Cxcl12 Deletion in Mesenchymal Cells Increases Bone Turnover and Attenuates the Loss of Cortical Bone Caused by Estrogen Deficiency in Mice

CXCL12 is abundantly expressed in reticular cells associated with the perivascular niches of the bone marrow (BM) and is indispensable for B lymphopoiesis. Cxcl12 promotes osteoclastogenesis and has been implicated in pathologic bone resorption. We had shown earlier that estrogen receptor α deletion in osteoprogenitors and estrogen deficiency in mice increase Cxcl12 mRNA and protein levels in the BM plasma, respectively. We have now generated female and male mice with conditional deletion of a Cxcl12 allele in Prrx1 targeted cells (Cxcl12^∆Prrx1) and show herein that they have a 90% decrease in B lymphocytes but increased erythrocytes and adipocytes in the marrow. Ovariectomy increased the expression of Cxcl12 and B-cell number in the Cxcl12^f/f control mice, but these effects were abrogated in the Cxcl12^∆Prrx1 mice. Cortical bone mass was not affected in Cxcl12^∆Prrx1 mice. Albeit, the cortical bone loss caused by ovariectomy was greatly attenuated. Most unexpectedly, the rate of bone turnover in sex steroid–sufficient female or male Cxcl12^∆Prrx1 mice was dramatically increased, as evidenced by a more than twofold increase in several osteoblast- and osteoclast-specific mRNAs, as well as increased mineral apposition and bone formation rate and increased osteoclast number in the endosteal surface. The magnitude of the Cxcl12^∆Prrx1-induced changes were much greater than those caused by ovariectomy or orchidectomy in the Cxcl12^f/f mice. These results strengthen the evidence that CXCL12 contributes to the loss of cortical bone mass caused by estrogen deficiency. Moreover, they reveal for the first time that in addition to its effects on hematopoiesis, CXCL12 restrains bone turnover—without changing the balance between resorption and formation—by suppressing osteoblastogenesis and the osteoclastogenesis support provided by cells of the osteoblast lineage. © 2020 American Society for Bone and Mineral Research.     

Deletion of FGFR3 in Osteoclast Lineage Cells Results in Increased Bone Mass in Mice by Inhibiting Osteoclastic Bone Resorption

Fibroblast growth factor receptor 3 (FGFR3) participates in bone remodeling. Both Fgfr3 global knockout and activated mice showed decreased bone mass with increased osteoclast formation or bone resorption activity. To clarify the direct effect of FGFR3 on osteoclasts, we specifically deleted Fgfr3 in osteoclast lineage cells. Adult mice with Fgfr3 deficiency in osteoclast lineage cells (mutant [MUT]) showed increased bone mass. In a drilled‐hole defect model, the bone remodeling of the holed area in cortical bone was also impaired with delayed resorption of residual woven bone in MUT mice. In vitro assay demonstrated that there was no significant difference between the number of tartrate‐resistant acid phosphatase (TRAP)‐positive osteoclasts derived from wild‐type and Fgfr3‐deficient bone marrow monocytes, suggesting that FGFR3 had no remarkable effect on osteoclast formation. The bone resorption activity of Fgfr3‐deficient osteoclasts was markedly decreased accompanying with downregulated expressions of Trap, Ctsk, and Mmp 9. The upregulated activity of osteoclastic bone resorption by FGF2 in vitro was also impaired in Fgfr3‐deficient osteoclasts, indicating that FGFR3 may participate in the regulation of bone resorption activity of osteoclasts by FGF2. Reduced adhesion but not migration in osteoclasts with Fgfr3 deficiency may be responsible for the impaired bone resorption activity. Our study for the first time genetically shows the direct positive regulation of FGFR3 on osteoclastic bone resorption. © 2016 American Society for Bone and Mineral Research.     

The Loss of Estrogen and Progesterone Receptor Gene Expression in Human Breast Cancer

Hormone responsiveness is a critical determinantof breast cancer progression and management, and theresponse to endocrine therapy is highly correlated withthe estrogen receptor (ER)³ and progesterone receptor (PR) status of tumor cells. Thus, keyareas of study in breast cancer are those mechanismsthat regulate ER and PR expression in normal andmalignant breast tissues. One-third of all breastcancers lack ER and PR; these conditions are associatedwith less differentiated tumors and poorer clinicaloutcome. In addition, approximately one-half ofER-positive tumors lack PR protein and patients withthis phenotype are less likely to respond tohormonal therapies than those whose tumors express bothreceptors. Since PR is induced by ER; its presence is amarker of a functional ER. In this review, we will discuss possible mechanisms for loss of ER andPR gene expression, especially structural changes withineach gene including deletions, polymorphisms ormethylation. Improved understanding of the pathways that lead to loss of ER and/or PR proteinsshould allow the development of better predictiveindicators as well as novel therapeutic approaches totarget these hormone-independent cancers.     

