Effect of hyperinsulinaemia-hyperaminoacidaemia on leg muscle protein synthesis and breakdown: reassessment of the two-pool arterio-venous balance model

Abstract

Accurate measurement of muscle protein turnover is critical for understanding the physiological processes underlying muscle atrophy and hypertrophy. Several mathematical approaches, used in conjunction with a tracer amino acid infusion, have been described to derive protein synthesis and breakdown rates from a two-pool (artery-vein) model. Despite apparently common underlying principles, these approaches differ significantly (some seem to not take into account arterio-venous shunting of amino acids, which comprises ∼80-90% of amino acids appearing in the vein) and most do not specify how tracer enrichment (i.e. mole percent excess (MPE) or tracer-to-tracee ratio (TTR)) and amino acid concentration (i.e. unlabelled only or total labelled plus unlabelled) should be expressed, which could have a significant impact on the outcome when using stable isotope labelled tracers. We developed equations that avoid these uncertainties and used them to calculate leg phenylalanine (Phe) kinetics in subjects who received a [(2) H5 ]Phe tracer infusion during postabsorptive conditions and during a hyperinsulinaemic-euglycaemic clamp with concomitant protein ingestion. These results were compared with those obtained by analysing the same data with previously reported equations. Only some of them computed the results correctly when used with MPE as the enrichment measure and total (tracer+tracee) Phe concentrations; errors up to several-fold in magnitude were noted when the same approaches were used in conjunction with TTR and/or unlabelled concentration only, or when using the other approaches (irrespective of how concentration and enrichment are expressed). Our newly developed equations should facilitate accurate calculation of protein synthesis and breakdown rates.

© 2015 The Authors. The Journal of Physiology © 2015 The Physiological Society.

Figures

Figure 1

Two pool arterio-venous protein turnover model Central principles to this model are that: (i) Phe uptake by the leg reflects Phe used for protein synthesis, (ii), labelled and unlabelled Phe are both taken up by the limb and shunted from the artery to the vein, (iii) only unlabelled Phe is released by the leg, and (iv) Phe appearance into the vein (other than Phe shunted from the artery) is release from protein breakdown. Abbreviations: A, artery/arterial; B, breakdown; C, concentration (in μm); F, flux rate (in μmol min−1); L, labelled amino acid; PF, plasma flow rate (in l min−1); S, synthesis; T, total amino acid (i.e. sum of labelled and unlabelled amino acid); TTR, tracer-to-tracee ratio; U, unlabelled amino acid; V, vein/venous.

Figure 2

Leg phenylalanine kinetics during postabsorptive conditions (open bars) and during a hyperinsulinaemic–euglycaemic clamp procedure with concomitant whey protein ingestion (filled bars) Hatched bars represent the difference between these two conditions. Data are means ± SEM. *Value significantly different from postabsorptive value (P < 0.05).

Figure 3

Change (from postabsorptive conditions) in Phe utilization for protein synthesis (filled bars) and release from protein breakdown (open bars) during the hyperinsulinaemic–euglycaemic clamp procedure with concomitant whey protein ingestion calculated by using individual subjects’ data summarized in Table 1 and the newly proposed and six different previously published approaches Phe turnover rates for Barrett et al. (1987) were calculated two ways: (i) protein breakdown as initially presented in their methods section, and (ii) with an extraction ratio applied that corrects for the contribution from arterial inflow. Changes in the synthesis rates for Greenhaff et al. (2008) were calculated for the purposes of this figure as the difference between net balance and breakdown because the rates calculated using the equations presented in the original paper result in synthesis rates that are ∼5- to 10-fold different from those derived by any of the other methods and therefore appear to be attributable to errors in the reported formula. Data are percentage change from postpabsorptive values ± SEM of the change. Abbreviations: CT, total (i.e. tracer and tracee) concentration; CU, unlabelled (i.e. tracee) concentration; ER, extraction ratio; MPE, mole percent excess; TTR, tracer-to-tracee ratio.