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Vitamin D Status and Muscle Function in Post-Menarchal Adolescent Girls
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The Journal of Clinical Endocrinology & Metabolism Feb 2009 Vol. 94, No. 2 559-563
Kate A. Ward, Geeta Das, Jacqueline L. Berry, Stephen A. Roberts, Rainer Rawer, Judith E. Adams and Zulf Mughal
Imaging Science and Biomedical Engineering (K.A.W., J.E.A.), and Health Methodology Research Group (S.A.R.), University of Manchester, Manchester M13 9PT, United Kingdom; Central Manchester Primary Care Trust (G.D.), Longsight Health Centre, Longsight, Manchester M13 0RR, United Kingdom; Vitamin D Research Group (J.L.B.), Medicine, University of Manchester, Manchester Royal Infirmary, Manchester M13 9WL, United Kingdom; Novotec Medical GmbH (R.R.), D-75172 Pforzheim, Germany; and Department of Paediatric Medicine (Z.M.), Saint Mary's Hospital for Women & Children, Central Manchester & Manchester Children's Hospitals National Health Service Trust, Manchester M13 0JH, United Kingdom
"In conclusion, vitamin D status is significantly associated with muscle power and force. These data highlight the importance of vitamin D status on muscle function in adolescent girls. Suboptimal force might have implications for long-term bone development. The long-term implications of these observations require further study.....
....From these data we conclude that vitamin D was significantly associated with muscle power and force in adolescent girls......Our data demonstrate that in a group of asymptomatic post-menarchal adolescents, serum 25(OH)D was positively related to muscle power, force, velocity, and jump height; PTH had a lesser effect upon muscle parameters. We have also confirmed the observations that there is an interdependence of muscle function (force and power) with anthropometric parameters; in our data this was predominantly weight (13, 17). Therefore, these data suggest that muscle contractility is affected by the girl's vitamin D status, those with low-serum 25(OH)D concentration generated less power, and so jump height and velocity were lower than those with higher concentrations of 25(OH)D. In this study the majority of girls had an EFI of less than 100% (median 88%, range 36-143%)......PTH status may also contribute to myopathy associated with vitamin D deficiency; patients with primary hyperparathyroidism often complain of muscle fatigue and weakness...
.....We used JM to assesses the maximal force developed in seconds; this type of force is generated through anaerobic metabolism by the type II fibers that are those responsible for fast anaerobic contraction (26) and those affected by vitamin D deficiency (27). We cannot directly ascertain how 25(OH)D status affects muscle function in our population. The mechanism is likely to be multifactorial due to genomic (protein synthesis) and nongenomic (Ca and P transport) effects of 25(OH)D's active metabolite, 1,25 dihydroxy vitamin D, which interacts with the vitamin D receptor. In a RCT of vitamin D supplementation, El-Hajj Fuleihan et al. (28) observed an increase in whole body lean mass (a surrogate measure of muscle mass) in pre-menarchal girls who received the supplement. Thus, it is plausible that the better muscle function that we have observed is mediated through the effect of vitamin D on muscle mass. Finally, the effects on muscle might be mediated through hypophosphatemia, which occurs in vitamin D deficiency due to the secondary hyperparathyroidism (29). However, we did not observe any relationships between serum phosphorous concentrations and the parameters measured by JM....
....The main limitation of the current study is that the data are cross-sectional, and, therefore, we cannot establish the temporal nature of any of the relationships described. We cannot exclude any effect of body composition (BMI or less likely height) from these data, or whether the effects are due to a direct effect of 25(OH)D, or its metabolites, on muscle. However, after adjusting for both BMI and height in the model, the effect sizes differed very little (data not shown), thus, suggesting some effect of 25(OH)D on muscle (30, 31). Finally, we did not measure 1,25 dihydroxy vitamin D, the biologically active form of vitamin D, but others have reported that 25(OH)D is appropriate for the study of muscle (8)."
