Pre-proenkephalin 1 is Downregulated Under Unloading and is Involved in Osteoblast Biology.

Pre-proenkephalin 1 (Penk1) is a pro-neuropeptide that belongs to the typical opioid peptide's family, having analgesic properties. We previously found Penk1 to be the most downregulated gene in a whole gene profiling analysis performed in osteoblasts subjected to microgravity as a model of mechanical unloading. In this work, Penk1 downregulation was confirmed in the bones of two in vivo models of mechanical unloading: tail-suspended and botulinum toxin A (botox)-injected mice. Consistently, in the sera from healthy volunteers subjected to bed rest, we observed an inverse correlation between PENK1 and bed rest duration. These results prompted us to investigate a role for this factor in bone. Penk1 was highly expressed in mouse bone, but its global deletion failed to impact bone metabolism in vivo. Indeed, Penk1 knock out (Penk1-/-) mice did not show an overt bone phenotype compared to the WT littermates. Conversely, in vitro Penk1 gene expression progressively increased during osteoblast differentiation and its transient silencing in mature osteoblasts by siRNAs upregulated the transcription of the Sost1 gene encoding sclerostin, and decreased Wnt3a and Col1a1 mRNAs, suggesting an altered osteoblast activity due to an impairment of the Wnt pathway. In line with this, osteoblasts treated with the Penk1 encoded peptide, Met-enkephalin, showed an increase of Osx and Col1a1 mRNAs and enhanced nodule mineralization. Interestingly, primary osteoblasts isolated from Penk1-/- mice showed lower metabolic activity, ALP activity, and nodule mineralization, as well as a lower number of CFU-F compared to osteoblasts isolated from WT mice, suggesting that, unlike the transient inhibition, the chronic Penk1 deletion affects both osteoblast differentiation and activity. Taken together, these results highlight a role for Penk1 in the regulation of the response of the bone to mechanical unloading, potentially acting on osteoblast differentiation and activity in a cell-autonomous manner.

Dietary Intake of Athletes at the World Masters Athletics Championships as Assessed by Single 24 h Recall.

Proper dietary intake is important for masters athletes because of the physiological changes that occur with aging and the unique nutritional needs when competing at high levels. We evaluated the dietary intake of masters athletes competing at the World Masters Athletics Championships (outdoor games, Tampere, Finland, 2022, and indoor games, Torun, Poland, 2023). A total of 43 athletes (16 females and 27 males, mean age 59.2 ± 10.3 y, height 168 ± 8 cm, and body mass 62.3 ± 10.8 kg) participating in endurance (n = 21), sprint (n = 16), jumping (2), multi-component (e.g., decathlon; n = 3), and throwing (n = 1) events provided 24 h dietary recalls while participating in the games. Carbohydrate intake was below the recommended levels for endurance athletes. Protein intake was below the recommended levels for masters athletes, except for female athletes involved in power events (i.e., sprinters and jumpers). Other nutrient intakes that were below the recommended levels included vitamins D and E, calcium, potassium, vitamin A (except for female endurance athletes), folate (except for female power athletes), vitamin C for female endurance athletes, vitamin K and fiber for males, and zinc for endurance athletes. We conclude that while competing at world championships, many athletes are not consuming the recommended levels of carbohydrates, protein, and micronutrients. Athletes attending these games would benefit from increased nutritional support.

The impact of bed rest on human skeletal muscle metabolism.

Insulin sensitivity and metabolic flexibility decrease in response to bed rest, but the temporal and causal adaptations in human skeletal muscle metabolism are not fully defined. Here, we use an integrative approach to assess human skeletal muscle metabolism during bed rest and provide a multi-system analysis of how skeletal muscle and the circulatory system adapt to short- and long-term bed rest (German Clinical Trials: DRKS00015677). We uncover that intracellular glycogen accumulation after short-term bed rest accompanies a rapid reduction in systemic insulin sensitivity and less GLUT4 localization at the muscle cell membrane, preventing further intracellular glycogen deposition after long-term bed rest. We provide evidence of a temporal link between the accumulation of intracellular triglycerides, lipotoxic ceramides, and sphingomyelins and an altered skeletal muscle mitochondrial structure and function after long-term bed rest. An intracellular nutrient overload therefore represents a crucial determinant for rapid skeletal muscle insulin insensitivity and mitochondrial alterations after prolonged bed rest.

Human gut microbiome and metabolite dynamics under simulated microgravity.

