Khosla, Sundeep

Osteoporosis is an important clinical problem, affecting more than 50% of people over age 50 yr. Estrogen signaling is critical for maintaining proper bone density, and the identification of an endogenous selective estrogen receptor (ER) modulator, 27-hydroxycholesterol (27HC), suggests a mechanism by which nutritional/metabolic status can influence bone biology. With its levels directly correlated with cholesterol, a new possibility emerges wherein 27HC links estrogen and cholesterol signaling to bone homeostasis. In these studies, we found that increasing concentrations of 27HC, both by genetic and pharmacological means, led to decreased bone mineral density that was associated with decreased bone formation and increased bone resorption. Upon manipulation of endogenous estrogen levels, many of the responses to elevated 27HC were altered in such a way as to implicate ER as a likely mediator. In a model of postmenopausal bone loss, some pathologies associated with elevated 27HC were exacerbated by the absence of endogenous estrogens, suggesting that 27HC may act both in concert with and independently from classic ER signaling. These data provide evidence for interactions between estrogen signaling, cholesterol and metabolic disease, and osteoporosis. Patients with high cholesterol likely also have higher than average 27HC, perhaps putting them at a higher risk for bone loss and fracture. More studies are warranted to fully elucidate the mechanism of action of 27HC in bone and to identify ways to modulate this pathway therapeutically.

Abstract While female mice do not have the equivalent of a menopause, they do undergo reproductive senescence. Thus, to dissociate the effects of aging versus estrogen deficiency on age-related bone loss, we sham-operated, ovariectomized, or ovariectomized and estrogen-replaced female C57/BL6 mice at 6 months of age and followed them to age 18 to 22 months. Lumbar spines and femurs were excised for analysis, and bone marrow hematopoietic lineage negative (lin–) cells (enriched for osteoprogenitor cells) were isolated for gene expression studies. Six-month-old intact control mice were euthanized to define baseline parameters. Compared with young mice, aged/sham-operated mice had a 42% reduction in lumbar spine bone volume/total volume (BV/TV), and maintaining constant estrogen levels over life in ovariectomized/estrogen-treated mice did not prevent age-related trabecular bone loss at this site. By contrast, lifelong estrogen treatment of ovariectomized mice completely prevented the age-related reduction in cortical volumetric bone mineral density (vBMD) and thickness at the tibial diaphysis present in the aged/sham-operated mice. As compared with cells from young mice, lin– cells from aged/sham-operated mice expressed significantly higher mRNA levels for osteoblast differentiation and proliferation marker genes. These data thus demonstrate that, in mice, age-related loss of cortical bone in the appendicular skeleton, but not loss of trabecular bone in the spine, can be prevented by maintaining constant estrogen levels over life. The observed increase in osteoblastic differentiation and proliferation marker gene expression in progenitor bone marrow cells from aged versus young mice may represent a compensatory mechanism in response to ongoing bone loss. © 2010 American Society for Bone and Mineral Research.

Abstract Sex steroids are important regulators of bone turnover, but the mechanisms of their effects on bone remain unclear. Sclerostin is an inhibitor of Wnt signaling, and circulating estrogen (E) levels are inversely associated with sclerostin levels in postmenopausal women. To directly test for sex steroid regulation of sclerostin levels, we examined effects of E treatment of postmenopausal women or selective withdrawal of E versus testosterone (T) in elderly men on circulating sclerostin levels. E treatment of postmenopausal women (n = 17) for 4 weeks led to a 27% decrease in serum sclerostin levels [versus +1% in controls (n = 18), p < .001]. Similarly, in 59 elderly men, we eliminated endogenous E and T production and studied them under conditions of physiologic T and E replacement, and then following withdrawal of T or E, we found that E, but not T, prevented increases in sclerostin levels following induction of sex steroid deficiency. In both sexes, changes in sclerostin levels correlated with changes in bone-resorption, but not bone-formation, markers (r = 0.62, p < .001, and r = 0.33, p = .009, for correlations with changes in serum C-terminal telopeptide of type 1 collagen in the women and men, respectively). Our studies thus establish that in humans, circulating sclerostin levels are reduced by E but not by T. Moreover, consistent with recent data indicating important effects of Wnts on osteoclastic cells, our findings suggest that in humans, changes in sclerostin production may contribute to effects of E on bone resorption. © 2011 American Society for Bone and Mineral Research.

Abstract Recent studies have demonstrated an important role for circulating serotonin in regulating bone mass in rodents. In addition, patients treated with selective serotonin reuptake inhibitors (SSRIs) have reduced areal bone mineral density (aBMD). However, the potential physiologic role of serotonin in regulating bone mass in humans remains unclear. Thus we measured serum serotonin levels in a population-based sample of 275 women and related these to total-body and spine aBMD assessed by dual-energy X-ray absorptiometry, femur neck total and trabecular volumetric BMD (vBMD) and vertebral trabecular vBMD assessed by quantitative computed tomography (QCT), and bone microstructural parameters at the distal radius assessed by high-resolution peripheral QCT (HRpQCT). Serotonin levels were inversely associated with body and spine aBMD (age-adjusted R = −0.17 and −0.16, P < .01, respectively) and with femur neck total and trabecular vBMD (age-adjusted R = −0.17 and −0.25, P < .01 and < .001, respectively) but not lumbar spine vBMD. Bone volume/tissue volume, trabecular number, and trabecular thickness at the radius were inversely associated with serotonin levels (age-adjusted R = −0.16, −0.16, and −0.14, P < .05, respectively). Serotonin levels also were inversely associated with body mass index (BMI; age-adjusted R = −0.23, P < .001). Multivariable models showed that serotonin levels remained significant negative predictors of femur neck total and trabecular vBMD, as well as trabecular thickness at the radius, after adjusting for age and BMI. Collectively, our data provide support for a physiologic role for circulating serotonin in regulating bone mass in humans. © 2010 American Society for Bone and Mineral Research

