Petra Frings-Meuthen

Low-Grade Metabolic Acidosis May Be the Cause of Sodium Chloride-Induced Exaggerated Bone Resorption

Stepwise increase in NaCl intake in healthy male test subjects led to a low‐grade metabolic acidosis. This was most likely the cause for increased bone resorption during high sodium chloride intake, as determined by analyzing bone resorption markers.

High sodium chloride intake exacerbates immobilization-induced bone resorption and protein losses

We examined, in immobilization, the effect of a diet high in sodium chloride (NaCl) on bone markers, nitrogen balance, and acid-base status. Eight healthy male test subjects participated in a 14-day head-down-tilt bed rest (HDBR) study. During the bed rest period they received, in a randomized crossover design, a high (7.7 meq Na(+)/kg body wt per day) and a low (0.7 meq Na(+)/kg body wt per day) NaCl diet. As expected, 24-h excretion of urinary calcium was significantly greater in the high-NaCl-intake HDBR phase than in the low-NaCl-intake HDBR phase (P < 0.001). High NaCl intake caused a 43-50% greater excretion of the bone resorption markers COOH- (CTX) and NH(2)- (NTX) terminal telopeptide of type I collagen in HDBR than low NaCl in HDBR (CTX/NTX: P < 0.001). Serum concentrations of the bone formation markers bone-specific alkaline phosphatase (bAP) and NH(2)-terminal propeptide of type I procollagen (PINP) were identical in both NaCl intake phases. High NaCl intake led to a more negative nitrogen balance in HDBR (P < 0.001). Changes were accompanied by increased serum chloride concentration (P = 0.008), reduced blood bicarbonate (P = 0.017), and base excess (P = 0.009) whereas net acid excretion was lower during high than during low NaCl intake in immobilization (P < 0.001). High NaCl intake during immobilization exacerbates disuse-induced bone and muscle loss by causing further protein wasting and an increase in bone resorption. Changes in the acid-base status, mainly caused by disturbances in electrolyte metabolism, seem to determine NaCl-induced degradation processes.

Changes in intervertebral disc morphology persist 5 mo after 21-day bed rest

As part of the nutrition-countermeasures (NUC) study in Cologne, Germany in 2010, seven healthy male subjects underwent 21 days of head-down tilt bed rest and returned 153 days later to undergo a second bout of 21-day bed rest. As part of this model, we aimed to examine the recovery of the lumbar intervertebral discs and muscle cross-sectional area (CSA) after bed rest using magnetic resonance imaging and conduct a pilot study on the effects of bed rest in lumbar muscle activation, as measured by signal intensity changes in T(2)-weighted images after a standardized isometric spinal extension loading task. The changes in intervertebral disc volume, anterior and posterior disc height, and intervertebral length seen after bed rest did not return to prebed-rest values 153 days later. While recovery of muscle CSA occurred after bed rest, increases (P ≤ 0.016) in multifidus, psoas, and quadratus lumborum muscle CSA were seen 153 days after bed rest. A trend was seen for greater activation of the erector spinae and multifidus muscles in the standardized loading task after bed rest. Greater reductions of multifidus and psoas CSA muscle and greater increases in multifidus signal intensity with loading were associated with incidence of low back pain in the first 28 days after bed rest (P ≤ 0.044). The current study contributes to our understanding of the recovery of the lumbar spine after 21-day bed rest, and the main finding was that a decrease in spinal extensor muscle CSA recovers within 5 mo after bed rest but that changes in the intervertebral discs persist.