The Artificial Gravity Bed Rest - European Space Agency (AGBRESA) study was the first joint bed rest study by ESA, DLR, and NASA that examined the effect of simulated weightlessness on the human body and assessed the potential benefits of artificial gravity as a countermeasure in an analog of long-duration spaceflight. In this study, we investigated the impact of simulated microgravity on the gut microbiome of 12 participants during a 60-day head-down tilt bed rest at the :envihab facilities. Over 60 days of simulated microgravity resulted in a mild change in the gut microbiome, with distinct microbial patterns and pathway expression in the feces of the countermeasure group compared to the microgravity simulation-only group. Additionally, we found that the countermeasure protocols selectively increased the abundance of beneficial short-chain fatty acids in the gut, such as acetate, butyrate, and propionate. Some physiological signatures also included the modulation of taxa reported to be either beneficial or opportunistic, indicating a mild adaptation in the microbiome network balance. Our results suggest that monitoring the gut microbial catalog along with pathway clustering and metabolite profiling is an informative synergistic strategy to determine health disturbances and the outcome of countermeasure protocols for future space missions.

The future of spaceflight will involve missions beyond the International Space Station or the Moon and astronaut's health will be challenged by a harsh space environment for longer periods. In the last decade, the intestine has gained importance in dictating overall physiology and we explore it as an additional indicator of health during our ground-based bed rest study simulating microgravity for 60 days. Through the analysis of fecal proteins, we compile the catalog of microbes colonizing the gut of the 12 participants along with the implicated biological activity of the proteins and another 9 lipid analytes. We found specific microbes associated with recovery or healthy status in our subjects to be increased during spaceflight countermeasure conditions and inverse observations in subjects subjected to perilous spaceflight simulation. Our approach improves the functional characterization of the gut by the use of noninvasive methodology correlating the microbial composition of human stool samples with physiological status.

Oral fructose intake does not improve exercise, visual, or cognitive performance during acute normobaric hypoxia in healthy humans.

Introduction: The ability to metabolize fructose to bypass the glucose pathway in near-anaerobic conditions appears to contribute to the extreme hypoxia tolerance of the naked-mole rats. Therefore, we hypothesized that exogenous fructose could improve endurance capacity and cognitive performance in humans exposed to hypoxia. Methods: In a randomized, double-blind, crossover study, 26 healthy adults (9 women, 17 men; 28.8 ± 8.1 (SD) years) ingested 75 g fructose, 82.5 g glucose, or placebo during acute hypoxia exposure (13% oxygen in a normobaric hypoxia chamber, corresponding to oxygen partial pressure at altitude of ~3,800 m) on separate days. We measured exercise duration, heart rate, SpO2, blood gasses, and perceived exertion during a 30-min incremental load test followed by Farnsworth-Munsell 100 Hue (FM-100) color vision testing and the unstable tracking task (UTT) to probe eye-hand coordination performance. Results: Exercise duration in hypoxia was 21.13 ± 0.29 (SEM) min on fructose, 21.35 ± 0.29 min on glucose, and 21.35 ± 0.29 min on placebo (p = 0.86). Heart rate responses and perceived exertion did not differ between treatments. Total error score (TES) during the FM-100 was 47.1 ± 8.0 on fructose, 45.6 ± 7.6 on glucose and 53.3 ± 9.6 on placebo (p = 0.35) and root mean square error (RMSE) during the UTT was 15.1 ± 1.0, 15.1 ± 1.0 and 15.3 ± 0.9 (p = 0.87). Discussion: We conclude that oral fructose intake in non-acclimatized healthy humans does not acutely improve exercise performance and cognitive performance during moderate hypoxia. Thus, hypoxia tolerance in naked mole-rats resulting from oxygen-conserving fructose utilization, cannot be easily reproduced in humans.

Case Report: Muscle Wasting during Severe Sustained Hypoxia in Two Professional Mountaineers.