Abstract
Context: There has been a resurgence of vitamin D deficiency among infants, toddlers, and adolescents in the United Kingdom. Myopathy is an important clinical symptom of vitamin D deficiency, yet it has not been widely studied.
Objective: Our objective was to investigate the relationship of baseline serum 25 hydroxyvitamin D [25(OH)D] concentration and PTH with muscle power and force.
Design: This was a cross-sectional study.
Setting: The study was community based in a secondary school.
Participants: A total of 99 post-menarchal 12- to 14-yr-old females was included in the study.
Main Outcome Measures: Jumping mechanography to measure muscle power, velocity, jump height, and Esslinger Fitness Index from a two-legged counter movement jump and force from multiple one-legged hops was performed. Body height, weight, and serum concentrations of 25(OH)D, PTH, and calcium were measured.
Results: Median serum 25(OH)D concentration was 21.3 nmol/liter (range 2.5-88.5) and PTH 3.7 pmol/liter (range 0.47-26.2). After correction for weight using a quadratic function, there was a positive relationship between 25(OH)D and jump velocity (P = 0.002), jump height (P = 0.005), power (P = 0.003), Esslinger Fitness Index (P = 0.003), and force (P = 0.05). There was a negative effect of PTH upon jump velocity (P = 0.04).
Conclusion: From these data we conclude that vitamin D was significantly associated with muscle power and force in adolescent girls.
Introduction
There has been a resurgence of vitamin D deficiency among infants, toddlers, and adolescents in the United Kingdom (1, 2, 3, 4, 5, 6). Extremely low levels of 25 hydroxyvitamin D [25(OH)D] (measurable parameter of an individual's vitamin D status) in adolescents cause nonspecific limb pain, lower limb and pelvic deformities, tetany, and convulsions (4). In addition to these symptoms, vitamin D deficiency can cause myopathy (4, 7), which tends to be more marked in proximal muscles (8). Myopathy associated with vitamin D deficiency has been less well studied (9). In children, myopathy can lead to nonparticipation in physical education or organized sports. Muscles play a crucial role in bone development and maintenance of their integrity (10, 11). Thus, chronic vitamin D deficiency might influence skeletal development and mineralization though promotion of gastrointestinal absorption of calcium (Ca) and phosphorous, and ensuring optimal muscle action on the skeleton.
Jumping mechanography (JM) is designed to measure muscle force and power by deriving measurements from an individual's ground reaction forces (12); the method has only recently been introduced as a tool to assess muscle function, and has been validated in children, athletes, and the frail elderly (13, 14, 15). The proximal muscles required to jump (quadriceps, gastrocnemius, soleus) are those most often affected in subjects with vitamin D deficiency (8), and, therefore, JM provides a way of assessing this. Compared with the more common functional techniques used to assess proximal muscle function such as chair rising and timed up and go, JM had the smallest short-term error, is least influenced by learning effect, and had the largest interindividual variation (14, 15). Using a dynamic measure such as JM might be more functionally relevant than static measures such as isometric or dynamic isokinetic tests (15).
Therefore, the aim of the current study was to use JM to study the effects of vitamin D status, as reflected by serum 25(OH)D concentrations on muscle function. Because vitamin D deficiency leads to secondary hyperparathyroidism, the effect of serum PTH on muscle function was also studied. The data used in this study were collected during the screening phase of a double-blind randomized controlled trial (RCT) of vitamin D supplementation in a group of girls at risk for vitamin D deficiency. The specific pretrial hypotheses were that baseline serum 25(OH)D concentration would be positively, and PTH concentration negatively, related to muscle power and force measured using JM.
Discussion
We have used a novel outcome measure of JM to investigate how skeletal muscle function in the lower limb is affected by vitamin D and PTH status. Our data demonstrate that in a group of asymptomatic post-menarchal adolescents, serum 25(OH)D was positively related to muscle power, force, velocity, and jump height; PTH had a lesser effect upon muscle parameters. We have also confirmed the observations that there is an interdependence of muscle function (force and power) with anthropometric parameters; in our data this was predominantly weight (13, 17). Therefore, these data suggest that muscle contractility is affected by the girl's vitamin D status, those with low-serum 25(OH)D concentration generated less power, and so jump height and velocity were lower than those with higher concentrations of 25(OH)D. In this study the majority of girls had an EFI of less than 100% (median 88%, range 36-143%).