Lipocalin 2: A new mechanoresponding gene regulating bone homeostasis

Mechanical loading represents a crucial factor in the regulation of skeletal homeostasis. Its reduction causes loss of bone mass, eventually leading to osteoporosis. In a previous global transcriptome analysis performed in mouse calvarial osteoblasts subjected to simulated microgravity, the most upregulated gene compared to unit gravity condition was Lcn2, encoding the adipokine Lipocalin 2 (LCN2), whose function in bone metabolism is poorly known. To investigate the mechanoresponding properties of LCN2, we evaluated LCN2 levels in sera of healthy volunteers subjected to bed rest, and found a significant time‐dependent increase of this adipokine compared to time 0. We then evaluated the in vivo LCN2 regulation in mice subjected to experimentally‐induced mechanical unloading by (1) tail suspension, (2) muscle paralysis by botulin toxin A (Botox), or (3) genetically‐induced muscular dystrophy (MDX mice), and observed that Lcn2 expression was upregulated in the long bones of all of them, whereas physical exercise counteracted this increase. Mechanistically, in primary osteoblasts transfected with LCN2‐expression‐vector (OBs‐Lcn2) we observed that Runx2 and its downstream genes, Osterix and Alp, were transcriptionally downregulated, and alkaline phosphatase (ALP) activity was less prominent versus empty‐vector transduced osteoblasts (OBs‐empty). OBs‐Lcn2 also exhibited an increase of the Rankl/Opg ratio and IL‐6 mRNA, suggesting that LCN2 could link poor differentiation of osteoblasts to enhanced osteoclast stimulation. In fact, incubation of purified mouse bone marrow mononuclear cells with conditioned media from OBs‐Lcn2 cultures, or their coculture with OBs‐Lcn2, improved osteoclastogenesis compared to OBs‐empty, whereas treatment with recombinant LCN2 had no effect. In conclusion, our data indicate that LCN2 is a novel osteoblast mechanoresponding gene and that its regulation could be central to the pathological response of the bone tissue to low mechanical forces.

Alkaline Salts to Counteract Bone Resorption and Protein Wasting Induced by High Salt Intake: Results of a Randomized Controlled Trial

High sodium chloride (NaCl) intake can induce low-grade metabolic acidosis (LGMA) and may thus influence bone and protein metabolism. We hypothesized that oral potassium bicarbonate (KHCO(3)) supplementation may compensate for NaCl-induced, LGMA-associated bone resorption and protein losses. Eight healthy male subjects participated in a randomized trial with a crossover design. Each of two study campaigns consisted of 5 d of dietary and environmental adaptation followed by 10 d of intervention and 1.5 d of recovery. In one study campaign, 90 mmol KHCO(3)/d were supplemented to counteract NaCl-induced LGMA, whereas the other campaign served as a control with only high NaCl intake. When KHCO(3) was ingested during high NaCl intake, postprandial buffer capacity ([HCO(3)(-)]) increased (P = 0.002). Concomitantly, urinary excretion of free potentially bioactive glucocorticoids [urinary free cortisol (UFF) and urinary free cortisone (UFE)] was reduced by 14% [∑(UFF,UFE); P = 0.024]. Urinary excretion of calcium and bone resorption marker N-terminal telopeptide of type I collagen was reduced by 12 and 8%, respectively (calcium, P = 0.047; N-terminal bone collagen telopeptide, P = 0.044). There was a trend of declining net protein catabolism when high NaCl was combined with KHCO(3) (P = 0.052). We conclude that during high salt intake, the KHCO(3)-induced postprandial shift to a more alkaline state reduces metabolic stress. This leads to decreased bone resorption and protein degradation, which in turn might initiate an anticatabolic state for the musculoskeletal system in the long run.

21 Days head-down bed rest induces weakening of cell-mediated immunity – Some spaceflight findings confirmed in a ground-based analog

