement was formulated to yield similar amounts of total energy
      (kcal) and fat (g), and was double-blinded to laboratory personnel and participants for
      group, appearance, taste, texture, and packaging. The WPC supplement was formulated using an
      agglomerated, 80% WPC (Hilmar™ 8010, Hilmar Ingredients; Hilmar, CA, USA). The WPH supplement
      was formulated using an agglomerated, partially hydrolyzed [12.5% degree of hydrolysate
      (12.5% DH), yielding approximately 67% of peptides as <5 kilodaltons (kDa) in molecular
      weight] 80% whey protein concentrate (Hilmar™ 8360, Hilmar Ingredients); SPC used an
      agglomerated, 80% soy protein concentrate (ALPHA® 5812, Solae, LLC; St. Louis, MO, USA); LEU
      used an agglomerated, L-Leucine (L-Leucine USP, Glambia Nutritionals; Carlsbad, CA, USA) and
      non-GMO, corn-derived maltodextrin (MALTRIN®-M100; Grain Processing Corporation; Muscantine,
      IA, USA); and, the PLA group was formulated using maltodextrin (MALTRIN®-M100; Grain
      Processing Corporation). All five supplements were manufactured at JW Nutritional, LLC
      (Allen, TX, USA), a United States Food and Drug Administration cGMP-compliant facility
      independently audited and pre-qualified by Obvium*Q, LLC (Phoenix, AZ, USA), a GMP regulatory
      compliance firm. Personnel at JW Nutritional, LLC and a third-party organization (Lockwood,
      LLC; Draper, UT, USA) formulated and maintained blinding of groups, and each supplement was
      assigned a randomly generated item number. Manufacturing batch records for production of each
      of the five supplements were reviewed by a trained, independent expert in dietary supplement
      quality control and assurance before approval for use within the present study. All
      supplements were independently validated for nutritional facts and total amino acids using
      validated, approved methods at Covance Laboratories, Inc. (Madison, WI, USA), a pre-qualified
      third-party analytical laboratory, and results reviewed by a third-party organization
      (Lockwood, LLC) prior to the supplements being approved for use within the present study.
      Once analysis was complete, a third-party organization (Lockwood, LLC) representative not
      involved in the study released the code for all treatments.
    

Intervention name(s): Placebo; L-leucine + maltodextrin; Whey protein concentrate; Hydrolyzed whey protein concentrate; Soy protein concentrate
Intervention type(s): Dietary Supplement; Dietary Supplement; Dietary Supplement; Dietary Supplement; Dietary Supplement
Intervention description(s): Supplementation twice daily (1 serving dose per consumption) of maltodextin while engaging in resistance training 3 times per week for a 12-week period. Supplementation occurred during training and non-training days.; Supplementation twice daily (1 serving dose per consumption) of L-leucine + maltodextin while engaging in resistance training 3 times per week for a 12-week period. Supplementation occurred during training and non-training days.; Supplementation twice daily (1 serving dose per consumption) of whey protein concentrate while engaging in resistance training 3 times per week for a 12-week period. Supplementation occurred during training and non-training days.; Supplementation twice daily (1 serving dose per consumption) of hydrolyzed whey protein concentrate while engaging in resistance training 3 times per week for a 12-week period. Supplementation occurred during training and non-training days.; Supplementation twice daily (1 serving dose per consumption) of soy protein concentrate while engaging in resistance training 3 times per week for a 12-week period. Supplementation occurred during training and non-training days.
Allocation: Randomized
Intervention model: Parallel Assignment
Masking: Double (Participant, Investigator)
Primary purpose: Treatment