The majority of girls in our study have force less than 3 N/kg body weight. Under normal circumstances, the maximum force generated when jumping is approximately three to 3.5 times an individual's body weight (20). Given that the forces generated by muscles drive bone development, our data suggest that the bones are not being maximally loaded, which might affect the development of peak bone strength. We recently showed that South Asians, who should have attained peak bone strength, had less mineral content and thinner cortices than Europeans; this could, in part, be due to the lack of optimization of muscle force-generating capacity during childhood (21).
PTH status may also contribute to myopathy associated with vitamin D deficiency; patients with primary hyperparathyroidism often complain of muscle fatigue and weakness (22), and show improved muscle strength after surgery (22, 23). Furthermore, an association between high-serum PTH levels and sarcopenia had been reported (24). However, Glerup et al. (8) showed only small associations between muscle strength and PTH concentration in individuals with poor vitamin D status. In our population, PTH seems to have little effect upon muscle function; there is a negative relationship with velocity of jump, but no other parameters.
In the current study, none of the girls had symptoms of low 25(OH)D status, yet we have shown the effects of 25(OH)D upon muscle function. We previously reported that the Gowers' sign (25), used to detect proximal muscle myopathy in children, was not related to 25(OH)D status (3). In other words, jumping appears to be detecting a subclinical effect of 25(OH)D on muscle function, suggesting that it is more sensitive than the Gowers' sign; this concurs with previous observations in the elderly (14). There has been a resurgence of vitamin D deficiency in South Asian and Middle Eastern girls living in the United Kingdom. Of our screened population attending this school, 75% have low 25(OH)D levels (<15 ng/ml); we had specifically targeted this asymptomatic population to see whether we could detect a subclinical effect of 25(OH)D on muscle function in these girls. In a more 25(OH)D replete population, the association may differ; we tested for a threshold effect (data not shown) in our population, but none was detected.
We used JM to assesses the maximal force developed in seconds; this type of force is generated through anaerobic metabolism by the type II fibers that are those responsible for fast anaerobic contraction (26) and those affected by vitamin D deficiency (27). We cannot directly ascertain how 25(OH)D status affects muscle function in our population. The mechanism is likely to be multifactorial due to genomic (protein synthesis) and nongenomic (Ca and P transport) effects of 25(OH)D's active metabolite, 1,25 dihydroxy vitamin D, which interacts with the vitamin D receptor. In a RCT of vitamin D supplementation, El-Hajj Fuleihan et al. (28) observed an increase in whole body lean mass (a surrogate measure of muscle mass) in pre-menarchal girls who received the supplement. Thus, it is plausible that the better muscle function that we have observed is mediated through the effect of vitamin D on muscle mass. Finally, the effects on muscle might be mediated through hypophosphatemia, which occurs in vitamin D deficiency due to the secondary hyperparathyroidism (29). However, we did not observe any relationships between serum phosphorous concentrations and the parameters measured by JM.
The main limitation of the current study is that the data are cross-sectional, and, therefore, we cannot establish the temporal nature of any of the relationships described. We cannot exclude any effect of body composition (BMI or less likely height) from these data, or whether the effects are due to a direct effect of 25(OH)D, or its metabolites, on muscle. However, after adjusting for both BMI and height in the model, the effect sizes differed very little (data not shown), thus, suggesting some effect of 25(OH)D on muscle (30, 31). Finally, we did not measure 1,25 dihydroxy vitamin D, the biologically active form of vitamin D, but others have reported that 25(OH)D is appropriate for the study of muscle (8).
In conclusion, vitamin D status is significantly associated with muscle power and force. These data highlight the importance of vitamin D status on muscle function in adolescent girls. Suboptimal force might have implications for long-term bone development. The long-term implications of these observations require further study.