Several studies indicate a weakening of cell-mediated immunity (CMI) and reactivation of latent herpes viruses during spaceflight. We tested the hypothesis that head-down bed rest (HDBR), a ground-based analog of spaceflight, mimics the impact of microgravity on human immunity. Seven healthy young males underwent two periods of 3 weeks HDBR in the test facility of the German Aerospace Center. As a nutritional countermeasure aimed against bone demineralisation, 90 mmol potassium bicarbonate (KHCO(3)) was administered daily in a crossover design. Blood samples were drawn on five occasions. Whole blood was stimulated with antigen i.e. Candida albicans, purified protein derivative (PPD) tuberculin, tetanus toxoid and Cytomegalovirus (CMV) (CMV-QuantiFERON). Flow cytometric analysis included CD4(+)CD25(+)CD127(-)FOXP3(+) regulatory T cells (Tregs), γδ T cells, B cells, NK cells and dendritic cells. In one of the two bed rest periods, we observed a significant decrease in production of interleukin-2 (IL-2), interferon-γ (IFN-γ) and tumor necrosis factor-α (TNF-α) following phytohemagglutinin (PHA) stimulation, with a rapid normalization being observed after HDBR. The cytokine levels showed a V-shaped pattern that led to a relativeTh2-shift in cytokine balance. Only three individuals responded to the specific T cell antigens without showing signs of an altered response during HDBR, nor did we observe reactivation of CMV or Epstein-Barr virus (EBV). Of unknown significance, dietary supplementation with KHCO(3) counteracted the decrease in IL-2 levels during HDBR, while there was no impact on other immunological parameters. We conclude that discrete alterations in CMI may be induced by HDBR in selected individuals.

Elevated serum soluble CD200 and CD200R as surrogate markers of bone loss under bed rest conditions

CD200 is a transmembrane protein that belongs to the immunoglobulin family of proteins and is ubiquitously expressed on a variety of cell types. Upon interaction with its receptors (CD200Rs) expressed on myeloid-derived cells and T lymphocytes, an immunoregulatory signal is delivered to receptor-expressing cells. Previous studies have implicated a role for CD200:CD200R in the regulation of the expression of mRNA markers of osteoclastogenesis/osteoblastogenesis, following interaction of CD200 (on osteoblast precursors) with CD200R1 (on osteoclast precursors). Signaling of CD200R1 is hypothesized to attenuate osteoclastogenesis. We have investigated whether levels of soluble forms of CD200 and/or CD200R1 (sCD200, sCD200R1) are altered in volunteers undergoing 6° head down tilt bed rest to mimic conditions of microgravity known to be associated with preferential osteoclastogenesis and whether countermeasures, reported to be beneficial in attenuation of bone loss under microgravity conditions, would lead to altered sCD200 and sCD200R1 levels. Our data suggest that, as predicted, sCD200 levels fall under bed rest conditions while sCD200R1 levels rise. In subjects undergoing 30-minute per day continuous centrifugation protocols, as a countermeasure to attenuate changes which may lead to bone loss, these alterations in sCD200 and sCD200R1 levels seen under conditions of bed rest were abolished or attenuated. Our results suggest that measurement of sCD200 and/or sCD200R1 may prove a useful and rapid means of monitoring subjects at risk of bone loss and/or accessing the efficacy of treatment regimes designed to counter bone loss.

Short-term high dietary calcium intake during bedrest has no effect on markers of bone turnover in healthy men.

OBJECTIVE Immobilization and space flight are causes of disuse osteoporosis. Increasing calcium intake may counteract this disuse-induced bone loss. METHODS We conducted two bedrest experiments (crossover design: bedrest versus ambulatory control) in a metabolic ward, studying the effect of 1000 mg/d of calcium intake (study A, length of intervention 14 d) compared with that of a high calcium intake of 2000 mg/d (study B, 6 d) on markers of bone turnover. Both studies were randomized, controlled studies with the subjects staying under well-controlled environmental conditions (study A, 9 male subjects, age 23.6+/-3.0 y; study B, 8 male subjects, age 25.5+/-2.9 y). Blood was drawn to analyze serum calcium, parathyroid hormone, procollagen type I C-terminal propeptide, and bone alkaline phosphatase. Urine (24-h) was collected for analysis of calcium, C-terminal telopeptide of collagen type I, and N-terminal telopeptide of collagen type I. RESULTS In both studies, serum calcium levels remained unchanged. Procollagen type I C-terminal propeptide was lower (P=0.03) in the bedrest phase than in the ambulatory phase in study A and tended to be lower (P=0.08) in bedrest in study B, whereas bone alkaline phosphatase was not affected in either study. Urinary calcium excretion was greater during bedrest than during the ambulatory phase (study A, P=0.005; study B, P=0.002). C-terminal telopeptide of collagen type I excretion was also greater during bedrest in both studies (study A, P<0.001; study B, P<0.001). CONCLUSION Doubling calcium intake to 2000 mg/d does not prevent increased bone resorption induced by bedrest.

