Replacement of dietary carbohydrate with protein increases fat mass and reduces hepatic triglyceride synthesis and content in female obese Zucker rats.

Previous studies have demonstrated both energy restriction (ER) and higher protein (HP), lower carbohydrate (LC) diets downregulate hepatic de novo lipogenesis. Little is known about the independent and combined impact of ER and HP/LC diets on tissue-specific lipid kinetics in leptin receptor-deficient, obese rodents. This study investigated the effects of ER and dietary macronutrient content on body composition; hepatic, subcutaneous adipose tissue (SAT), and visceral AT (VAT) lipid metabolic flux (2 H2 O-labeling); and blood and liver measures of cardiometabolic health in six-week-old female obese Zucker rats (Leprfa+/fa+ ). Animals were randomized to a 10-week feeding intervention: ad libitum (AL)-HC/LP (76% carbohydrate/15% protein), AL-HP/LC (35% protein/56% carbohydrate), ER-HC/LP, or ER-HP/LC. ER groups consumed 60% of the feed consumed by AL. AL gained more fat mass than ER (P-energy = 0.012) and HP/LC gained more fat mass than HC/LP (P-diet = 0.025). Hepatic triglyceride (TG) concentrations (P-interaction = 0.0091) and absolute hepatic TG synthesis (P-interaction = 0.012) were lower in ER-HP/LC versus ER-HC/LP. ER had increased hepatic, SAT, and VAT de novo cholesterol fractional synthesis, absolute hepatic cholesterol synthesis, and serum cholesterol (P-energy≤0.0035). A HP/LC diet, independent of energy intake, led to greater gains in fat mass. A HP/LC diet, in the context of ER, led to reductions in absolute hepatic TG synthesis and TG content. However, ER worsened cholesterol metabolism. Increased adipose tissue TG retention with the HP/LC diet may reflect improved lipid storage capacity and be beneficial in this genetic model of obesity.

REDD1 deletion and treadmill running increase liver hepcidin and gluconeogenic enzymes in male mice.

The iron-regulatory hormone hepcidin is transcriptionally up-regulated by gluconeogenic signals. Recent evidence suggeststhat increases in circulating hepcidin may decrease dietary iron absorption following prolonged exercise, however evidence is limited on whether gluconeogenic signals contribute to post-exercise increases in hepcidin. Mice with genetic knockout of regulated in development and DNA response-1 (REDD1) display greater glycogen depletion following exercise, possibly indicating greater gluconeogenesis. The objective of the present study was to determine liver hepcidin, markers of gluconeogenesis and iron metabolism in REDD1 knockout and wild-type mice following prolonged exercise. Twelve-week-old male REDD1 knockout and wild-type mice were randomised to rest or 60 min treadmill running with 1, 3 or 6 h recovery (n = 5-8/genotype/group). Liver gene expression of hepcidin (Hamp) and gluconeogenic enzymes (Ppargc1a, Creb3l3, Pck1, Pygl) were determined by qRT-PCR. Effects of genotype, exercise and their interaction were assessed by two-way ANOVAs with Tukey's post-hoc tests, and Pearson correlations were used to assess the relationships between Hamp and study outcomes. Liver Hamp increased 1- and 4-fold at 3 and 6 h post-exercise, compared to rest (P-adjusted < 0⋅009 for all), and was 50% greater in REDD1 knockout compared to wild-type mice (P = 0⋅0015). Liver Ppargc1a, Creb3l3 and Pck1 increased with treadmill running (P < 0⋅0001 for all), and liver Ppargc1a, Pck1 and Pygl were greater with REDD1 deletion (P < 0⋅02 for all). Liver Hamp was positively correlated with liver Creb3l3 (R = 0⋅62, P < 0⋅0001) and Pck1 (R = 0⋅44, P = 0⋅0014). In conclusion, REDD1 deletion and prolonged treadmill running increased liver Hamp and gluconeogenic regulators of Hamp, suggesting gluconeogenic signalling of hepcidin with prolonged exercise.

Effects of energy balance on cognitive performance, risk-taking, ambulatory vigilance and mood during simulated military sustained operations (SUSOPS).

