Nitric oxide (NO) is a short-lived free radical that plays an important regulatory role in several biological processes. Cytokines such as interleukin-1, tumor necrosis factor, and interferon-gamma have been shown to stimulate NO production in many cells types. Although these cytokines are known to have potent effects on bone remodeling and osteoblast function, the role of NO as an effector molecule in bone has been little studied. Here we investigate the effects of cytokines and calciotropic hormones on NO production by human osteoblast-like cells (hOB) and the role of NO as a modulator of osteoblast growth. Unstimulated hOB produced little NO, as reflected by measurement of nitrite concentrations in hOB-conditioned medium. NO production was not significantly altered by PTH and 1,25-dihydroxyvitamin D or human recombinant interleukin-1 beta (10 U/ml), tumor necrosis factor-alpha (25 ng/ml), and interferon-gamma (100 U/ml) individually. Combinations of all three cytokines at these concentrations, however, dramatically increased both NO generation and cGMP production. The stimulatory effect of cytokines on NO production began 12 h after exposure and was inhibited by cycloheximide, actinomycin-D, dexamethasone, and the competitive inhibitor of NO synthase L-NG-monomethylarginine. Reverse transcription/polymerase chain reaction analysis of hOB RNA, followed by direct sequencing of the amplified products, showed that hOB express the inducible, rather than the endothelial or neuronal, forms of NO synthase. Cytokine-induced increases in NO production were associated with a marked inhibition of [3H]thymidine uptake to less than 10% of that observed in control cultures. Abrogation of NO synthesis with L-NG-monomethylarginine under these conditions significantly increased [3H]thymidine uptake to approximately 20% of the control value, suggesting that NO may partly be responsible for the inhibition of osteoblast proliferation induced by these cytokines. Our data indicate that proinflammatory cytokines induce NO production in osteoblast-like cells and show and that this mediator plays a role in regulating cell growth. These findings may have important implications for the pathogenesis and management of bone loss in diseases associated with cytokine activation, such as rheumatoid arthritis.

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Diabetic osteopenia is a recognised but neglected complication of diabetes mellitus. Reduced bone mass and an overall twofold increase in fracture rate has been shown to occur in Type I (insulin-dependent) diabetes mellitus [1]. The situation in Type II (non-insulin-dependent) diabetes mellitus is less clear-cut, with reports of increased, decreased or unaltered bone mass reflecting the underlying heterogeneity of Type II diabetes [2]. The pathogenesis of diabetic osteopenia remains unclear. The reduction in bone mass could be due, in part, to a failure to reach peak bone mass especially in younger subjects with Type I diabetes [3] and partly to abnormal bone turnover [4]. In adults net bone loss has been found with a Diabetologia (1998) Summary Osteopenia is a recognised complication of diabetes mellitus which could be due to abnormal bone turnover or disturbances in the calcium/parathyroid hormone/vitamin D axis or both. Genetic factors also play an important part in determining bone mass although this has not been studied in diabetes.Recently a polymorphism of the collagen type 1 a 1 (COL1A1) gene has been shown to be associated with low bone mass in British women. To identify subjects with diabetes who may be at risk of developing osteoporosis and fractures, we analysed bone mineral density in relation to the biochemical markers of bone turnover, calcium homeostasis and the COL1A1 genotype in a group of premenopausal women with Type I (insulin-dependent) diabetes mellitus (n = 31), Type II (non-insulin dependent) diabetes mellitus (n = 21) and control subjects (n = 20). Bone mineral density was lower at the femoral neck in the subjects with Type I diabetes (p = 0.08) as were serum 25-hydroxyvitamin D compared with control subjects (p = 0.023) and this was negatively correlated with serum collagen type 1 C-terminal propeptide (r = ±0.56, p < 0.001). Bone mineral density in Type II diabetes was not different from control subjects, after correction for body mass index. Bone resorption was, however, raised in the Type II diabetic subjects as reflected by the higher urinary deoxypyridinoline values (p = 0.016) and lower collagen type 1 C-terminal propeptide:deoxypyridinoline ratio (p = 0.04). In the whole group studied, subjects with the COL1A1 s' genotype had lower bone mineral density at the femoral neck (p = 0.01) which was partly attributable to a lower body mass index. Following multiple regression analysis body mass index and collagen type 1 C-terminal propeptide concentrations remained determinants of bone mass at all three sites, whereas genotype appeared to be a predictor of bone mass at the femoral neck only. We conclude that measurement of these variables could prove useful in firstly identifying those diabetic women at risk of osteoporosis and secondly guiding therapeutic intervention. [Diabetologia (1998)

Physical activity during the first three decades of life may increase peak bone mass and reduce future osteoporosis risk. The aim of this study was to determine the extent to which different components of physical activity may influence bone mineral status within a representative population sample of young men and women. Bone mineral density (BMD) and content (BMC) were determined at the lumbar spine and femoral neck in 242 men and 212 women, aged 20 -25 years, by dual-energy X-ray absorptiometry. Physical activity was assessed by a self-report questionnaire designed to measure the frequency and duration of physical activity and its components (i.e., work, non-sports leisure, sports-related activities, and peak strain sports activities). Potential confounding factors such as height, weight, diet, and smoking habits were also assessed. In multivariate linear regression models, sports activity and peak strain sports activity undertaken by men were strongly associated with both lumbar spine BMD (␤ ‫؍‬ , respectively) (all p < 0.01), but work and non-sports leisure activities were not. In women, there were no associations between bone measurements and any component of physical activity. In models involving all subjects the gender/sports activity, but not the gender/peak strain, interaction term was statistically significant. Sports activity explained 10.4% of the observed variance in lumbar spine BMD in men, but <1% in women. These results demonstrate the importance of sports activities, especially those involving high peak strain, in determining peak bone status in young men. Failure to observe this association in women reflects their lower participation in such activities, but they may have the same capacity to benefit from these activities as men. Intervention studies are warranted to determine whether peak bone density in women can be improved by participating, during childhood and adolescence, in sports activities involving high peak strain. 0