J.L. Berry

Vitamin D status and muscle function in post-menarchal adolescent girls

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.

A randomized, controlled trial of vitamin D supplementation upon musculoskeletal health in postmenarchal females

CONTEXT There has been a resurgence of vitamin D deficiency rickets throughout the developed world, with infants and adolescents being primarily affected. Adolescence is a crucial period for muscle and bone mineral accumulation. OBJECTIVE The aim was to determine the effect of vitamin D supplementation on the adolescent musculoskeletal system. DESIGN AND SETTING We conducted a community-based, double-blind, randomized controlled trial in a secondary school. PARTICIPANTS Postmenarchal 12- to 14-yr-old females participated in the trial. Ninety-nine were screened, 73 were included in randomized controlled trial, and 69 completed the trial. There were no adverse events. INTERVENTION Four doses of 150,000 IU vitamin D(2) (ergocalciferol) were given over 1 yr. MAIN OUTCOME MEASURES Dual-energy x-ray absorptiometry, peripheral quantitative computed tomography, and jumping mechanography were used. RESULTS At follow-up, 25-hydroxyvitamin D [25(OH)D] status was 56.0 ± 8.9 nmol/liter in the intervention group and 15.8 ± 6.6 nmol/liter in controls. There were no effects of supplementation on bone; however, for muscle function, efficiency of movement improved in the vitamin D-treated group. There was an interaction between baseline 25(OH)D concentration and response to vitamin D supplementation for muscle jump velocity. CONCLUSIONS Despite improvements in 25(OH)D status, treatment with vitamin D(2) was not shown to increase mineral accretion, bone geometry or strength, muscle force, or power. There were greater increases in jump velocity in girls with the lowest baseline 25(OH)D concentrations. Lack of effect of intervention after the period of peak mineral and muscle mass accretion suggests that earlier action is required.

Hypovitaminosis D among Healthy Adolescent Girls attending an Inner City School.

Aims: To determine the prevalence of hypovitaminosis D among healthy adolescent schoolgirls attending an inner city multiethnic girls’ school. Methods: Fifty one (28%) of 182 girls (14 white, 37 non-white; median age 15.3 years, range 14.7–16.6) took part in the study. Biochemical parameters, dietary vitamin D intake, muscle function parameters, duration of daily sunlight exposure (SE), and percentage of body surface area exposed (%BSA) were measured. Results: Thirty seven (73%) girls were vitamin D deficient (25-hydroxyvitamin D (25OHD) <30 nmol/l) and 9 (17%) were severely deficient (25OHD <12.5 nmol/l). The median (range) 25OHD concentration of white girls (37.3 nmol/l (18.3–73.3)) was higher than that of non-white girls (14.8 nmol/l (5.8–42.8)). The median (range) concentration of parathyroid hormone in white girls (2.8 pmol/l (1.0–3.7)) was lower than that of non-white girls (3.4 pmol/l (1.7–34.2)). Serum Ca, inorganic phosphate, alkaline phosphatase, and 1,25-dihydroxyvitamin D were not different in white and non-white girls. For the whole group, 25OHD concentration was related to the estimated SE and %BSA, but not to estimated intake of vitamin D. In white girls, the estimated SE and %BSA were significantly higher than that of non-white girls. The median times taken to complete the Gower’s manoeuvre and grip strength were not different in the two groups; these variables were not related to serum 25OHD. Conclusions: Hypovitaminosis D is common among healthy adolescent girls; non-white girls are more severely deficient. Reduced sunshine exposure rather than diet explains the difference in vitamin D status of white and non-white girls.

The role of sunlight exposure in determining the vitamin D status of the UK white Caucasian adult population.

Background  Vitamin D is necessary for bone health and is potentially protective against a range of malignancies. Opinions are divided on whether the proposed optimal circulating 25‐hydroxyvitamin D [25(OH)D] level (≥ 32 ng mL−1) is an appropriate and feasible target at population level.

The anomalous behaviour of exogenous 25-hydroxyvitamin D in competitive binding assays.