Sustained operations (SUSOPS) require military personnel to conduct combat and training operations while experiencing physical and cognitive stress and limited sleep. These operations are often conducted in a state of negative energy balance and are associated with degraded cognitive performance and mood. Whether maintaining energy balance can mitigate these declines is unclear. This randomized crossover study assessed the effects of energy balance on cognitive performance, risk-taking propensity, ambulatory vigilance, and mood during a simulated 72-h SUSOPS. METHODS: Ten male Soldiers (mean ± SE; 22.4 ± 1.7 y; body weight 87.3 ± 1.1 kg) completed two, 72-h simulated SUSOPS in random order, separated by 7 days of recovery. Each SUSOPS elicited ∼4500 kcal/d total energy expenditure and restricted sleep to 4 h/night. During SUSOPS, participants consumed either an energy-balanced or restricted diet that induced a 43 ± 3% energy deficit. A cognitive test battery was administered each morning and evening to assess: vigilance, working memory, grammatical reasoning, risk-taking propensity, and mood. Real-time ambulatory vigilance was assessed each morning, evening, and night via a wrist-worn monitoring device. RESULTS: Participants exhibited heightened risk-taking propensity (p = 0.047) with lower self-reported self-control (p = 0.021) and fatigue (p = 0.013) during energy deficit compared to during energy balance. Vigilance accuracy (p < 0.001) and working memory (p = 0.040) performance decreased, and vigilance lapses increased (p < 0.001) during SUSOPS, but did not differ by diet. Percentage of correct responses to ambulatory vigilance stimuli varied during SUSOPS (p = 0.019) independent of diet, with generally poorer performance during the morning and night. Total mood disturbance (p = 0.001), fatigue (p < 0.001), tension (p = 0.003), and confusion (p = 0.036) increased whereas vigor decreased (p < 0.001) during SUSOPS, independent of diet. CONCLUSION: Prolonged physical activity combined with sleep restriction is associated with impaired vigilance, memory, and mood state. Under such conditions, maintaining energy balance prevents increased risk-taking and improves self-control, but does not improve other aspects of cognitive function or mood. Given the small sample in the present study, replication in a larger cohort is warranted.

Iron-enriched Aspergillus oryzae as an alternative to iron sulphate to limit iron accumulation, growth and motility of the enteric pathogen S. Typhimurium.

Excess unabsorbed iron in the gastrointestinal tract may select for enteric pathogens and increase the incidence and severity of infectious disease. Aspergillus oryzae (Ao) is a filamentous fungus that has the ability to accumulate and store large amounts of iron, and when used as a supplement or fortificant, has similar absorption to ferrous sulphate (FeSO4) in humans. The objective of this study was to determine the effect of iron-enriched Ao (Ao iron) compared with FeSO4 on iron accumulation, growth and motility of the Gram-negative enteric pathogen, S. Typhimurium. S. Typhimurium was cultured in media containing no added iron or 1 µM elemental iron as either Ao iron or FeSO4. S. Typhimurium cultured with FeSO4 accumulated more iron than those cultured with Ao iron. Genes regulated by the iron-activated transcriptional repressor, Fur, did not differ between control and Ao iron, but decreased in S. Typhimurium cultured with FeSO4 compared with both groups. Growth of S. Typhimurium was greater when cultured with FeSO4 compared with Ao iron and control. S. Typhimurium swam faster, had greater acceleration and travelled further when cultured with FeSO4 compared with Ao iron and control; swim speed, acceleration and distance travelled did not differ between Ao iron and control. These findings provide evidence that Ao iron reduces the virulence of a common enteric pathogen in vitro. Further research is required to determine whether iron-enriched Ao is a suitable iron supplement to improve iron delivery in areas with a high infection burden.

Mild to Moderate Food Deprivation Increases Hepcidin and Results in Hypoferremia and Tissue Iron Sequestration in Mice.

