An exploration of the methods to determine the protein-specific synthesis and breakdown rates in vivo in humans.

Lars Holm,Rie H Nielsen,Kasper Dideriksen,Gerrit Hall,Grith Højfeldt,Simon Doessing,Marcus Moberg,Søren Reitelseder,Eva Blomstrand,Rasmus L Bechshoeft

doi:10.14814/phy2.14143

Abstract

The present study explores the methods to determine human in vivo protein‐specific myofibrillar and collagenous connective tissue protein fractional synthesis and breakdown rates. We found that in human myofibrillar proteins, the protein‐bound tracer disappearance method to determine the protein fractional breakdown rate (FBR) (via 2H2O ingestion, endogenous labeling of 2H‐alanine that is incorporated into proteins, and FBR quantified by its disappearance from these proteins) has a comparable intrasubject reproducibility (range: 0.09–53.5%) as the established direct‐essential amino acid, here L‐ring‐13C6‐phenylalanine, incorporation method to determine the muscle protein fractional synthesis rate (FSR) (range: 2.8–56.2%). Further, the determination of the protein breakdown in a protein structure with complex post‐translational processing and maturation, exemplified by human tendon tissue, was not achieved in this experimentation, but more investigation is encouraged to reveal the possibility. Finally, we found that muscle protein FBR measured with an essential amino acid tracer prelabeling is inappropriate presumably because of significant and prolonged intracellular recycling, which also may become a significant limitation for determination of the myofibrillar FSR when repeated infusion trials are completed in the same participants.

Highlights

The study of protein mass adaptation in whole body and limb, in structures such as skeletal muscle and tendons, and in various tissues and organs during health and disease requires knowledge of the dynamics of protein turnover (Garlick and Millward 1972; Millward et al 1975; Young1976; Matthews 1993)
We found that in human myofibrillar proteins, the protein-bound tracer disappearance method to determine the protein fractional breakdown rate (FBR) has a comparable intrasubject reproducibility as the established direct-essential amino acid, here L-ring-13C6-phenylalanine, incorporation method to determine the muscle protein fractional synthesis rate (FSR)
We found that muscle protein FBR measured with an essential amino acid tracer prelabeling is inappropriate presumably because of significant and prolonged intracellular recycling, which may become a significant limitation for determination of the myofibrillar FSR when repeated infusion trials are completed in the same participants

Summary

Introduction

The study of protein mass adaptation in whole body and limb, in structures such as skeletal muscle and tendons, and in various tissues and organs during health and disease requires knowledge of the dynamics of protein turnover (Garlick and Millward 1972; Millward et al 1975; Young1976; Matthews 1993). Valid measures of protein turnover involving both protein synthesis and breakdown rates are mandatory. These measures are possible with the use of stable isotopically labeled amino acid tracers. Using such tracers, many approaches exist and care must be taken with designing complex studies involving repeated tracer exposure in, for example, cross-over studies. Physiological Reports published by Wiley Periodicals, Inc. on behalf of The Physiological Society and the American Physiological Society Muscle biopsies were obtained from both legs, alternating within each trial, which we showed in Study 1 add some variability, though evenly across all trials

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