D. J. Millward

The effect of protein deprivation and starvation on the rate of protein synthesis in tissues of the rat

1. The fractional rate of protein synthesis was measured in tissues of rats in vivo by continuous infusion of [14C]tyrosine. In growing animals proteins of liver and kidney were renewed at a rate greater than 50% per day, those in skeletal muscle, brain and heart at a rate between 13 and 23% per day. 2. Protein synthesis was also measured in liver, kidney, heart, brain and skeletal muscle of rats either given a protein-free diet for 21 days or starved for 2 days. During the first 2 days no clear differences between the effects of these two regimes could be detected. 3. Gastrocnemius muscle did not lose tissue protein till after 9 days without protein in the diet. The rate of protein synthesis was halved after 1 day and halved again after 21 days without protein. It was deduced that the rate of protein breakdown in muscle had declined also. 4. In liver the loss of protein was immediate without any apparent change in the fractional rate of protein synthesis. Between 2 and 21 days of dietary protein deprivation the liver lost protein slowly but the fractional rate of protein synthesis was increased. It is proposed that lack of protein in the diet also causes an increase in the rate of liver protein breakdown.

Growth and zinc homeostasis in the severely Zn-deficient rat

Male weanling rats were fed on diets either adequate (55 mg/kg), or severely deficient (0.4 mg/kg) in zinc, either ad lib. or in restricted amounts in four experiments. Measurements were made of growth rates and Zn contents of muscle and several individual tissues. Zn-deficient rats exhibited the expected symptoms of deficiency including growth retardation, cyclic changes in food intake and body-weight. Zn deficiency specifically reduced whole body and muscle growth rates as indicated by the fact that (a) growth rates were lower in ad lib.-fed Zn-deficient rats compared with rats pair-fed on the control diet in two experiments, (b) Zn supplementation increased body-weights of Zn-deficient rats given a restricted amount of diet at a level at which they maintained weight if unsupplemented, (c) Zn supplementation maintained body-weights of Zn-deficient rats fed a restricted amount of diet at a level at which they lost weight if unsupplemented (d) since the ratio, muscle mass: body-weight was lower in the Zn-deficient rats than in the pair-fed control groups, the reduction in muscle mass was greater than the reduction in body-weight. Zn concentrations were maintained in muscle, spleen and thymus, reduced in comparison to some but not all control groups in liver, kidney, testis and intestine, and markedly reduced in plasma and bone. In plasma, Zn concentrations varied inversely with the rate of change of body-weight during the cyclic changes in body-weight. Calculation of the total Zn in the tissues examined showed a marked increase in muscle Zn with a similar loss from bone, indicating that Zn can be redistributed from bone to allow the growth of other tissues. The magnitude of the increase in muscle Zn in the severely Zn-deficient rat, together with the magnitude of the total losses of muscle tissue during the catabolic phases of the cycling, indicate that in the Zn-deficient rat Zn may be highly conserved in catabolic states.

Dose response of protein turnover in rat skeletal muscle to triiodothyronine treatment

Skeletal muscle protein turnover has been examined in thyroidectomized rats treated with 0, 0.3, 0.75, 2, 20 and 100 micrograms triidothyronine/day for 7 days by implanted osmotic minipump. Protein synthesis in gastrocnemius, plantaris and soleus muscle were measured in vivo by the constant infusion method and protein degradation estimated as the difference between gross and net rates of synthesis. Serum levels of triidothyronine (T3) and insulin were also measured in addition to oxygen consumption rates in some cases. Compared with untreated intact rats muscle growth rates were unchanged at 0.3, 0.75 and 2 micrograms T3/day and, judging by plasma T3 levels, 0.75 microgram T3/day was a replacement dose. Slowing of growth was evident in the untreated thyroidectomized rats mid-way through the 7 day experimental period (6-7 days after throidectomy). High doses of T3 (20 and 100 micrograms/day) promptly supressed growth but there was subsequent recovery. Protein synthesis and degradation were generally lower in the hypothyroid state and normal or elevated in the hyperthyroid state. The changes in protein synthesis were mediated by changes in both RNA concentration and RNA activity (protein synthesis per unit RNA). Gastrocnemius and plantaris muscles were most responsive in the hypothyroid range. Since protein synthesis is particularly depressed in these muscles in malnutrition, the fall in protein degradation induced by the lowered thyroid status in this condition will be an important adaptive response to conserve protein. The increased protein turnover in the hyperthyroid rats was most marked in the soleus muscle and it is argued that this is necessary to allow the changes in protein composition and metabolic character which occur in response to hyperthyroidism in this muscle.