BACKGROUND: Short-term starvation and severe food deprivation (FD) reduce dietary iron absorption and restricts iron to tissues, thereby limiting the amount of iron available for erythropoiesis. These effects may be mediated by increases in the iron regulatory hormone hepcidin; however, whether mild to moderate FD has similar effects on hepcidin and iron homeostasis is not known. OBJECTIVES: To determine the effects of varying magnitudes and durations of FD on hepcidin and indicators of iron status in male and female mice. METHODS: Male and female C57BL/6J mice (14 wk old; n = 170) were randomly assigned to consume AIN-93M diets ad libitum (AL) or varying magnitudes of FD (10%, 20%, 40%, 60%, 80%, or 100%). FD was based on the average amount of food consumed by the AL males or females, and food was split into morning and evening meals. Mice were euthanized at 48 h and 1, 2, and 3 wk, and hepcidin and indicators of iron status were measured. Data were analyzed by Pearson correlation and one-way ANOVA. RESULTS: Liver hepcidin mRNA was positively correlated with the magnitude of FD at all time points (P < 0.05). At 3 wk, liver hepcidin mRNA increased 3-fold with 10% and 20% FD compared with AL and was positively associated with serum hepcidin (R = 0.627, P < 0.0001). Serum iron was reduced by ∼65% (P ≤ 0.01), and liver nonheme iron concentrations were ∼75% greater (P ≤ 0.01) with 10% and 20% FD for 3 wk compared with AL. Liver hepcidin mRNA at 3 wk was positively correlated with liver Bmp6 (R = 0.765, P < 0.0001) and liver gluconeogenic enzymes (R = >0.667, P < 0.05) but not markers of inflammation (P > 0.05). CONCLUSIONS: FD increases hepcidin in male and female mice and results in hypoferremia and tissue iron sequestration. These findings suggest that increased hepcidin with FD may contribute to the disturbances in iron homeostasis with undernutrition.

Circulating and skeletal muscle microRNA profiles are more sensitive to sustained aerobic exercise than energy balance in males.

MicroRNAs (miRNAs) regulate molecular processes governing muscle metabolism. Physical activity and energy balance influence both muscle anabolism and substrate metabolism, but whether circulating and skeletal muscle miRNAs mediate those effects remains unknown. This study assessed the impact of sustained physical activity with participants in energy balance (BAL) or deficit (DEF) on circulating and skeletal muscle miRNAs. Using a randomized cross-over design, 10 recreational active healthy males (mean ± SD, 22 ± 5 years, 87 ± 11 kg) completed 72 h of high aerobic exercise-induced energy expenditures in BAL (689 ± 852 kcal/day) or DEF (-2047 ± 920 kcal/day). Blood and muscle samples were collected under rested/fasted conditions before (PRE) and immediately after 120 min load carriage exercise bout at the end (POST) of the 72 h. Trials were separated by 7 days. Circulating and skeletal muscle miRNAs were measured using microarray RT-qPCR. Independent of energy status, 36 circulating miRNAs decreased (P < 0.05), while 10 miRNAs increased and three miRNAs decreased in skeletal muscle (P < 0.05) at POST compared to PRE. Of these, miR-122-5p, miR-221-3p, miR-222-3p and miR-24-3p decreased in circulation and increased in skeletal muscle. Two circulating (miR-145-5p and miR-193a-5p) and four skeletal muscle (miR-21-5p, miR-372-3p, miR-34a-5p and miR-9-5p) miRNAs had time-by-treatment effects (P < 0.05). These data suggest that changes in miRNA profiles are more sensitive to increased physical activity compared to energy status, and that changes in circulating miRNAs in response to high levels of daily aerobic exercise are not reflective of changes in skeletal muscle miRNAs. KEY POINTS: Circulating and skeletal muscle miRNA profiles are more sensitive to high levels of aerobic exercise-induced energy expenditure compared to energy status. Changes in circulating miRNA in response to high levels of daily sustained aerobic exercise are not reflective of changes in skeletal muscle miRNA.

A Prolonged Bout of Running Increases Hepcidin and Decreases Dietary Iron Absorption in Trained Female and Male Runners.