PURPOSE: Chronic exposure to hypoxia can induce muscle wasting in unaccustomed individuals. Detailed assessment of the effects of hypoxia on muscle tissue adaptation in elite mountaineers has not been performed. This study aims to assess muscle volume after exposure to normobaric hypoxia. METHODS: Two professional mountaineers (A and B) participated in a 35-d intervention of graded normobaric hypoxia with the aim of 14 d exposure to 8% oxygen corresponding to 7112-m altitude. Volume of the shank, thigh, and hip muscles was assessed by magnetic resonance imaging pre- and postintervention. Dietary intake and physical activity were monitored throughout the study from food images and accelerometry analysis, together with blood analysis and anthropometric measurements. RESULTS: Hypoxia reduced total leg muscle volume by 3.3% ± 6.0% in A and by 9.4% ± 7.3% in B. A lost 288 g and B 642 g of muscle mass, whereas dietary intake only declined by ~23% in the last intervention week. Arterial oxygen saturation declined from 95% and 86% to 77% and 72% in A and B, respectively. In hypoxia, participants could not maintain their physical activity levels. Notably, muscle loss varied substantially across muscle groups amounting to 5.4% ± 3.0%, 8.3% ± 5.2%, and 4.1% ± 8.6% for hip, thigh, and shank muscles, respectively. CONCLUSIONS: Our results indicate that hypoxia and resultant reductions in physical activity and caloric intake lead to substantial loss of muscle mass that was accentuated in proximal muscle as opposed to distal muscles. Surprisingly, thigh muscle wasting during this intervention is comparable with that observed during strict 56-d bed rest.

Bone metabolism during strict head-down tilt bed rest and exposure to elevated levels of ambient CO2.

Astronauts on the International Space Station are exposed to levels of atmospheric carbon dioxide (CO2) above typical terrestrial levels. We explored the possibility that increased levels of ambient CO2 further stimulate bone resorption during bed rest. We report here data from 2 ground-based spaceflight analog studies in which 12 male and 7 female subjects were placed in a strict 6° head-down tilt (HDT) position for either 30 days at 0.5% ambient CO2 or 60 days with nominal environmental exposure to CO2. Bone mineral density (BMD) and bone mineral content (BMC) were determined using dual-energy X-ray absorptiometry (DXA). Blood and urine were collected before and after HDT for biochemical analysis. No change was detected in either BMD or BMC, as expected given the study duration. Bone resorption markers increased after bed rest as expected; however, elevated CO2 had no additive effect. Elevated CO2 did not affect concentrations of minerals in serum and urine. Serum parathyroid hormone and 1,25-dihydroxyvitamin D were both reduced after bed rest, likely secondary to calcium efflux from bone. In summary, exposure to 0.5% CO2 for 30 days did not exacerbate the typical bone resorption response observed after HDT bed rest. Furthermore, results from these strict HDT studies were similar to data from previous bed rest studies, confirming that strict 30-60 days of HDT can be used to evaluate changes in bone metabolism. This is valuable in the continuing effort to develop and refine efficacious countermeasure protocols to mitigate bone loss during spaceflight in low-Earth orbit and beyond.

Effects of short-term hypercaloric nutrition on orthostatic tolerance in healthy individuals: a randomized controlled crossover study.

Reduced-caloric intake lowers blood pressure through sympathetic inhibition, and worsens orthostatic tolerance within days. Conversely, hypercaloric nutrition augments sympathetic activity and blood pressure. Because dietary interventions could be applied in patients with syncope, we tested the hypothesis that short-term hypercaloric dieting improves orthostatic tolerance. In a randomized crossover trial, 20 healthy individuals (7 women, 26.7 ± 8 years, 22.6 ± 2 kg/m2) followed a 4-day hypercaloric (25% increase of energy intake by fat) or normocaloric nutritional plan, with a washout period of at least 23 days between interventions. We then performed head-up tilt table testing with incremental lower body negative pressure while recording beat-by-beat blood pressure and heart rate. The primary endpoint was orthostatic tolerance defined as time to presyncope. Time to presyncope during combined head-up tilt and lower body negative pressure did not differ between hypercaloric and normocaloric dieting (median 23.19 versus 23.04 min, ratio of median 1.01, 95% CI of ratio 0.5-1.9). Heart rate, blood pressure, heart rate variability, and blood pressure variability in the supine position and during orthostatic testing did not differ between interventions. We conclude that 4 days of moderate hypercaloric nutrition does not significantly improve orthostatic tolerance in healthy individuals. Nevertheless, given the important interaction between energy balance and cardiovascular autonomic control in the brain, caloric intake deserves more attention as a potential contributor and treatment target for orthostatic intolerance.

Assessing the effects of artificial gravity in an analog of long-duration spaceflight: The protocol and implementation of the AGBRESA bed rest study.