Results
A total of 99 girls aged 12-14 yr participated in the study. Of those, 68% (n = 67) were of South Asian origin (Indian Pakistani and Bangladeshi), 12% were British Black (n = 12), and 20% (n = 20) were white-Caucasian. All girls performed at least one two-LJ or one set of one-LJ's satisfactorily to allow their inclusion in this study. None of the girls had symptoms associated with vitamin D deficiency.
Table 1 summarizes the demographical details of the study population; also included are summary statistics of the muscle function data. Serum concentrations of Ca, P, and alkaline phosphatase were normal. Median serum 25(OH)D concentration was 21.3 nmol/liter (range 2.5-88.5), and PTH concentration was 3.7 pmol/liter (range 0.47-26.2). Serum 25(OH)D concentration was positively, and weight negatively, correlated with all outcome variables (Table 2). Body height was positively correlated with jumping height and power.
After correction for weight, there was a positive relationship between 25(OH)D and velocity (P = 0.002), jump height (P = 0.006), power (P = 0.004), EFI (P = 0.003), and force (P = 0.04) (Table 3 and Fig. 1). There were no interactions between body weight and 25(OH)D status.
In general the effects were somewhat smaller for PTH than seen in 25(OH)D and failed to reach statistical significance in this sample, although there was a significant negative relationship between PTH and velocity of jump (P = 0.04) (Table 3).
The magnitude of these relations with 25(OH)D and PTH was largely unchanged if we additionally control for BMI and/or height, suggesting that the mechanism of action is independent of both growth and obesity.
Subjects and Methods
Participants
Participants attended an all girls, inner city multiethnic school in Manchester, United Kingdom, and were being screened for inclusion in a randomized trial of vitamin D supplementation. The school was chosen because we had previously found that a high level (>70%) of individuals had low 25(OH)D levels (<37.5 nmol/liter) without the presence of symptoms (3). Inclusion criteria were that the girls were post-menarchal, free from disabilities that precluded jumping, and that they did not suffer from chronic childhood conditions.
The school doctor (G.D.) introduced the study to the participants. All parents of girls, aged 12-14 yr, were then contacted by telephone (G.D.) to discuss the trial and confirm whether their daughters could participate in the RCT. Informed consent was gained from all parents and verbal assent gained from participants themselves. The study received ethical approval from Covance Clinical Research Unit Independent Ethics Committee and was registered with the European Clinical Trials Database (EudraCT No. 2005-004729-25).
Anthropometric measurements
Standing height was measured using a microtoise (Chasmors Ltd., London, UK). Before jumping, weight was measured using the JM platform during quiet stance.
JM
Peak jumping power, jump height, and velocity were measured using the Leonardo Mechanography Ground Reaction Force Platform, software version 4.1 (Novotec Medical GmbH, Pforzheim, Germany). Each participant performed three maximum countermovement jumps with arms moving freely and at least a 60-sec rest between each jump. Each jump was two footed, and participants were instructed to jump as high as possible. The jump of greatest height was used for data analysis.
JM measures ground reaction forces (N), and by integrating these, computes vertical velocity (m/sec) (16); power (kW) is the product of force and velocity. Jump height (m) can be calculated from velocity (m/sec) and time taken (sec) for jump. The Esslinger Fitness Index (EFI) is a comparison of power per kg of body weight to an age and gender-matched reference population (15, 17) and is expressed as a percentage. In children the reference population consisted of 312 individuals (177 females) from Germany (17). The precision, measured as coefficient of variation, of muscle power measurement in adults is 3.6% (14) and children 3.7% (17); peak jumping force [from two leg jumps (LJs)] in children was 8.9% (17).
Maximum voluntary force (N/kg) is measured from multiple one-LJs. The participant is instructed to hop as fast and hard as possible, landing only on the forefoot; data are expressed as force relative to body weight (force times body weight). In this study one-LJ measurements were taken on the dominant foot. Participants did three sets of one-LJ and the one with highest force selected.