The effects of severe zinc deficiency on protein turnover in muscle and thymus.

Measurements have been made of protein turnover, RNA and DNA in thymus and skeletal muscle from rats fed on a zinc-deficient diet (ZD) for 10 and 17 d, in pair-fed controls (CI) and in muscle from rats fed on the ZD diet for 24 d and then fed on restricted amounts of the deficient diet with (RIZS) or without (RIZD) Zn supplementation, for 8 d. In thymus the ZD diet induced a loss of DNA and protein which was not observed with the CI rats. Accumulation of RNA was less affected but protein synthesis was reduced. In muscle the accumulation of DNA and protein was slowed by the ZD diet, particularly in glycolytic muscles compared with oxidative muscles, and Zn supplementation increased DNA and protein. Protein synthesis and RNA concentrations were reduced in the ZD rats compared with the CI rats, but Zn supplementation at constant restricted food intake did not increase protein synthesis. Muscle protein synthesis per unit RNA varied markedly in the ZD rats after 10 d when the characteristic cycling of the food intakes and body-weight was most pronounced, the highest values being observed in the anabolic phase of the cycle although these were less than values for well-fed controls. The variability was inversely correlated with the plasma Zn levels. The extent of the variability was much less after 17 d and was not apparent in the food-restricted ZD animals. Protein degradation in muscle, assessed as the difference between overall and net protein synthesis, was faster in the ZD rats compared with the CI rats and fluctuated considerably, partly accounting for the cyclic changes in muscle after 10 d, and was entirely responsible after 17 d. The concentration of muscle-free 3-methylhistidine and its urinary excretion rate indicated inconsistent results which could not be satisfactorily interpreted. Plasma insulin was reduced in the ZD rats compared with the CI rats and was insensitive to food intake in contrast to urinary corticosterone excretion which was inversely correlated with the cyclic changes in body-weight and food intake. Furthermore, adrenalectomized rats exhibited increased mortality and reduced cycling of body-weight and food intake. Thus Zn deficiency impairs growth by a combination of reduced food intake, a reduced anabolic response to food due to a reduced capacity for protein synthesis and reduced activation of protein synthesis, possibly reflecting impaired insulin secretion, and an increased catabolic response to the reduced intake in which corticosterone may play a role.

Whole-body protein and amino acid turnover in man: what can we measure with confidence?

Measurements of whole-body protein turnover are made either to assess the actual rate of one or more of the components of protein and amino acid turnover in a particular set of circumstances quantitatively, or to evaluate the qualitative responses to some administered stimulus (nutrient, hormone etc.). The achievement of either of these objectives still remains problematic despite considerable efforts to investigate and improve the methodology. Many of these problems are well known, having been reviewed on several occasions (Waterlow et al. 1978; Garlick & Clugston, 1981; Bier et al. 1985; Millward & Rivers, 1988). Since we are currently engaged in studies in which we have deployed several stable isotopic methods simultaneously, the results of these studies enable us to illustrate most of the major problems which are still outstanding and to address the question: What can we measure with confidence?

The effect of dietary protein and energy restriction on heat production and growth costs in the young rat.