A comprehensive strategy is required to mitigate risks to astronauts' health, well-being, and performance. This strategy includes developing countermeasures to prevent or reduce adverse responses to the stressors astronauts encounter during spaceflight, such as weightlessness. Because artificial gravity (AG) by centrifugation simultaneously affects all physiological systems, AG could mitigate the effects of weightlessness in multiple systems. In 2019, NASA and the German Aerospace Center conducted a 60-days Artificial Gravity Bed Rest Study with the European Space Agency (AGBRESA). The objectives of this study were to 1) determine if 30 min of AG daily is protective during head down bed rest, and 2) compare the protective effects of a single daily bout (30 min) of AG versus multiple daily bouts (6 × 5 min) of AG (1 Gz at the center of mass) on physiological functions that are affected by weightlessness and by head-down tilt bed rest. The AGBRESA study involved a comprehensive suite of standard and innovative technologies to characterize changes in a broad spectrum of physiological systems. The current article is intended to provide a detailed overview of the methods used during AGBRESA.

Iron metabolism regulation in females and males exposed to simulated microgravity: results from the randomized trial Artificial Gravity Bed Rest-European Space Agency (AGBRESA).

BACKGROUND: Iron metabolism imbalance could contribute to physical deconditioning experienced by astronauts due to its essential role in energy metabolism, cellular respiration, and oxygen transport. OBJECTIVES: In this clinical exploratory study, we wanted to determine whether artificial gravity (AG) training modulated iron metabolism, RBC indices, and body lean mass in healthy male and female participants exposed to head-down tilt (HDT) bed rest, the reference ground-based model of microgravity. METHODS: We recruited 8 healthy female and 16 healthy male participants who were all exposed to HDT bed rest for 60 d. In addition, they were assigned to 3 experimental groups (n = 8/each): controls, continuous AG training in a short-arm centrifuge (1 × 30 min/d), and intermittent AG training (6 × 5 min/d). RESULTS: The iron metabolism responses to simulated microgravity of the AG training groups did not differ significantly from the responses of controls. Independently from AG, we found that both serum iron concentrations (+31.3%, P = 0.027) and transferrin saturation levels (+28.4%, P = 0.009) increased in males after 6 d of HDT bed rest, as well as serum hepcidin concentrations (+36.9%, P = 0.005). The increase of transferrin saturation levels persisted after 57 d of HDT bed rest (+13.5%, P = 0.026), suggesting that long-term exposure to microgravity sustainably increases serum iron availability in males, and consequently the risk of iron excess or misdistribution. In females, 6 and 57 d of HDT bed rest did not significantly change serum iron, transferrin saturation, or hepcidin levels. CONCLUSIONS: The data from this exploratory study suggest that 1) AG training does not influence the iron metabolism responses to microgravity; and 2) iron metabolism parameters, especially iron availability for cells, are significantly increased in males, but not in females, exposed to long-term simulated microgravity. Because of the small sample size of females, we nevertheless must be cautious before concluding that iron metabolism could differently respond to microgravity in females. This trial was registered at https://www.drks.de as DRKS00015677.

Age-Related Decline in Vertical Jumping Performance in Masters Track and Field Athletes: Concomitant Influence of Body Composition.

Vertical jumping power declines with advancing age, which is theoretically explicable by loss of muscle mass and increases in body fat. However, the results of previous cross-sectional studies remain inconsistent on these relationships. The present study included 256 masters athletes who competed at the 2018 track and field world championships in Málaga, Spain. We assessed body composition with bioelectrical impedance (Inbody S10) and vertical jumping power with a Leonardo ground reaction force platform. Relationships between age, jumping power, and body composition were analyzed by correlation and regression analyses. Hierarchical multiple regression analysis was used to evaluate effects of each factor on vertical jumping power. Age-related rates of decreases in maximal power and jump height were similar between male and female athletes. Percent fat-free mass and percent body fat were negatively and positively, respectively, associated with age in masters athletes and were comparable to those previously observed in the general population. Moreover, these effects in body composition can, to a great extent, explain the age-related decline in jumping power, an effect that seems at least partly independent of age. Finally, the multiple regression model to determine independent predictors of vertical jump performance yielded an overall R 2 value of 0.75 with the inclusion of (1) athletic specialization in power events, (2) percent fat-free mass, and (3) phase angle. However, partial regression yielded significant effects of age, but not gender, on peak power, even when adjusting for athletic specialization, percent fat-free mass, and phase angle. We concluded that loss of skeletal muscle mass and changes in bio-impedance phase angle are important contributors to the age-related reduction in anaerobic power, even in adults who maintain high levels of physical activity into old age. However, age per se remains a significant predictor of vertical jump performance, further demonstrating deteriorated muscle quality at old age (sarcosthenia).

