José M. Olmos

Seasonal deficiency of vitamin D in children : A potential target for osteoporosis-preventing strategies?

Peak bone mass attained after skeletal growth is a major determinant of the risk of developing osteoporosis later in life, hence the importance of nutritional factors that contribute to bone mass gain during infancy and adolescence. An adequate supply of vitamin D is essential for normal bone homeostasis. This study was undertaken to determine what the levels are of 25‐hydroxyvitamin D (25(OH)D) that may be considered desirable in children and to assess if normal children maintain these levels throughout the year. Vitamin D metabolites and parathyroid hormone (PTH) serum levels were measured in 21 children in March and October, prior to and after the administration of a daily supplement of 25(OH)D (40 μg for 7 consecutive days). There were inverse correlations between basal 25(OH)D levels and supplementation‐induced changes in serum 1,25(OH)2D (r = 0.57, p < 0.05) and PTH (r = 0.41, p < 0.05). When basal levels of 25(OH)D were below 20 ng/ml, the supplement induced an increase in serum 1,25(OH)2D; with basal 25(OH)D under 10–12 ng/ml, the supplement also decreased serum PTH. The lowest serum level of 25(OH)D in 43 normal children studied in summer was 13 ng/ml. Those results suggested that the lowest limit for desirable levels of 25(OH)D in children was somewhere between 12 and 20 ng/ml. However, 31% of 51 normal children studied in winter had levels below 12 ng/ml, and 80% had levels lower than 20 ng/ml. Those children are likely to have suboptimal bioavailability of vitamin D, which might hamper their achievement of an adequate peak bone mass. Since cutaneous synthesis of vitamin D is rather limited in winter, oral vitamin D supplementation should be considered.

Trend in hip fracture epidemiology over a 14-year period in a Spanish population

Spain lacks detailed data on hip fracture trends despite being the country with the greatest increase in the pensioner-to-provider ratio in Europe. We reproduced a study on hip fracture incidence in a region of northern Spain (Cantabria) carried out 14 years ago to determine whether a secular trend to change is taking place. If such a trend could be found, our objective was to determine whether the effect is solely due to ageing or whether additional variables are involved. We assessed the incidence of hip fracture in patients aged ≥50 years through clinical records from Emergency Units and Orthopedic Surgical Units of all hospitals in the region of Cantabria in 1988 and 2002. A total of 318 new cases of hip fracture were recorded in 1988 and 490 in 2002 (54% increase; p<0.001). No significant changes were noticed following an adjustment for age. Women accounted for the increase in crude hip fracture incidence [246 women and 72 men suffered a hip fracture in 1988 compared to 404 women and 86 men in 2002 (64% increase in women and 19% increase in men; p<0.005 and not significant, respectively)]. The female:male ratio was 3.4 in 1988 versus 4.7 in 2002; following age-adjustment, no significant changes were found (1.8 in 1988 and 1.9 in 2002). The increase in crude hip fracture incidence was greater at cervical (versus trochanteric) sites. Patient residence, time of the year, site of fracture, kind of injury, previous contralateral hip fracture, length of stay, and peri-operative mortality did not differ significantly. In conclusion, a crude hip fracture incidence increase of about 50% in the northern Spanish region of Cantabria has taken place over the last 14 years. This effect does not persist after adjustments have been made for age. The crude rate increase occurred mainly at the expense of women, with a more noticeable rise in cervical fractures as opposed to trochanteric lesions.