BACKGROUND: Declines in iron status are frequently reported in those who regularly engage in strenuous physical activity. A possible reason is increases in the iron regulatory hormone hepcidin, which functions to inhibit dietary iron absorption and can be induced by the inflammatory cytokine interleukin-6 (IL-6). OBJECTIVES: The current study aimed to determine the impact of a prolonged bout of running on hepcidin and dietary iron absorption in trained female and male runners. METHODS: Trained female and male collegiate cross country runners (n = 28, age: 19.7 ± 1.2 y, maximal oxygen uptake: 66.1 ± 6.1 mL $\cdot$ kg -1$\cdot$ min-2, serum ferritin: 21.9 ± 13.3 ng/mL) performed a prolonged run (98.8 ± 14.7 min, 21.2 ± 3.8 km, 4.7 ± 0.3 min/km) during a team practice. Participants consumed a stable iron isotope with a standardized meal 2 h postrun and blood was collected 1 h later. The protocol was repeated 2 wk later except participants abstained from exercise (rest). RBCs were collected 15 d after exercise and rest to determine isotope enrichment. Differences between exercise and rest were assessed by paired t tests and Wilcoxon matched-pairs signed rank tests. Data are means ± SDs. RESULTS: Plasma hepcidin increased 51% after exercise (45.8 ± 34.4 ng/mL) compared with rest (30.3 ± 27.2 ng/mL, P = 0.0010). Fractional iron absorption was reduced by 36% after exercise (11.8 ± 14.6 %) compared with rest (18.5 ± 14.4 %, P = 0.025). Plasma IL-6 was greater after exercise (0.660 ± 0.354 pg/mL) than after rest (0.457 ± 0.212 pg/mL, P < 0.0001). Exploratory analyses revealed that the increase in hepcidin with exercise may be driven by a response in males but not females. CONCLUSIONS: A prolonged bout of running increases hepcidin and decreases dietary iron absorption compared with rest in trained runners with low iron stores. The current study supports that IL-6 contributes to the increase in hepcidin with prolonged physical activity, although future studies should explore potential sex differences in the hepcidin response.This trial was registered at Clinicaltrials.gov as NCT04079322.

Iron deficient diets modify the gut microbiome and reduce the severity of enteric infection in a mouse model of S. Typhimurium-induced enterocolitis.

Enteric infections are widespread in infants and children living in low-resource settings. Iron availability in the gastrointestinal tract may modify the gut microbiome and impact the incidence and severity of enteropathy. This study was designed to determine the effect of an iron-deplete compared to an iron-rich environment in the lower intestine on the gut microbiome, and whether iron availability in the lower intestine affects the host immune response and severity of enteric infection in young mice. Weanling C57BL/6 female mice were fed an iron deficient (Fe-, <6 ppm iron) or an iron fortified (Fe+, 300 ppm iron) diet for 6 weeks. Mice were pretreated with streptomycin prior to oral inoculation of Salmonella enterica subspecies enterica serovar Typhimurium to induce enteric infection (Sal+) or saline control (Sal-). Cecal iron concentrations were 55-fold greater with Fe+Sal- compared to Fe-Sal-. Microbiome sequencing revealed shifts in gut microbiota with dietary iron and enteric infection. There was ∼30% more S. Typhimurium in the cecum of Fe+Sal+ compared to Fe-Sal+. Plasma hepcidin increased with dietary iron and enteric infection, but was greatest in Fe+Sal+. Plasma lipocalin-2 and spleen size relative to bodyweight were greater in Fe+Sal+ compared to Fe+Sal-, Fe-Sal- and Fe-Sal+, and Fe+Sal+ lost more bodyweight compared to Fe-Sal+. Unabsorbed iron in the lower intestine modifies the gut microbiome and promotes a more severe enteropathy. These findings could suggest the need for alternative iron supplementation strategies in areas where enteric infection are common.

Slc39a4 in the small intestine predicts zinc absorption and utilization: a comprehensive analysis of zinc transporter expression in response to diets of varied zinc content in young mice.

Zinc homeostasis is primarily maintained by zinc transporters that regulate zinc uptake and efflux in the small intestine; however, the relative contribution of the many zinc transporters identified (Slc39a1-14, Slc30a1-10) to dietary zinc absorption and utilization remains unknown. The objective of this study was to determine the expression of Slc39a1-14 and Slc30a1-10 in the small intestine and their relative contribution to dietary zinc absorption in mice. Five-week-old male C57BL/6J mice were fed modified AIN-93G diets containing <1, 30, or 100ppm zinc (n=15 mice/diet). Following 1 week of feeding, mice were given an oral gavage containing 67Zn and liver and plasma isotope appearance was determined 6-h later by ICP-MS. Expression of Slc39a1-14 and Slc30a1-10 was determined in mucosa from duodenum, jejunum, and ileum. Plasma and liver total zinc concentrations were not different after one week of feeding (P>.05). Liver and plasma appearance of 67Zn was greater in mice fed <1ppm compared to the 30ppm (P<.0001) and 100ppm (P<.0001) zinc diets. With the exception of Slc39a2, Slc39a12, Slc30a3, and Slc30a8, the remaining zinc transporters were expressed across all diets and intestinal segments. Expression of Slc39a4, Slc39a11, and Slc30a6 changed with diet (Pdiet<.05 for all); expression of Slc39a5, Slc39a7, Slc39a11, Slc39a14, Slc30a1, Slc30a2, Slc30a4, Slc30a5, Slc30a7, and Slc30a10 changed by intestinal segment (Psegment<.05 for all). Slc39a4 was the only transporter positively associated with liver (r2=0.316, P<.001) and plasma (r2=0.189, P<.01) 67Zn appearance. Although most zinc transporters are expressed in the small intestine, intestinal Slc39a4 predicts fractional zinc absorption and utilization in young mice.