On the combined effects of normobaric hypoxia and bed rest upon bone and mineral metabolism: Results from the PlanHab study

Bone losses are common as a consequence of unloading and also in patients with chronic obstructive pulmonary disease (COPD). Although hypoxia has been implicated as an important factor to drive bone loss, its interaction with unloading remains unresolved. The objective therefore was to assess whether human bone loss caused by unloading could be aggravated by chronic hypoxia. In a cross-over designed study, 14 healthy young men underwent 21-day interventions of bed rest in normoxia (NBR), bed rest in hypoxia (HBR), and hypoxic ambulatory confinement (HAmb). Hypoxic conditions were equivalent to 4000m altitude. Bone metabolism (NTX, P1NP, sclerostin, DKK1) and phospho-calcic homeostasis (calcium and phosphate serum levels and urinary excretion, PTH) were assessed from regular blood samples and 24-hour urine collections, and tibia and femur bone mineral content was assessed by peripheral quantitative computed tomography (pQCT). Urinary NTX excretion increased (P<0.001) to a similar extent in NBR and HBR (P=0.69) and P1NP serum levels decreased (P=0.0035) with likewise no difference between NBR and HBR (P=0.88). Serum total calcium was increased during bed rest by 0.059 (day D05, SE 0.05mM) to 0.091mM (day D21, P<0.001), with no additional effect by hypoxia during bed rest (P=0.199). HAmb led, at least temporally, to increased total serum calcium, to reduced serum phosphate, and to reduced phosphate and calcium excretion. In conclusion, hypoxia did not aggravate bed rest-induced bone resorption, but led to changes in phospho-calcic homeostasis likely caused by hyperventilation. Whether hyperventilation could have mitigated the effects of hypoxia in this study remains to be established.

High-Intensity Jump Training Is Tolerated during 60 Days of Bed Rest and Is Very Effective in Preserving Leg Power and Lean Body Mass : An Overview of the Cologne RSL Study

Purpose Space agencies are looking for effective and efficient countermeasures for the degrading effects of weightlessness on the human body. The aim of this study was to assess the effects of a novel jump exercise countermeasure during bed rest on vitals, body mass, body composition, and jump performance. Methods 23 male participants (29±6 years, 181±6 cm, 77±7 kg) were confined to a bed rest facility for 90 days: a 15-day ambulatory measurement phase, a 60-day six-degree head-down-tilt bed rest phase (HDT), and a 15-day ambulatory recovery phase. Participants were randomly allocated to the jump training group (JUMP, n = 12) or the control group (CTRL, n = 11). A typical training session consisted of 4x10 countermovement jumps and 2x10 hops in a sledge jump system. The training group had to complete 5–6 sessions per week. Results Peak force for the reactive hops (3.6±0.4 kN) as well as jump height (35±4 cm) and peak power (3.1±0.2 kW) for the countermovement jumps could be maintained over the 60 days of HDT. Lean body mass decreased in CTRL but not in JUMP (-1.6±1.9 kg and 0±1.0 kg, respectively, interaction effect p = 0.03). Resting heart rate during recovery was significantly increased for CTRL but not for JUMP (interaction effect p<0.001). Conclusion Participants tolerated the near-daily high-intensity jump training and maintained high peak forces and high power output during 60 days of bed rest. The countermeasure was effective in preserving lean body mass and partly preventing cardiac deconditioning with only several minutes of training per day.