BONE DENSITOMETRY AND BIOCHEMICAL BONE REMODELING MARKERS IN TYPE 1 DIABETES MELLITUS

Bone mineral density (BMD) at the lumbar spine, quantified by dual energy X-ray absorptiometry, and biochemical bone remodeling markers (serum alkaline phosphatase, osteocalcin, tartrate-resistant acid phosphatase and urinary hydroxyproline) have been studied in 94 patients with diabetes mellitus aged 18-62 years. BMD was normal (1.13 +/- 0.02 g/cm2 in patients vs. 1.16 +/- 0.12 g/cm2 in controls), although it was found to be negatively correlated with HbA1, microalbuminuria, age and the duration of the disease. Serum alkaline phosphatase (188 +/- 75 I.U./l vs. 168 +/- 42 I.U./1; P < 0.03), serum tartrate-resistant acid phosphatase (14.3 +/- 4.3 I.U./l vs. 11.7 +/- 3.7 I.U./l; P < 0.0001) and urinary hydroxyproline (0.018 +/- 0.016 mmol/mmol creatinine vs. 0.011 +/- 0.008 mmol/mmol creatinine; P < 0.001) were higher in diabetics than in controls. Serum osteocalcin was lower (2.5 +/- 1.3 ng/ml vs. 3.4 +/- 1.2 ng/ml; P < 0.0001). No relationship was found between bone remodeling markers and BMD. It is concluded that lumbar BMD is normal in type 1 diabetic patients, although the degree of metabolic control, age and duration of the disease may affect it. In the light of the biochemical markers, bone remodeling may be disturbed in diabetes, but such disturbance seems to be unimportant regarding BMD.

Metabolic syndrome and bone metabolism: the Camargo Cohort Study

Objectives: The aims of this study were to compare in participants with and without metabolic syndrome (1) bone mineral density (BMD), (2) prevalent vertebral and nonvertebral fractures, and (3) calciotropic hormones and bone turnover markers and to examine the association of each component of metabolic syndrome with bone parameters. Methods: A cross-sectional study (495 men and 1,013 women) from the Camargo Cohort Study was conducted. A multivariable regression approach was used to analyze the relationship between the components of metabolic syndrome and bone parameters. Results: Women with metabolic syndrome had higher age-adjusted BMD at all localizations (P < 0.0001) than did women without metabolic syndrome. Adjusting for body mass index canceled out this difference at the spine and femoral neck, although borderline significance persisted at the total hip. Moreover, in regression analyses, waist circumference (P < 0.0001) and hypertension (P between 0.002 and <0.0001) highly correlated with BMD at the three sites. However, no significant differences in BMD were found in men between those with and without metabolic syndrome. No differences in the prevalence of vertebral or nonvertebral fractures between participants with metabolic syndrome and controls were found for either sex. 25-Hydroxyvitamin D was significantly lower (P < 0.0001) and parathyroid hormone was significantly higher (P < 0.0001) in women with metabolic syndrome than in women without metabolic syndrome, whereas no differences were seen in men. Propeptide of type I collagen and C-terminal telopeptide of type I collagen were significantly lower in participants with metabolic syndrome than in controls in either sex. Conclusions: Women with metabolic syndrome show higher BMD than controls do, mainly driven by their higher body weight. Bone remodeling in these women is lower. Despite the greater bone mass and lower bone turnover, fracture prevalence is not reduced, suggesting worse bone quality and/or higher tendency to fall. No differences in BMD or fractures were seen in men, suggesting that the impact of metabolic syndrome on bone is sex dependent.

Expression of Opioid Receptors in Osteoblast-Like MG-63 Cells, and Effects of Different Opioid Agonists on Alkaline Phosphatase and Osteocalcin Secretion by These Cells