The Vitamin D International External Quality Assessment Scheme (DEQAS) was established in 1989 to monitor the performance of assays for 25-hydroxyvitamin D (25-OHD) and 1,25-dihydroxyvitamin D (I,25(OH)(2)D). This is achieved through the quarterly distribution of five samples of human serum. Results are used to calculate an All-Laboratory Trimmed Mean and a Method Mean for each of the methods used by participants. In July 2005, participants were asked to assay serum to which 50.9 nmol of either 25-OHD(3) or 25-OHD(2) had been added as ethanolic solutions. The final concentration of ethanol in the serum was 0.7%. The distribution also included a sample of the original serum (OS) containing 0.7% pure ethanol. The percentage recoveries of exogenous 25-OHD(3) (R1) and 25-OHD(2) (R2) were calculated for each method. Results (OS nM, R1 and R2) were as follows: DiaSorin RIA (n=53); 39.2, 82.1%, 83.3%, DiaSorin Liason (n=16); 36.8, 81.4%, 88.6%, IDS RIA (n=21); 36.4, 54.2%, 29.1%, IDS OCTEIA (n=16); 47.3, 78.8%, 56.4%, Nichols Advantage (n=21); 58.9, 46.4%, 43.2%, HPLC (n=9); 42.6, 112.2%, 97.1%, LC-MS (n=4); 34.0, 111.5%, 118.1%. The IDS RIA and Nichols assays gave unexpectedly low recoveries. This does not appear to be a calibration problem or the effect of ethanol.

Measurement of Plasma 1,25 Dihydroxyvitamin D Using a Novel Immunoextraction Technique and Immunoassay with Iodine Labelled Vitamin D Tracer

We evaluated a novel assay for the measurement of 1,25 dihydroxyvitamin D (1,25 (OH)2D). Immunoextraction of 1,25 (OH)2D is performed using a mini column containing a solid-phase monoclonal antibody followed by radioimmunoassay (RIA) using an 125I-labelled 1,25 (OH)2D derivative tracer and Sac-cell separation. The mean recovery of 1,25(OH)2D3 was 101%, linearity was excellent, inter- and intra-assay coefficients of variation were 9, 8 and 13% and 11, 10 and 14% at low, medium and high concentrations of 1,25(OH)2D3, respectively. The cross-reactivity of vitamin D metabolites was <0·0015% for 25-hydroxyvitamin D3, 24, 25 dihydroxyvitamin D3 and dihydrotachysterol and 0·54% for lα calcidol. 1,25 dihydroxyvitamin D2 cross-reactivity was 79%. The detection limit of the assay was 5pmol/L. Comparison with a commercial radio receptor assay (RRA) and an in-house RIA gave regression equations of y = 0·94x+11·8 (r = 0·98) and y = 0·91x-1·7 (r = 0.95), respectively, with no major discrepancies between the methods in all patient groups studied. Plasma concentrations of 1,25 (OH)2D obtained with the assay were as follows: normal, unsupplemented subjects: mean 88, range 48–155 pmol/L, n = 68, patients with chronic renal failure: mean 11, range 3–36 pmol/L, n = 27, primary hyperparathyroidism: mean 198, range 130–299 pmol/L, n = 23, Pagets disease: mean 92, range 42–149 pmol/L, n = 24, osteomalacia: mean 43, range 27–61 pmol/L, n = 9. A minimum sample volume of 300 μL is required, the hands-on time is significantly less than other commercial assays and the measuring procedure is gamma counting rather than scintillation counting. The assay offers several advantages over previous methods and should allow more laboratories to offer measurement of 1,25 (OH)2D as part of their repertoire.

Guinea‐pig isolated trachealis: the effects of charybdotoxin on mechanical activity, membrane potential changes and the activity of plasmalemmal K+‐channels

1 A study has been made, in guinea‐pig isolated trachealis, of the effects of charybdotoxin in modulating (a) the activity of large conductance K+‐channels, (b) the spontaneous electrical activity of intact cells and (c) the mechanical effects of some bronchodilator drugs. 2 Single smooth muscle cells were isolated from guinea‐pig trachealis by enzymic digestion and were studied by the patch clamp recording technique. Recordings were made from outside‐out plasmalemmal patches when the medium bathing the external surface of the patches contained 1.2 mm Ca2+ and 6 mm K+ while that bathing the cytosolic surface contained 0.1 μm Ca2+ and 140 mm K+. Charybdotoxin (100 nm), applied to the external surface of patches held at 0 mV, abolished the unitary currents associated with the opening of large conductance K+‐channels. 3 Opened segments of guinea‐pig trachea were used for the simultaneous recording of membrane potential and tension changes. In these experiments charybdotoxin (100 nm) caused the conversion of spontaneous electrical slow waves into spike‐like action potentials. This effect was accompanied by a very small reduction in resting membrane potential. 4 Tissue bath recording showed that charybdotoxin (100 nm) increased the spontaneous mechanical tone of the tissue, antagonized (2.8 fold in each case) the relaxant actions of isoprenaline and theophylline but did not antagonize the relaxant actions of cromakalim or RP 49356. 5 It is concluded that charybdotoxin is an effective inhibitor of large conductance K+‐channels in guinea‐pig trachealis cells. The ability of charybdotoxin to convert spontaneous slow waves into spike‐like action potentials suggests that the large, charybdotoxin‐sensitive, K+‐channels play an important role in determining the strong outward rectifying behaviour of the cells. The ability of charybdotoxin to antagonize isoprenaline and theophylline, but not to antagonize cromakalim and RP 49356, suggests that opening of the large conductance, charybdotoxin‐sensitive K+‐channel is implicated in the action of the former but not the latter pair of bronchodilator drugs.