Divergent effects of sex and calcium/vitamin D supplementation on serum magnesium and markers of bone structure and function during initial military training.

Maintaining Mg status may be important for military recruits, a population that experiences high rates of stress fracture during initial military training (IMT). The objectives of this secondary analysis were to (1) compare dietary Mg intake and serum Mg in female and male recruits pre- and post-IMT, (2) determine whether serum Mg was related to parameters of bone health pre-IMT, and (3) whether Ca and vitamin D supplementation (Ca/vitamin D) during IMT modified serum Mg. Females (n 62) and males (n 51) consumed 2000 mg of Ca and 25 µg of vitamin D/d or placebo during IMT (12 weeks). Dietary Mg intakes were estimated using FFQ, serum Mg was assessed and peripheral quantitative computed tomography was performed on the tibia. Dietary Mg intakes for females and males pre-IMT were below the estimated average requirement and did not change with training. Serum Mg increased during IMT in females (0·06 ± 0·08 mmol/l) compared with males (-0·02 ± 0·10 mmol/l; P < 0·001) and in those consuming Ca/vitamin D (0·05 ± 0·09 mmol/l) compared with placebo (0·001 ± 0·11 mmol/l; P = 0·015). In females, serum Mg was associated with total bone mineral content (BMC, ß = 0·367, P = 0·004) and robustness (ß = 0·393, P = 0·006) at the distal 4 % site, stress-strain index of the polaris axis (ß = 0·334, P = 0·009) and robustness (ß = 0·420, P = 0·004) at the 14 % diaphyseal site, and BMC (ß = 0·309, P = 0·009) and stress-strain index of the polaris axis (ß = 0·314, P = 0·006) at the 66 % diaphyseal site pre-IMT. No significant relationships between serum Mg and bone measures were observed in males. Findings suggest that serum Mg may be modulated by Ca/vitamin D intake and may impact tibial bone health during training in female military recruits.

Energy Restriction Suppresses Muscle Protein Synthesis, and High Protein Diets Extend Protein Half-Lives Across the Muscle Proteome in Obese Female Zucker Rats.

BACKGROUND: Effects of high protein (HP) diets and prolonged energy restriction (ER) on integrated muscle protein kinetics have not been determined. OBJECTIVE: The objective of this study was to measure protein kinetics in response to prolonged ER and HP on muscle protein synthesis (MPS; absolute rates of synthesis) and muscle protein breakdown (MPB; half-lives) for proteins across the muscle proteome. METHODS: Female 6-wk-old obese Zucker rats (Leprfa+/fa+, n = 48) were randomly assigned to one of four diets for 10 wk: ad libitum-standard protein (AL-SP; 15% kcal from protein), AL-HP (35% kcal from protein), ER-SP, and ER-HP (both fed 60% feed consumed by AL-SP). During week 10, heavy/deuterated water (2H2O) was administered by intraperitoneal injection, and isotopic steady-state was maintained via 2H2O in drinking water. Rats were euthanized after 1 wk, and mixed-MPS as well as fractional replacement rate (FRR), relative concentrations, and half-lives of individual muscle proteins were quantified in the gastrocnemius. Data were analyzed using 2-factor (energy × protein) ANOVAs and 2-tailed t-tests or binomial tests as appropriate. RESULTS: Absolute MPS was lower in ER than AL for mixed-MPS (-29.6%; P < 0.001) and MPS of most proteins measured [23/26 myofibrillar, 48/60 cytoplasmic, and 46/60 mitochondrial (P < 0.05)], corresponding with lower gastrocnemius mass in ER compared with AL (-29.4%; P < 0.001). Although mixed-muscle protein half-life was not different between groups, prolonged half-lives were observed for most individual proteins in HP compared with SP in ER and AL (P < 0.001), corresponding with greater gastrocnemius mass in HP than SP (+5.3%; P = 0.043). CONCLUSIONS: ER decreased absolute bulk MPS and most individual MPS rates compared with AL, and HP prolonged half-lives of most proteins across the proteome. These data suggest that HP, independent of energy intake, may reduce MPB, and reductions in MPS may contribute to lower gastrocnemius mass during ER by reducing protein deposition in obese female Zucker rats.