We have previously shown that several stressful situations associated with tissue injury determine a decrease in serum osteocalcin concentration. Since reduced osteocalcin production is a marker of decreased osteoblastic activity, this finding could be related to the pathogenesis of osteoporosis secondary to some diseases. Endogenous opioids are involved in stress response. Proenkephalin-derived peptides have been shown to inhibit alkaline phosphatase activity, another marker of bone formation, in the murine cell line ROS-17/2.8. On the other hand, serum osteocalcin has been reported as being low in heroin abusers. We have therefore thought it of interest to study the presence of opioid receptors in the human osteoblast-like cell line MG-63, and to evaluate the effects of different opioid agonists on the secretion of alkaline phosphatase and osteocalcin by these cells. The presence of opioid receptors was studied by means of RT-PCR and immunohistochemistry. RT-PCR studies suggested the presence of specific mRNA for the three types of receptors, and immunohistochemistry clearly showed their occurrence. Osteocalcin synthesis was significantly inhibited by high concentrations of the mu agonists morphine and (D-Ala2,N-MePhe4,Gly5-ol)-enkephalin although no changes were seen with the delta agonist (D-Ala2,D-leu5)-enkephalin. Morphine-induced osteocalcin inhibition was abolished when osteoblastic cells were incubated simultaneously with naloxone, whereas it was potentiated when cells were preincubated with naloxone. None of the opioid agonists modified the secretion of alkaline phosphatase. In conclusion, human osteoblast-like cells MG-63 express the three types of opioid receptors. Endogenous opioids may be involved in the reduction of osteocalcin observed in stressful situations associated with tissue injury.

Bone turnover markers in Spanish postmenopausal women The Camargo cohort study

BACKGROUND This cross-sectional study was performed to determine the reference ranges for two bone turnover markers--aminoterminal propeptide of type I collagen (P1NP) and C-terminal telopeptide of type I collagen (beta-CrossLaps, beta-CTX)--in normal postmenopausal Spanish women as determined in serum by automated methods. METHODS A community-based population of 1080 healthy postmenopausal women was evaluated. Data regarding risk factors for osteoporosis and fractures were collected by means of a structured questionnaire. Fasting serum levels of P1NP, beta-CTX, 25-Hydroxivitamin D (25OHD), and intact parathyroid hormone (iPTH) were measured on the Elecsys 2010 automated analyzer (Roche). BMD at lumbar spine, femoral neck and total hip was determined by DXA. RESULTS The mean age of subjects was 63+/-9. Logarithmic transformation of both markers was performed to allow for normal distributions. Mid-95% ranges for P1NP and beta-CTX were 19-100 ng/ml and 0.112-1.018 ng/ml, respectively. Mean values of P1NP (47.7+/-19.9 ng/ml) were similar to those previously determined by the manufacturer of the assays, whereas beta-CTX mean values (0.387+/-0.197 ng/ml) were lower. Both markers were higher among osteoporotic women. CONCLUSIONS Values obtained from this well-characterized population study provide reference ranges for serum automated P1NP and beta-CTX in normal Spanish postmenopausal women.

Wnt receptors, bone mass, and fractures: gene-wide association analysis of LRP5 and LRP6 polymorphisms with replication

OBJECTIVE Genes explaining the susceptibility to osteoporosis have not been fully elucidated. Our objective was to explore the association of polymorphisms capturing common variations of the lipoprotein receptor-related protein (LRP) 5 and 6 genes, encoding two Wnt receptors, with femoral neck bone mineral density (BMD) and osteoporotic fractures of the spine and the hip. DESIGN Cross-sectional, case-control, and replication genetic association study. METHODS Thirty-nine tagging and functional single nucleotide polymorphisms (SNPs) were analyzed in a group of 1043 postmenopausal women and 394 women with hip fractures. The results were replicated in a different group of 342 women. RESULTS Three SNPs of the LRP6 gene were associated with BMD (nominal uncorrected P values <0.05) in the discovery cohort. One showed a significant association after multiple test correction; two of them were also associated in the replication cohort, with a combined standardized mean difference of 0.51 (P=0.009) and 0.47 (P<0.003) across rs11054704 and rs2302685 genotypes. In the discovery cohort, several LRP5 SNPs were associated with vertebral fractures (odds ratio (OR) 0.67; P=0.01), with hip fractures (unadjusted ORs between 0.59 and 1.21; P=0.005-0.033, but not significant after multiple test adjustment or age adjustment), and with height and the projected femoral neck area, but not with BMD. Transcripts of LRP5 and LRP6 were similarly abundant in bone samples. CONCLUSIONS In this study, we found common polymorphisms of LRP5 associated with osteoporotic fractures, and polymorphisms of the LRP6 gene associated with BMD, thus suggesting them as likely candidates to contribute to the explaination of the hereditary influence on osteoporosis.