Resting Energy Expenditure of Master Athletes: Accuracy of Predictive Equations and Primary Determinants.

Resting energy expenditure (REE) is determined mainly by fat-free mass (FFM). FFM depends also on daily physical activity. REE normally decreases with increased age due to decreases in FFM and physical activity. Measuring REE is essential for estimating total energy expenditure. As such, there are a number of different equations in use to predict REE. In recent years, an increasing number of older adults continue to participate in competitive sports creating the surge of master athletes. It is currently unclear if these equations developed primarily for the general population are also valid for highly active, older master athletes. Therefore, we tested the validity of six commonly-used equations for predicting REE in master athletes. In conjunction with the World Masters Athletic Championship in Malaga, Spain, we measured REE in 113 master athletes by indirect calorimetry. The most commonly used equations to predict REE [Harris & Benedict (H&B), World Health Organization (WHO), Müller (MÜL), Müller-FFM (MÜL-FFM), Cunningham (CUN), and De Lorenzo (LOR)] were tested for their accuracies. The influences of age, sex, height, body weight, FFM, training hours per week, phase angle, ambient temperature, and athletic specialization on REE were determined. All estimated REEs for the general population differed significantly from the measured ones (H&B, WHO, MÜL, MÜL-FFM, CUN, all p < 0.005). The equation put forward by De Lorenzo provided the most accurate prediction of REE for master athletes, closely followed by FFM-based Cunningham's equation. The accuracy of the remaining commonly-used prediction equations to estimate REE in master athletes are less accurate. Body weight (p < 0.001), FFM (p < 0.001), FM (p = 0.007), sex (p = 0.045) and interestingly temperature (p = 0.004) are the significant predictors of REE. We conclude that REE in master athletes is primarily determined by body composition and ambient temperature. Our study provides a first estimate of energy requirements for master athletes in order to cover adequately athletes' energy and nutrient requirements to maintain their health status and physical performance.

Fetuin-A as a Potential Biomarker of Metabolic Variability Following 60 Days of Bed Rest.

Background: Fetuin-A is a hepatokine linked to the development of insulin resistance. The purpose of this study was to determine if 60 days head-down-tilt (HDT) bed rest increased circulating fetuin-A and if it was linked to whole body insulin sensitivity (IS). Additionally, we examined whether reactive jump training (RJT) could alleviate the metabolic changes associated with bed rest. Methods: 23 young men (29 ± 6 years, 181 ± 6 cm, 77 ± 7 kg) were randomized to a control (CTRL, n = 11) or RJT group (JUMP, n = 12) and exposed to 60 days of bed rest. Before and after bed rest, body composition and V . O 2 ⁢ p ⁢ e ⁢ a ⁢ k were measured and an oral glucose tolerance test was performed to estimate IS. Circulating lipids and fetuin-A were measured in fasting serum. Results: Body weight, lean mass, and V . O 2 ⁢ p ⁢ e ⁢ a ⁢ k decreased in both groups following bed rest, with greater reductions in CTRL (p < 0.05). There was a main effect of time, but not the RJT intervention, for the increase in fetuin-A, triglycerides (TG), area under the curve for glucose (AUCG) and insulin (AUCI), and the decrease in Matsuda and tissue-specific IS (p < 0.05). Fetuin-A increased in participants who became less insulin sensitive (p = 0.019). In this subgroup, liver IS and adipose IS decreased (p < 0.05), while muscle IS was unchanged. In a subgroup, where IS did not decrease, fetuin-A did not change. Liver IS increased (p = 0.012), while muscle and adipose tissue IS remained unchanged. Conclusions: In this study, we report an increase in circulating fetuin-A following 60 days of bed rest, concomitant with reduced IS, which could not be mitigated by RJT. The amount of fetuin-A released from the liver may be an important determinant of changes in whole body IS. In this regard, it may also be a useful biomarker of individual variation due to inactivity or lifestyle interventions.

The LunHab project: Muscle and bone alterations in male participants following a 10 day lunar habitat simulation.