25(OH)D and PTH measurements
Nonfasting blood samples (5-7 ml) were taken at the same time as jumping measurements to determine total 25(OH)D and PTH levels. Serum was stored at -20 C until measurement. Thawed samples were whirly mixed, and 1 ml serum was taken for extraction of vitamin D metabolites. Each sample was made up to 3 ml with 0.9% saline, and 20 μl 25-hydroxy[26,27-methyl-3H]cholecalciferol recovery tracer (TRK 655; GE Healthcare UK Ltd., Amersham Bucks, UK) was added to each tube, mixed, and allowed to equilibrate for 30 min. After equilibration, 3 ml acetonitrile was added, and each tube was whirly mixed for 30 sec before centrifugation at 2800 rpm, at 4 C for 15 min (Sigma Labs 4K15, Shrewsbury, Shropshire, UK). One milliliter of distilled water was added to each sample before application to C18 Sep Pak Cartridges (Waters UK, Elstree, Herts, UK) preconditioned with 2 ml methanol and 5 ml water. After application, cartridges were washed with 3 ml 65% methanol/water before elution with 3 ml acetonitrile. Samples were dried down under a stream of nitrogen and transferred to limited volume inserts using HPLC column solvent. Samples were blown down again, and exactly 250 μl column solvent was added to each vial then capped.
Separation of metabolites was by straight-phase HPLC (Waters UK) run overnight using a 5 μm, 4.6 x 255-mm Hewlett-Packard Zorbax-Sil Column (Hicrom, Reading, Berkshire, UK) eluted with hexane-propan2ol (98:2), and quantified by UV absorbance at 265 nm and corrected for recovery. Run time for each sample was approximately 16 min, ensuring good separation. Sensitivity was 2 ng/ml and interassay variation 6%. The assay laboratory is accredited to International Organization for Standardization 9001:2000 and International Organization for Standardization 13485:2003, and participates successfully in the Vitamin D External Quality Assessment Scheme. Serum intact PTH was measured using the IDS intact PTH ELISA kit(Immunodiagnostic Systems Ltd., Boldon, Tyne and Wear, UK) normal adult reference range 0.8-3.9 pmol/liter, sensitivity 0.06 pmol/liter, and intraassay and interassay coefficients of variation 4 and 6%, respectively (manufacturer's values).
Serum concentrations of Ca adjusted for albumin and inorganic phosphate (P) were measured using the Roche Modular P unit (Roche Diagnostics, Ltd., Burgess Hill, West Sussex, UK). Reference ranges for corrected Ca and P were 2.2-2.6 and 0.7-1.4 mmol/liter, respectively.
Data analysis
Baseline demographical and jumping details for the group are presented in Table 1. Pearson's correlation coefficients were calculated to determine the relationship between body habitus, serum 25(OH)D, and PTH concentrations and jumping variables: muscle power (W), velocity of jump (m/sec), jump height (m), EFI, and force per kg of body weight (N/kg).
Preliminary exploration of the data indicated that serum 25(OH)D and PTH concentrations were better represented on a logarithmic scale. Due to basic physical relationships, the jumping parameters are related directly to the amount of mass that is being moved in the jump, i.e. the body weight. Simple normalization (as in, e.g. Refs. 14 and 15) forces a particular relationship, and a priori it is unlikely that a simple linear relationship will hold across the whole weight range. Therefore, the data were adjusted for this effect by adding a weight function to the regression model, and preliminary analysis indicated that a simple linear function was indeed inadequate to represent the relationships of the jumping variables with weight but that a quadratic function was adequate. Analysis of covariance adjusting for weight (as a quadratic function) was used to determine whether vitamin D status [log-transformed 25(OH)D or PTH levels] affects muscle function. The effect of vitamin D was expressed as a slope per doubling of the serum 25(OH)D concentration. An interaction between weight and 25(OH)D was also tested. To address potential causal relationships, additional exploratory analyses, including body mass index (BMI) and height in the models [both expressed as age-specific SD scores relative to national standards (18, 19)], were performed but are not presented in detail.
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