The Vitamin D Debate: Translating Controlled Experiments into Reality for Human Sun Exposure Times

Exposure to sunlight, specifically the ultraviolet radiation, has both positive and negative health effects. Maximizing the benefits (vitamin D synthesis) while minimizing the damage is a multifaceted problem in which many of the elements are poorly quantified. Here we show how rigorously conducted large sample size laboratory studies of the effect of ultraviolet radiation dose on vitamin D status can be applied to real‐life situations. This was achieved by modeling the radiation incident on different surfaces for different solar locations, and equating with the controlled exposures in the laboratory studies. Results from both model and experimental data show that relatively short exposures of a modest amount of unprotected skin to summer sunlight in northern climes, on a regular basis during lunchtime hours, increases vitamin D to sufficiency status (≥20 ng mL−1) in the white Caucasian population. While both sun exposure conditions and human skin responses are variable in real life, these quantitative findings provide a guide for authorities devising sunlight exposure recommendations.

A sensitive radioimmunoassay using a monoclonal antibody that is equipotent for ercalcitriol and calcitriol (1,25-dihydroxy vitamin D2 and D3).

A monoclonal antibody has been used in a sensitive radioimmunoassay that measures 1,25-dihydroxyvitamin D2 (ercalcitriol) and 1,25-dihydroxyvitamin D3 (calcitriol) with equal potency. This important characteristic has not been reported for any other radioimmunoassay for 1,25-dihydroxyvitamin D. The two forms can be assayed in human serum together or individually after HPLC separation. Sample preparation entails acetonitrile extraction followed by C18-Sep-pak chromatography and HPLC. The assay measures 98% of added analyte, and achieves inter- and intra-assay coefficients of variation of 10.7% at 34 pg/ml and 7.8% at 81 pg/ml respectively. The limit of detection is 1.25 pg/tube and 50% displacement of bound ligand is achieved at 14 pg/tube. The reference interval is 20-50 pg/ml, mean 35. The correlation between results from the monoclonal radioimmunoassay and an established polyclonal antibody method was r = 0.98, slope 0.99. The assay has particular application in patients treated with vitamin D2 since 1,25-dihydroxyvitamin D2 can now be measured accurately in the presence of 1,25-dihydroxyvitamin D3.

Regulators of chondrocyte differentiation in tibial dyschondroplasia: an in vivo and in vitro study

Tibial dyschondroplasia (TD) is a disorder of endochondral bone growth and results in the retention of a mass of unmineralized, avascular cartilage extending into the metaphysis. We have studied various parameters of chondrocyte differentiation, both in isolated chick chondrocytes and growth plate sections, in an attempt to determine whether the inhibition in chondrocyte differentiation seen in TD is a consequence of an inherent incapability of chondrocytes to differentiate terminally and mineralize. Results from in vitro experiments indicated that both normal and lesion chondrocytes synthesized a matrix that stained with antibodies to types II and X collagen and displayed foci of mineralization. Alkaline phosphatase activity in lesion chondrocytes was significantly increased in comparison to that in normal hypertrophic chondrocytes. In addition, normal and lesion chondrocytes in culture synthesized transforming growth factor-beta and 24,25(OH)2D3 but not 1,25(OH)2D3. There was no significant difference in the production rate of these growth regulators between normal and lesion chondrocytes. In contrast, in growth plate sections, alkaline phosphatase activity was markedly reduced in the lesion chondrocytes and sites of mineralization were not evident. Type II collagen was located throughout the growth plate and lesion, but type X collagen was not present within the lesion except at sites of vascularization. These results indicate that, in culture, lesion chondrocytes have the ability to differentiate terminally and mineralize, and suggest that the primary abnormality in TD is related to a developmental fault which is only operative in vivo. This may include a defect in cartilage vascularization and/or impairment of chondrocyte differentiation by mechanisms that have not yet been elucidated but may involve the abnormal production of regulatory factors.