NEW FINDINGS: What is the central question of this study? It is well established that muscle and bone atrophy in conditions of inactivity or unloading, but there is little information regarding the effect of a hypoxic environment on the time course of these deconditioning physiological systems. What is the main finding and its importance? The main finding is that a horizontal 10 day bed rest in normoxia results in typical muscle atrophy, which is not aggravated by hypoxia. Changes in bone mineral content or in metabolism were not detected after either normoxic or hypoxic bed rest. ABSTRACT: Musculoskeletal atrophy constitutes a typical adaptation to inactivity and unloading of weightbearing bones. The reduced-gravity environment in future Moon and Mars habitats is likely to be hypobaric hypoxic, and there is an urgent need to understand the effect of hypoxia on the process of inactivity-induced musculoskeletal atrophy. This was the principal aim of the present study. Eleven males participated in three 10 day interventions: (i) hypoxic ambulatory confinement; (ii) hypoxic bed rest; and (iii) normoxic bed rest. Before and after the interventions, the muscle strength (isometric maximal voluntary contraction), mass (lean mass, by dual-energy X-ray absorptiometry), cross-sectional area and total bone mineral content (determined with peripheral quantitative computed tomography) of the participants were measured. Blood and urine samples were collected before and on the 1st, 4th and 10th day of the intervention and analysed for biomarkers of bone resorption and formation. There was a significant reduction in thigh and lower leg muscle mass and volume after both normoxic and hypoxic bed rests. Muscle strength loss was proportionately greater than the loss in muscle mass for both thigh and lower leg. There was no indication of bone loss. Furthermore, the biomarkers of resorption and formation were not affected by any of the interventions. There was no significant effect of hypoxia on the musculoskeletal variables. Short-term normoxic (10 day) bed rest resulted in muscular deconditioning, but not in the loss of bone mineral content or changes in bone metabolism. Hypoxia did not modify these results.

Calcium Isotopes in Human Urine as a Diagnostic Tool for Bone Loss: Additional Evidence for Time Delays in Bone Response to Experimental Bed Rest.

The calcium (Ca) isotopic composition in urine during bed rest has been demonstrated to be systematically light, indicating a negative bone mineral balance (i.e., bone loss). Here we present new Ca isotope data on urine during the "nutritional countermeasures" (NUC) bed rest study. We analyzed the Ca isotopic composition of 24 h pooled urine samples from seven healthy male subjects during baseline data collection (BDC), head-down-tilt bed rest and recovery. Additionally, we analyzed urine from two follow-up examinations after the regeneration phase. We observed a change in Ca isotopic composition during the bed rest phase, indicative of bone loss with a time delay of 10 to 21 days. We also observe that the Ca isotopic composition of urine is strongly dependent on the individual Ca metabolism and varies between subjects. We relate this individuality in Ca metabolism to differences in the amounts of Ca being recycled in the kidneys. Previous studies have shown that the more Ca is reabsorbed in the kidneys the more enriched the urine becomes in heavy isotopes of calcium. The Ca isotopic composition of urine is thus modified by more than one process and cannot be used in a straightforward manner to monitor net bone mineral balance. To overcome this problem, we propose a new baseline approach for using Ca isotopes, which effectively cancels out the effects of individual renal Ca reabsorption. This allows us to detect bone loss in patients without ambiguity by combining measurements of the Ca isotopic composition of urine and daily Ca excretion rate and comparing these to data collected on healthy individuals with a normal steady-state bone balance.

Alkalinization with potassium bicarbonate improves glutathione status and protein kinetics in young volunteers during 21-day bed rest.

BACKGROUND & AIMS: Physical inactivity is associated with lean body mass wasting, oxidative stress and pro-inflammatory changes of cell membrane lipids. Alkalinization may potentially counteract these alterations. We evaluated the effects of potassium bicarbonate supplementation on protein kinetics, glutathione status and pro- and anti-inflammatory polyunsaturated fatty acids (PUFA) in erythrocyte membranes in humans, during experimental bed rest. METHODS: Healthy, young, male volunteers were investigated at the end of two 21-day bed rest periods, one with, and the other without, daily potassium bicarbonate supplementation (90 mmol × d-1), according to a cross-over design. Oxidative stress in erythrocytes was evaluated by determining the ratio between reduced (GSH) and oxidized glutathione (GSSG). Glutathione turnover and phenylalanine kinetics, a marker of whole body protein metabolism, were determined by stable isotope infusions. Erythrocyte membranes PUFA composition was analyzed by gas-chromatography. RESULTS: At the end of the two study periods, urinary pH was 10 ± 3% greater in subjects receiving potassium bicarbonate supplementation (7.23 ± 0.15 vs. 6.68 ± 0.11, p < 0.001). Alkalinization increased total glutathione concentrations by 5 ± 2% (p < 0.05) and decreased its rate of clearance by 38 ± 13% (p < 0.05), without significantly changing GSH-to-GSSG ratio. After alkalinization, net protein balance in the postabsorptive state improved significantly by 17 ± 5% (p < 0.05) as well as the sum of n-3 PUFA and the n-3-to-n-6 PUFA ratio in erythrocyte membranes (p < 0.05). CONCLUSIONS: Alkalinization during long-term inactivity is associated with improved glutathione status, anti-inflammatory lipid pattern in cell membranes and reduction in protein catabolism at whole body level. This study suggests that, in clinical conditions characterized by inactivity, oxidative stress and inflammation, alkalinization could be a useful adjuvant therapeutic strategy.