Effects of energy balance on appetite and physiological mediators of appetite during strenuous physical activity: secondary analysis of a randomised crossover trial.

Energy deficit is common during prolonged periods of strenuous physical activity and limited sleep, but the extent to which appetite suppression contributes is unclear. The aim of this randomised crossover study was to determine the effects of energy balance on appetite and physiological mediators of appetite during a 72-h period of high physical activity energy expenditure (about 9·6 MJ/d (2300 kcal/d)) and limited sleep designed to simulate military operations (SUSOPS). Ten men consumed an energy-balanced diet while sedentary for 1 d (REST) followed by energy-balanced (BAL) and energy-deficient (DEF) controlled diets during SUSOPS. Appetite ratings, gastric emptying time (GET) and appetite-mediating hormone concentrations were measured. Energy balance was positive during BAL (18 (sd 20) %) and negative during DEF (-43 (sd 9) %). Relative to REST, hunger, desire to eat and prospective consumption ratings were all higher during DEF (26 (sd 40) %, 56 (sd 71) %, 28 (sd 34) %, respectively) and lower during BAL (-55 (sd 25) %, -52 (sd 27) %, -54 (sd 21) %, respectively; Pcondition < 0·05). Fullness ratings did not differ from REST during DEF, but were 65 (sd 61) % higher during BAL (Pcondition < 0·05). Regression analyses predicted hunger and prospective consumption would be reduced and fullness increased if energy balance was maintained during SUSOPS, and energy deficits of ≥25 % would be required to elicit increases in appetite. Between-condition differences in GET and appetite-mediating hormones identified slowed gastric emptying, increased anorexigenic hormone concentrations and decreased fasting acylated ghrelin concentrations as potential mechanisms of appetite suppression. Findings suggest that physiological responses that suppress appetite may deter energy balance from being achieved during prolonged periods of strenuous activity and limited sleep.

Sensitivity and reliability of zinc transporter and metallothionein gene expression in peripheral blood mononuclear cells as indicators of zinc status: responses to ex vivo zinc exposure and habitual zinc intake in humans.

Zn is an essential nutrient for humans; however, a sensitive biomarker to assess Zn status has not been identified. The objective of this study was to determine the reliability and sensitivity of Zn transporter and metallothionein (MT) genes in peripheral blood mononuclear cells (PBMCs) to Zn exposure ex vivo and to habitual Zn intake in human subjects. In study 1, human PBMCs were cultured for 24 h with 0-50 µm ZnSO4 with or without 5 µm N,N,N',N'-tetrakis(2-pyridylmethyl)ethylenediamine (TPEN), and mRNA expression of SLC30A1-10, SLC39A1-14, MT1 subtypes (A, B, E, F, G, H, L, M and X), MT2A, MT3 and MT4 mRNA was determined. In study 2, fifty-four healthy male and female volunteers (31·9 (sd 13·8) years, BMI 25·7 (sd 2·9) kg/m2) completed a FFQ, blood was collected, PBMCs were isolated and mRNA expression of selected Zn transporters and MT isoforms was determined. Study 1: MT1E, MT1F, MT1G, MT1H, MT1L, MT1M, MT1X, MT2A and SLC30A1 increased with increasing concentrations of Zn and declined with the addition of TPEN. Study 2: Average daily Zn intake was 16·0 (sd 5·3) mg/d (range: 9-31 mg/d), and plasma Zn concentrations were 15·5 (SD 2·8) µmol/l (range 11-23 µmol/l). PBMC MT2A was positively correlated with dietary Zn intake (r 0·306, P = 0·03) and total Zn intake (r 0·382, P < 0·01), whereas plasma Zn was not (P > 0·05 for both). Findings suggest that MT2A mRNA in PBMCs reflects dietary Zn intake in healthy adults and may be a component in determining Zn status.

