Influence of Leucine Enriched Amino Acids on Myofibrillar and Collagen Protein Synthesis

Study Overview
=================
Brief Summary
-----------------
The primary objective will be to determine the effect of leucine-enriched essential amino acids (LEAA) compared to carbohydrate placebo on dietary incorporation of [D5] Phenylalanine & [D5] Glycine into the three skeletal muscle protein pools (myofibrillar, sarcoplasmic and collagen), both following resistance exercise and at rest, with the two tracers provided as a 'intrinsically labeled' bolus. Other outcomes will relate to molecular regulation of protein synthesis.

Official Title
-----------------
Influence of Leucine Enriched Amino Acids on Myofibrillar and Collagen Protein Synthesis in Human Skeletal Muscle

Conditions
-----------------
Interventional

Intervention / Treatment
-----------------
* Dietary Supplement: Leucine enriched amino acids
* Dietary Supplement: Carbohydrate Placebo


Participation Criteria
=================
Eligibility Criteria
-----------------
Inclusion Criteria: Healthy, male and female, recreationally-active participants. Healthy will be defined as screened by the PAR-Q+ (The Physical Activity Readiness Questionnaire for everyone; Appendix B). Self-reported not having engaged in resistance exercise and/or lower body plyometrics for at least 3 months prior to the study. Participants will be aged 18-35 years old. Participants are willing to abide by the compliance rules of this study. Self-reported regular menstrual cycle (25-35d) within the last 3 months (female participants). Exclusion Criteria: Inability to adhere to any of the compliance rules judged by principle investigator or medical doctor. Self-reported regular tobacco use. Self-reported illicit drug use (e.g. growth hormone, testosterone, etc.). Individuals who have participated in studies within the past year involving any of the stable isotopes in the study. Use of birth control and discontinued use within the last 3 months (female participants).

Ages Eligible for Study
-----------------
Minimum Age: 18 Years
Maximum Age: 35 Years

Sexes Eligible for Study
-----------------
All

Accepts Healthy Volunteers
-----------------
Accepts Healthy Volunteers

Study Plan
=================
How is the study designed?
-----------------

Design Details

Primary Purpose: Treatment
Allocation: Randomized
Intervention Model: Crossover Assignment
Interventional Model Description: Crossover to receive LEAA or Carbohydrate placebo. Unilateral leg exercise to give 4 outcome. (1) LEAA-EX; (2) LEAA-REST; (3) PLACEBO-EX; (4) PLACEBO-REST
Masking: Triple


Arms and Interventions

| Participant Group/Arm | Intervention/Treatment |
| --- | --- |
| Active Comparator: Leucine enriched essential amino acids<br>Leucine enriched essential amino acids (4g total, of which 1.6g are leucine). 20g carbohydrate, 2g of protein free drink powder, and 5g of Splenda to mask taste. | Dietary Supplement: Leucine enriched amino acids<br>* Leucine enriched essential amino acids (4g total, of which 1.6g are leucine). 20g carbohydrate, 2g of protein free drink powder, and 5g of Splenda.<br>* Other names: LEAA;|
| Placebo Comparator: Carbohdyrate Placebo<br>Iso-caloric placebo (4g maltodextrin to replace LEAA). 20g carbohydrate, 2g of protein free drink powder, and 5g of Splenda to mask taste. | Dietary Supplement: Carbohydrate Placebo<br>* iso-caloric placebo (4g maltodextrin). 20g carbohydrate, 2g of protein free drink powder, and 5g of Splenda.<br>|

What is the study measuring?
-----------------
Primary Outcome Measures

| Outcome Measure | Measure Description | Time Frame |
| --- | --- | --- |
| Myofibrillar Protein synthesis | Rate of incorporation of orally ingested amino acid tracers (D5 Phe and D5 Gly) into myofibrillar proteins isolated from skeletal muscle biopsies 4h following drink (LEAA or PLA) consumption and resistance exercise or at rest. Enrichment will be quantified via tandem LC-MS/MS to derive a rate of incorporation over the 4h period. | 4 hours | 

Secondary Outcome Measures

| Outcome Measure | Measure Description | Time Frame |
| --- | --- | --- |
| Sarcoplasmic protein synthesis | Rate of incorporation of orally ingested amino acid tracers (D5 Phe and D5 Gly) into sarcoplasmic (i.e., soluble) proteins isolated from skeletal muscle biopsies 4h following drink consumption (LEAA or PLA) and resistance exercise or at rest. Enrichment will be quantified via tandem LC-MS/MS. | 4 hours | 
| Collagen protein synthesis | Rate of incorporation of orally ingested amino acid tracers (D5 Phe and D5 Gly) into collagen (i.e., non-soluble) proteins isolated from skeletal muscle biopsies 4h following drink consumption (LEAA or PLA) and resistance exercise or at rest. Enrichment will be quantified via tandem LC-MS/MS. | 4 hours | 
| Protein localization via immunofluorescence | Antibody-determined Phosphorylated RPS6 (S240/244) and mTOR protein localization in human muscle cross sections and longitudinal sections as examined via immunofluorescence. Muscle sections (both longitudinal and cross-sections) will be derived from snap-frozen muscle biopsies performed 4h following drink consumption (LEAA or PLA) with or without resistance exercise (unilateral). Localization will be performed as done by Hodson et al., (2022; AJP Cell). Particularly, the localization of these targets relative to the cell periphery and z-discs (longitudinal sections only) will be analyzed via colocalization analysis. | 4 hours | 

 Terms related to the study
=================
Keywords Provided by Centre Hospitalier Valida
-----------------
Leucine-enriched amino acids, Carbohydrate Placebo