1. The effect of dietary protein and energy restriction on heat production and growth costs has been examined in rats fed on a marginal (MP) or high (HP) protein diet, containing 9.2% or 22% respectively of the gross energy content as casein. Diets were given either ad lib. or at approximately 25, 50 or 75% of the ad lib. intake. 2. Heat production (kJ/kg body-weight (W)0.75 per d) was increased by 23% in rats fed on the MP diet ad lib., as compared with their HP controls (P less than 0.01). 3. Factorial analysis of the data showed that the overall cost of energy deposition (kJ/kJ; Ee) was elevated on the MP diet (MP 1.7, HP 1.28; P less than 0.001). Maintenance requirements (kJ/kg W0.75 per d) for zero energy balance were unchanged (MP 562, HP 573). 4. The partial energy cost of protein deposition (Ep) varied with dietary manipulation. If the partial energy cost of fat deposition (Ef) was assumed constant at 1.25 kJ/kJ, and maintenance requirements were assumed to vary with metabolic body size (W0.75), Ep was elevated on the MP diet. On both diets, Ep was reduced at low energy intakes. 5. The significance of these results is discussed in the context of current approaches to the analysis and interpretation of findings describing dietary induced changes in the rate of heat production.

FACTORS AFFECTING PROTEIN BREAKDOWN IN SKELETAL MUSCLE

Publisher Summary This chapter focuses on factors affecting the protein breakdown in skeletal muscle. Rates of protein breakdown have been determined in various skeletal muscles in the steady state and in anabolic and catabolic non-steady-state conditions. Protein breakdown was increased in muscle during anabolic and catabolic situations. The rate of protein breakdown in muscle fell when growth was interrupted by feeding a protein-free, low-protein, or low-energy diet, as well as in diabetic or hypophysectomized rats. Skeletal muscle is not a homogenous tissue. It has been known for some time that the rates of protein turnover are faster in cardiac and red tonic muscles than in white the phasic muscles. Although, the actual turnover rate of all three muscles tends to be faster in the fowl than the rat, perhaps reflecting the somewhat higher body temperature of the birds, the relative turnover rate of the PLD and heart is very similar to that of the white muscles and heart in the rat.

The effect of catabolic doses of corticosterone on heat production in the growing rat

The effect of corticosterone treatment on energy balance and heat production was investigated in growing rats. Animals were treated with daily subcutaneous injections of a vehicle containing 0, 50 or 100 mg corticosterone/kg for 5 d. Measurements of digestible energy intake and urinary energy losses showed that corticosterone treatment resulted in a depression of metabolizable energy intake due to elevated urinary energy losses resulting from massive glucosuria. Measurements of the metabolizable energy intake and the change in carcass energy indicated that at 50 mg/kg energy deposition and heat production were reduced, whilst at 100 mg/kg energy deposition was completely abolished with heat production increased. Postprandial oxygen consumption was unchanged at 50 mg/kg and increased at 100 mg/kg. Factorial analysis of these results based on reported values for the energy cost of protein and fat deposition indicated that (a) the depression of total heat production at 50 mg/kg could be entirely accounted for by the concomitant suppression of growth, and (b) the elevation of total and postprandial heat production at 100 mg/kg reflected a specific influence of corticosterone on thermogenesis. The significance of these findings is discussed in the light of reports that corticosterone in low doses suppresses heat production.

The Endocrine Response to Dietary Protein: the Anabolic Drive on Growth

In this review, I examine the endocrine response to dietary protein in the context of the anabolic drive on growth, our term for the regulatory influence of dietary protein on all aspects of the maintenance of optimal organ function and growth [21].

The nutritional sensitivity of the diurnal cycling of body protein enables protein deposition to be measured in subjects at nitrogen equilibrium

In adults in overall protein balance on intakes of protein equal to or greater than minimum requirements, the diurnal pattern of feeding and fasting results in cycling of body protein, with fasted losses and fed state gains of increasing amplitude with increasing habitual protein intake. Measurement of the slope of the fed state gain-intake relationship enables investigation of the ability of various patient groups to utilise dietary protein, without the need to impose negative nitrogen balance due to sub-maintenance protein intakes.