Short-Term Effects of Lupin vs. Whey Supplementation on Glucose and Insulin Responses to a Standardized Meal in a Randomized Cross-Over Trial.

Background: Whey protein is known to reduce postprandial glycaemia in people with type 2 diabetes mellitus. Lupin as a vegetable source of protein could be considered as an alternative, as the percentage of vegetarian and vegan consumers is raising. The present study compares the acute glycemic effects of whey and lupin in healthy volunteers following a carbohydrate-rich reference meal. Methods In cross-over design, three standardized meals (reference meal; reference meal + whey; reference meal + lupin) were provided to 12 healthy male and female volunteers, aged between 23 and 33, in a balanced, randomized order. Volunteers' blood glucose and insulin concentrations were analyzed at baseline and at seven time points following the ingestion of the meals. Results: The supplementation of whey or lupin significantly blunted the postprandial increase in blood glucose concentrations compared to the reference meal (p < 0.001). In the overall statistical analysis, this effect was comparable for whey and lupin [Δ AUC whey-lupin = 8%, 0-60 min area under the curve (0-60 min AUC), p = 0.937], with a blunting effect of -46% by whey (p = 0.005, 0-60 min AUC) and of -54% by lupin (p < 0.001, 0-60 min AUC). When comparing whey and lupin data only, the insulin increase was found to be more pronounced for whey protein than for lupin supplementation (Δ AUC whey-lupin = 39%, 0-60 min AUC, p = 0.022). However, when comparing the insulin response of each supplementation to the one of the reference meal, no differences could be detected (whey p = 0.259, 0-60 min AUC; lupin p = 0.275, 0-60 min AUC). Conclusions: Results suggest that lupin and whey can both lower the increase of postprandial blood glucose concentrations to a comparable extent, implying the usability of lupin to reduce postprandial glycaemia. However, the insulin response following the supplementations to a carbohydrate-rich meal seems to differ for these two protein sources.

Effects of high-protein intake on bone turnover in long-term bed rest in women.

Bed rest (BR) causes bone loss, even in otherwise healthy subjects. Several studies suggest that ambulatory subjects may benefit from high-protein intake to stimulate protein synthesis and to maintain muscle mass. However, increasing protein intake above the recommended daily intake without adequate calcium and potassium intake may increase bone resorption. We hypothesized that a regimen of high-protein intake (HiPROT), applied in an isocaloric manner during BR, with calcium and potassium intake meeting recommended values, would prevent any effect of BR on bone turnover. After a 20-day ambulatory adaptation to a controlled environment, 16 women participated in a 60-day, 6° head-down-tilt (HDT) BR and were assigned randomly to 1 of 2 groups. Control (CON) subjects (n = 8) received 1 g/(kg body mass·day)-1 dietary protein. HiPROT subjects (n = 8) received 1.45 g protein/(kg body mass·day)-1 plus an additional 0.72 g branched-chain amino acids per day during BR. All subjects received an individually tailored diet (before HDTBR: 1888 ± 98 kcal/day; during HDTBR: 1604 ± 125 kcal/day; after HDTBR: 1900 ± 262 kcal/day), with the CON group's diet being higher in fat and carbohydrate intake. High-protein intake exacerbated the BR-induced increase in bone resorption marker C-telopeptide (>30%) (p < 0.001) by the end of BR. Bone formation markers were unaffected by BR and high-protein intake. We conclude that high-protein intake in BR might increase bone loss. Further long-duration studies are mandatory to show how the positive effect of protein on muscle mass can be maintained without the risk of reducing bone mineral density.