Context: Aging, low dehydroepiandrosterone (DHEA), and testosterone are associated with increased adiposity and metabolic risk. Treatment with these hormones may improve these abnormalities. Objective: The objective of the study was to determine effects of aging, DHEA, or testosterone replacement on adiposity, meal fat partitioning, and postabsorptive lipolysis. Design: This was a cross-sectional, 2-yr, double-blind, randomized, placebo-controlled trial. Setting: The study was conducted in the general community. Patients: Elderly women and men (≥60 yr) with low DHEA sulfate (women and men) and bioavailable testosterone (men) concentrations and young adults. Interventions: Thirty elderly women each received 50 mg DHEA or placebo daily for 2 yr. Thirty elderly men received 75 mg DHEA, 29 received 5 mg testosterone (patch), and 32 received placebo daily for 2 yr. Thirty young women and 32 young men served as controls. Main Outcome Measures: In vivo measures of meal fat storage into sc fat, postabsorptive lipolysis, and regional adiposity at baseline and after treatment. Results: At baseline, the elderly had more body fat, greater systemic lipolysis (women, P = 0.0003; men, P < 0.0001) adjusted for resting energy expenditure, greater meal fat oxidation (women, P = 0.026; men, P = 0.0025), and less meal fat storage in sc fat (women, P = 0.0139; men, P= 0.0006). Although testosterone treatment increased meal fat storage into upper- vs. lower-body fat in elderly men, neither hormone affected regional adiposity, meal fat oxidation, or systemic lipolysis. Conclusions: Aging, in the context of low DHEA sulfate (women and men) and bioavailable testosterone (men) concentrations, is associated with changes in meal fat partitioning and postabsorptive lipolysis that are not corrected by DHEA and only partly corrected by testosterone replacement. DHEA or testosterone treatment of hormone-deficient elderly does not normalize adipose tissue lipolysis, but testosterone restores a “youthful” meal fat storage pattern in elderly men.

Abstract The incidence of distal forearm fractures peaks during the adolescent growth spurt, but the structural basis for this is unclear. Thus, we studied healthy 6- to 21-yr-old girls (n = 66) and boys (n = 61) using high-resolution pQCT (voxel size, 82 μm) at the distal radius. Subjects were classified into five groups by bone-age: group I (prepuberty, 6–8 yr), group II (early puberty, 9–11 yr), group III (midpuberty, 12–14 yr), group IV (late puberty, 15–17 yr), and group V (postpuberty, 18–21 yr). Compared with group I, trabecular parameters (bone volume fraction, trabecular number, and thickness) did not change in girls but increased in boys from late puberty onward. Cortical thickness and density decreased from pre- to midpuberty in girls but were unchanged in boys, before rising to higher levels at the end of puberty in both sexes. Total bone strength, assessed using microfinite element models, increased linearly across bone age groups in both sexes, with boys showing greater bone strength than girls after midpuberty. The proportion of load borne by cortical bone, and the ratio of cortical to trabecular bone volume, decreased transiently during mid- to late puberty in both sexes, with apparent cortical porosity peaking during this time. This mirrors the incidence of distal forearm fractures in prior studies. We conclude that regional deficits in cortical bone may underlie the adolescent peak in forearm fractures. Whether these deficits are more severe in children who sustain forearm fractures or persist into later life warrants further study.

Under most conditions, resorbed bone is nearly precisely replaced in location and amount by new bone. Thus, it has long been recognized that bone loss through osteoclast-mediated bone resorption and bone replacement through osteoblast-mediated bone formation are tightly coupled processes. Abundant data conclusively demonstrate that osteoblasts direct osteoclast differentiation. Key questions remain, however, as to how osteoblasts are recruited to the resorption site and how the amount of bone produced is so precisely controlled. We hypothesized that osteoclasts play a crucial role in the promotion of bone formation. We found that osteoclast conditioned medium stimulates human mesenchymal stem (hMS) cell migration and differentiation toward the osteoblast lineage as measured by mineralized nodule formation in vitro. We identified candidate osteoclast-derived coupling factors using the Affymetrix microarray. We observed significant induction of sphingosine kinase 1 (SPHK1), which catalyzes the phosphorylation of sphingosine to form sphingosine 1-phosphate (S1P), in mature multinucleated osteoclasts as compared with preosteoclasts. S1P induces osteoblast precursor recruitment and promotes mature cell survival. Wnt10b and BMP6 also were significantly increased in mature osteoclasts, whereas sclerostin levels decreased during differentiation. Stimulation of hMS cell nodule formation by osteoclast conditioned media was attenuated by the Wnt antagonist Dkk1, a BMP6-neutralizing antibody, and by a S1P antagonist. BMP6 antibodies and the S1P antagonist, but not Dkk1, reduced osteoclast conditioned media-induced hMS chemokinesis. In summary, our findings indicate that osteoclasts may recruit osteoprogenitors to the site of bone remodeling through SIP and BMP6 and stimulate bone formation through increased activation of Wnt/BMP pathways.