Casein Ingestion Does Not Increase Muscle Connective Tissue Protein Synthesis Rates

Abstract

Purpose: This study aimed to assess the effect of dietary protein ingestion on intramuscular connective tissue protein synthesis rates during overnight recovery from a single bout of resistance exercise.

Methods: Thirty-six healthy, young males were randomly assigned to one of three treatments. One group ingested 30 g intrinsically L-[1-C]-phenylalanine-labeled casein protein before sleep (PRO, n = 12). The other two groups performed a bout of resistance exercise in the evening and ingested either placebo (EX, n = 12) or 30 g intrinsically L-[1-C]-phenylalanine-labeled casein protein before sleep (EX + PRO, n = 12). Continuous intravenous infusions of L-[ring-H5]-phenylalanine and L-[1-C]-leucine were applied, and blood and muscle tissue samples were collected to assess connective tissue protein synthesis rates and dietary protein-derived amino acid incorporation in the connective tissue protein fraction.

Results: Resistance exercise resulted in higher connective tissue protein synthesis rates when compared with rest (0.086 ± 0.017%·h [EX] and 0.080 ± 0.019%·h [EX + PRO] vs 0.059 ± 0.016%·h [PRO]; P < 0.05). Postexercise casein protein ingestion did not result in higher connective tissue protein synthesis rates when compared with postexercise placebo ingestion (P = 1.00). Dietary protein-derived amino acids were incorporated into the connective tissue protein fraction at rest, and to a greater extent during recovery from exercise (P = 0.002).

Conclusion: Resistance exercise increases intramuscular connective tissue protein synthesis rates during overnight sleep, with no further effect of postexercise protein ingestion. However, dietary protein-derived amino acids are being used as precursors to support de novo connective tissue protein synthesis.

FIGURE 1

Experimental protocol. Thirty-six young male subjects performed a single bout of resistance exercise (EX and EX + PRO) or remained rested between 1945 and 2045 h. Subjects underwent an intravenous infusion of stable isotope amino acid tracers and ingested either 30 g intrinsically labeled casein protein (EX + PRO and PRO) or placebo (EX) at 2330 h. Skeletal muscle biopsies were collected before and after sleep to assess intramuscular connective tissue protein synthesis rates.

FIGURE 2

Overnight plasma total amino acid (A) and essential amino acid (B) concentrations (μmol·L⁻¹). The dotted line represents the ingestion of the presleep protein. Values represent mean ± SD. Data were analyzed with a two-way repeated-measures (within-subject factor: time; between-subject factor: treatment) ANOVA. Total amino acids: time effect, P < 0.001; treatment effect, P < 0.001; time–treatment interaction, P < 0.001. Essential amino acids: time effect, P < 0.001; treatment effect, P < 0.001; time–treatment interaction, P < 0.001. TAA, total amino acids; EAA, essential amino acids; PRO, presleep protein ingestion without prior exercise; EX, placebo ingestion with prior exercise; EX + PRO, protein ingestion with prior exercise.

FIGURE 3

Overnight plasma glycine concentrations (A), glycine iAUC (B), proline concentrations (C), and proline iAUC (D) (μmol·L⁻¹). The dotted line represents the ingestion of the presleep protein. Plasma concentrations over time are expressed as mean ± SD. Plasma iAUC is expressed as box and whisker plots with the median (line), mean (cross), interquartile range (box), and minimum and maximum values (tails), and treatments without a common letter differ, P < 0.05. Plasma concentrations over time were analyzed with a two-way repeated-measures (within-subject factor: time; between-subject factor: treatment) ANOVA. Plasma concentration iAUC was analyzed with a one-way (between-subject factor: treatment) ANOVA. Glycine concentrations: time effect, P < 0.001; treatment effect, P = 0.761; time–treatment interaction, P = 0.006. Glycine iAUC: main treatment effect, P = 0.008; EX vs PRO post hoc comparison, P = 0.006. Proline concentrations: time effect, P < 0.001; treatment effect, P < 0.001; time–treatment effect, P = 0.002. Proline iAUC: main treatment effect, P < 0.001; EX vs PRO and EX vs EX + PRO post hoc comparisons, P < 0.001. iAUC, incremental area under the curve; PRO, presleep protein ingestion without prior exercise; EX, placebo ingestion with prior exercise; EX + PRO, protein ingestion with prior exercise.

FIGURE 4

Overnight plasma L-[ring-²H₅]-phenylalanine (A), L-[1-¹³C]-leucine (B), and L-[1-¹³C]-phenylalanine (C) enrichments in MPE. The dotted line represents the ingestion of the presleep protein. Values represent mean ± SD. Data were analyzed with a two-way repeated-measures (within-subject factor: time; between-subject factor: treatment) ANOVA. L-[ring-²H₅]-phenylalanine enrichments: time effect, P < 0.001; treatment effect, P < 0.001; time–treatment effect, P < 0.001. L-[1-¹³C]-leucine enrichments: time effect, P < 0.001; treatment effect, P < 0.001; time–treatment effect, P < 0.001. L-[1-¹³C]-phenylalanine enrichments: time effect, P < 0.001; treatment effect, P < 0.001; time–treatment effect, P < 0.001. PRO, presleep protein ingestion without prior exercise; EX, placebo ingestion with prior exercise; EX + PRO, protein ingestion with prior exercise.

FIGURE 5

Overnight intramuscular connective tissue protein synthetic rates (FSR in %·h⁻¹) as calculated using L-[ring-²H₅]-phenylalanine or L-[1-¹³C]-leucine as tracer. The data are presented as box and whisker plots with the median (line), mean (cross), interquartile range (box), and minimum and maximum values (tails). Treatments without a common letter differ, P < 0.05. Data were analyzed with a one-way (between-subject factor: treatment) ANOVA. FSR based on L-[ring-²H₅]-phenylalanine; main treatment effect, P = 0.002. EX vs PRO and EX + PRO vs PRO post hoc comparisons, P < 0.005. FSR based on L-[1-¹³C]-leucine: main treatment effect, P = 0.166. PRO, presleep protein ingestion without prior exercise; EX, placebo ingestion with prior exercise; EX + PRO, protein ingestion with prior exercise.

FIGURE 6

Overnight L-[1-¹³C]-phenylalanine incorporation into intramuscular connective tissue protein in MPE. The data are presented as box and whisker plots with the median (line), mean (cross), interquartile range (box), and minimum and maximum values (tails). Treatments without a common letter differ, P < 0.05. The data were analyzed with an independent sample t-test, P = 0.002. PRO, presleep protein ingestion without prior exercise; EX + PRO, protein ingestion with prior exercise.