Effects of interleukin-4 on human osteoblast-like cells

Interleukin-4 (IL-4) modulates the activity of a variety of lymphoid, hemopoietic and mesenchymal cells, but little is known about its influence on bone cells. We have studied the effects of IL-4 on the human osteoblast-like cell line MG63. IL-4 (0.1-50 ng/ml) inhibited cell proliferation. The effect did not depend on cell density, but it was more marked in serum-free cultures than in the presence of serum. IL-4 also induced a dose-dependent increase in the expression of alkaline phosphatase stimulated by 1,25-dihydroxyvitamin D3, a marker of differentiated osteoblast activity. However, IL-4 did not modify the secretion of interleukin-6 or tumor necrosis factor. These results suggest that interleukin-4 may play a role as a modulator of osteoblast activity.

Stress decreases the serum level of osteocalcin

During a study undertaken to characterize the time course of the osteocalcin response to bone fractures, lower than normal values of serum osteocalcin (1.7 +/- 0.9 vs. 3.3 +/- 1.3 ng/ml, P < 0.001) were found in the basal samples (blood obtained 48-72 h after fracturing). Suspecting that such a decrease could be due to the stress response induced by the fracture, for corticosteroids are known to diminish serum osteocalcin, we extended our study to another two stressful situations of high stress: acute myocardial infarction and elective abdominal surgery. Indeed, the concentration of osteocalcin was also significantly diminished in both of these (2.0 +/- 0.9 ng/ml, P < 0.0005; 1.5 +/- 1.0 ng/ml, P < 0.0001). To further characterize this phenomenon, the time course of osteocalcin changes during the 24 h following abdominal surgery was studied in a second group of patients. The decrease was found to begin soon after surgery. In order to exclude the immobilization present in those three situations as the cause of the decrease in serum osteocalcin, a group of patients with retinal detachment was studied. Their serum osteocalcin levels were normal. It is concluded that serum osteocalcin levels decrease in stressful situations. Therefore, they should be interpreted cautiously when used as a marker of osteoblastic activity in this setting.

Effect of calcitriol on the secretion of prostaglandin E2, interleukin 1, and tumor necrosis factor α by human monocytes☆

Cells of the monocyte/macrophage lineage express specific receptors for calcitriol (1,25-dihydroxyvitamin D3) and secrete prostaglandins and several cytokines with potent effects on bone metabolism. The aim of this study was to determine the effect of calcitriol on the secretion of prostaglandin E2 (PGE2), interleukin-1 (IL-1), and tumor necrosis factor (TNF alpha). Monocyte-enriched peripheral blood mononuclear cells (PBMC) from healthy subjects were cultured in the presence or absence of calcitriol (10(-11)-10(-7) M) and several stimulating agents. After 24 h, PGE2, IL-1, and TNF alpha were measured in the culture supernatants or lysates with specific immunoassays. Calcitriol induced a biphasic effect on PGE2 production by unstimulated cells and increased PGE2 synthesis by cells stimulated with either endotoxin or tau-interferon (IFN-tau). On the other hand, calcitriol inhibited the production of TNF alpha by monocytes stimulated with either IFN-tau or phorbol esters. This effect was not prevented by the addition of indomethacin, IL-1, or IL-2. Under the conditions used, we observed no effect of calcitriol on IL-1 alpha or IL-1 beta production. These results indicate that calcitriol induces in vitro marked changes in the secretion of monocyte products with known activity on bone cells. Further studies are needed to elucidate whether some effects of calcitriol on bone metabolism are mediated by the interaction of the sterol with cells of the immune system.