Energy deficit increases hepcidin and exacerbates declines in dietary iron absorption following strenuous physical activity: a randomized-controlled cross-over trial.

BACKGROUND: Strenuous physical activity promotes inflammation and depletes muscle glycogen, which may increase the iron regulatory hormone hepcidin. Hepcidin reduces dietary iron absorption and may contribute to declines in iron status frequently observed following strenuous physical activity. OBJECTIVES: To determine the effects of strenuous physical activity on hepcidin and dietary iron absorption and whether energy deficit compared with energy balance modifies those effects. METHODS: This was a randomized, cross-over, controlled-feeding trial in healthy male subjects (n = 10, mean ± SD age: 22.4 ± 5.4 y, weight: 87.3 ± 10.9 kg) with sufficient iron status (serum ferritin 77.0 ± 36.7 ng/mL). Rest measurements were collected before participants began a 72-h simulated sustained military operation (SUSOPS), designed to elicit high energy expenditure, glycogen depletion, and inflammation, followed by a 7-d recovery period. Two 72-h SUSOPS trials were performed where participants were randomly assigned to consume either energy matched (±10%) to their individual estimated total daily energy expenditure (BAL) or energy at 45% of total daily energy expenditure to induce energy deficit (DEF). On the rest day and at the completion of BAL and DEF, participants consumed a beverage containing 3.8 mg of a stable iron isotope, and plasma isotope appearance was measured over 6 h. RESULTS: Muscle glycogen declined during DEF and was preserved during BAL (-188 ± 179 mmol/kg, P-adjusted < 0.01). Despite similar increases in interleukin-6, plasma hepcidin increased during DEF but not BAL, such that hepcidin was 108% greater during DEF compared with BAL (7.8 ± 12.2 ng/mL, P-adjusted < 0.0001). Peak plasma isotope appearance at 120 min was 74% lower with DEF (59 ± 38% change from 0 min) and 49% lower with BAL (117 ± 81%) compared with rest (230 ± 97%, P-adjusted < 0.01 for all comparisons). CONCLUSIONS: Strenuous physical activity decreases dietary iron absorption compared with rest. Energy deficit exacerbates both the hepcidin response to physical activity and declines in dietary iron absorption compared with energy balance. This trial was registered at clinicaltrials.gov as NCT03524690.

Randomized Trial Comparing Consumption of Military Rations to Usual Intake for 21 Consecutive Days: Nutrient Adequacy and Indicators of Health Status.

BACKGROUND: The US military Meal, Ready-to-Eat food ration is approved as a nutritionally adequate sole source of nutrition for ≤21 days. However, the ration continuously evolves, requiring periodic reassessment of its influence on nutritional status and health. OBJECTIVE: To determine the effects of consuming the US Armed Services Meal, Ready-to-Eat ration for 21 days, relative to usual diets, on nutrient intake, and indicators of nutritional status and cardiometabolic health. DESIGN: Parallel-arm, randomized, controlled trial, secondary analysis. PARTICIPANTS: Sixty healthy, weight stable, free-living adults from the Natick, MA, area participated between June 2015 and March 2017. INTERVENTION: Participants were randomized to consume their usual diet for 31days (CON), or a strictly controlled Meal, Ready-to-Eat-only diet for 21 days followed by their usual diet for 10 days (MRE). MAIN OUTCOME MEASURES: Nutrient intake (absolute and adjusted) throughout the study period, and indicators of nutrition status (vitamins B, D, folate, homocysteine, iron, magnesium, and zinc) and cardiometabolic health (glucose, insulin, and blood lipid levels) before (Day 0), during (Day 10 through Day 21), and after (Day 31) the intervention period. STATISTICAL ANALYSIS PERFORMED: Between-group differences over time were assessed using marginal models. Models for nutritional status and cardiometabolic health indicators were adjusted for age, initial body mass index, and baseline value of the dependent variable. RESULTS: Energy-adjusted fiber; polyunsaturated fatty acids; vitamins A, thiamin, riboflavin, B-6, C, D, and E; and magnesium and zinc intakes all increased in MRE during the intervention and were higher compared with CON (P<0.05), whereas relative protein intake decreased and was lower (P<0.05). Serum triglyceride concentrations averaged 19% (95% CI 0% to 41%) higher in MRE relative to CON during Days 10 to 31 (P=0.05). No statistically significant effects of diet on any other nutritional status or cardiometabolic health indicators were observed. CONCLUSIONS: Findings demonstrate that a Meal, Ready-to-Eat ration diet can provide a more micronutrient-dense diet than usual dietary intake aiding in maintenance of nutritional status over 21 days.

High-Altitude Acclimatization Suppresses Hepcidin Expression During Severe Energy Deficit.

Hennigar, Stephen R., Claire E. Berryman, Alyssa M. Kelley, Bradley J. Anderson, Andrew J. Young, James P. McClung, and Stefan M. Pasiakos. High-altitude acclimatization suppresses hepcidin expression during severe energy deficit. High Alt Med Biol. 21:232-236, 2020. Background: The erythropoietic cells in the bone marrow require iron to synthesize heme for incorporation into hemoglobin. Exposure to hypoxic conditions, such as extended sojourns to high altitude (HA), results in increased erythropoiesis and an increased physiological requirement for iron. In addition to increasing iron requirements, hypoxic conditions suppress appetite and often lead to decreased energy intake. The objective of this study was to determine the combined effects of severe energy deficit and hypoxia on hepcidin and measures of iron status in lowlanders sojourning to HA. Methods: Iron status indicators and hepcidin were determined in 17 healthy male volunteers (mean ± standard deviation, age 23 ± 6 years, body mass index 27 ± 4 kg/m2) fed a controlled diet (12 ± 1.2 mg iron/day) during a 20-day sojourn to 4300 m above sea level. Results: Chronic exposure to HA during severe energy deficit increased hematocrit by 12% (p < 0.01) and decreased serum hepcidin by 37% (p < 0.01) compared with baseline. Ferritin declined by 18% (p = 0.02) and transferrin saturation and soluble transferrin receptor increased by 55% and 83%, respectively (p < 0.01 for both) compared with baseline. Conclusions: HA acclimatization suppresses hepcidin expression to increase iron availability during severe energy deficit. Registered at ClinicalTrials.gov as NCT02731066.

Testosterone Administration During Energy Deficit Suppresses Hepcidin and Increases Iron Availability for Erythropoiesis.

CONTEXT: Severe energy deprivation markedly inhibits erythropoiesis by restricting iron availability for hemoglobin synthesis. OBJECTIVE: The objective of this study was to determine whether testosterone supplementation during energy deficit increased indicators of iron turnover and attenuated the decline in erythropoiesis compared to placebo. DESIGN: This was a 3-phase, randomized, double-blind, placebo-controlled trial. SETTING: The study was conducted at the Pennington Biomedical Research Center. PATIENTS OR OTHER PARTICIPANTS: Fifty healthy young males. INTERVENTION(S): Phase 1 was a 14-day free-living eucaloric controlled-feeding phase; phase 2 was a 28-day inpatient phase where participants were randomized to 200 mg testosterone enanthate/week or an isovolumetric placebo/week during an energy deficit of 55% of total daily energy expenditure; phase 3 was a 14-day free-living, ad libitum recovery period. MAIN OUTCOME MEASURE(S): Indices of erythropoiesis, iron status, and hepcidin and erythroferrone were determined. RESULTS: Hepcidin declined by 41%, indicators of iron turnover increased, and functional iron stores were reduced with testosterone administration during energy deficit compared to placebo. Testosterone administration during energy deficit increased circulating concentrations of erythropoietin and maintained erythropoiesis, as indicated by an attenuation in the decline in hemoglobin and hematocrit with placebo. Erythroferrone did not differ between groups, suggesting that the reduction in hepcidin with testosterone occurs through an erythroferrone-independent mechanism. CONCLUSION: These findings indicate that testosterone suppresses hepcidin, through either direct or indirect mechanisms, to increase iron turnover and maintain erythropoiesis during severe energy deficit. This trial was registered at www.clinicaltrials.gov as #NCT02734238.