Reduced Serum IGF‐1 Associated With Hepatic Osteodystrophy Is a Main Determinant of Low Cortical but Not Trabecular Bone Mass

Hepatic osteodystrophy is multifactorial in its pathogenesis. Numerous studies have shown that impairments of the hepatic growth hormone/insulin‐like growth factor‐1 axis (GH/IGF‐1) are common in patients with non‐alcoholic fatty liver disease, chronic viral hepatitis, liver cirrhosis, and chronic cholestatic liver disease. Moreover, these conditions are also associated with low bone mineral density (BMD) and greater fracture risk, particularly in cortical bone sites. Hence, we addressed whether disruptions in the GH/IGF‐1 axis were causally related to the low bone mass in states of chronic liver disease using a mouse model of liver‐specific GH‐receptor (GHR) gene deletion (Li‐GHRKO). These mice exhibit chronic hepatic steatosis, local inflammation, and reduced BMD. We then employed a crossing strategy to restore liver production of IGF‐1 via hepatic IGF‐1 transgene (HIT). The resultant Li‐GHRKO‐HIT mouse model allowed us to dissect the roles of liver‐derived IGF‐1 in the pathogenesis of osteodystrophy during liver disease. We found that hepatic IGF‐1 restored cortical bone acquisition, microarchitecture, and mechanical properties during growth in Li‐GHRKO‐HIT mice, which was maintained during aging. However, trabecular bone volume was not restored in the Li‐GHRKO‐HIT mice. We found increased bone resorption indices in vivo as well as increased basal reactive oxygen species and increased mitochondrial stress in osteoblast cultures from Li‐GHRKO and the Li‐GHRKO‐HIT compared with control mice. Changes in systemic markers such as inflammatory cytokines, osteoprotegerin, osteopontin, parathyroid hormone, osteocalcin, or carboxy‐terminal collagen cross‐links could not fully account for the diminished trabecular bone in the Li‐GHRKO‐HIT mice. Thus, the reduced serum IGF‐1 associated with hepatic osteodystrophy is a main determinant of low cortical but not trabecular bone mass. © 2017 American Society for Bone and Mineral Research.     

Estrogen receptor‐α is required for the osteogenic response to mechanical loading in a ligand‐independent manner involving its activation function 1 but not 2

Estrogen receptor‐α (ERα) is crucial for the adaptive response of bone to loading but the role of endogenous estradiol (E2) for this response is unclear. To determine in vivo the ligand dependency and relative roles of different ERα domains for the osteogenic response to mechanical loading, gene‐targeted mouse models with (1) a complete ERα inactivation (ERα^?/?), (2) specific inactivation of activation function 1 (AF‐1) in ERα (ERαAF‐1⁰), or (3) specific inactivation of ERαAF‐2 (ERαAF‐2⁰) were subjected to axial loading of tibia, in the presence or absence (ovariectomy [ovx]) of endogenous E2. Loading increased the cortical bone area in the tibia mainly as a result of an increased periosteal bone formation rate (BFR) and this osteogenic response was similar in gonadal intact and ovx mice, demonstrating that E2 (ligand) is not required for this response. Female ERα^?/? mice displayed a severely reduced osteogenic response to loading with changes in cortical area (?78% ± 15%, p < 0.01) and periosteal BFR (?81% ± 9%, p < 0.01) being significantly lower than in wild‐type (WT) mice. ERαAF‐1⁰ mice also displayed a reduced response to mechanical loading compared with WT mice (cortical area ?40% ± 11%, p < 0.05 and periosteal BFR ?41% ± 8%, p < 0.01), whereas the periosteal osteogenic response to loading was unaffected in ERαAF‐2⁰ mice. Mechanical loading of transgenic estrogen response element (ERE)‐luciferase reporter mice did not increase luciferase expression in cortical bone, suggesting that the loading response does not involve classical genomic ERE‐mediated pathways. In conclusion, ERα is required for the osteogenic response to mechanical loading in a ligand‐independent manner involving AF‐1 but not AF‐2. © 2013 American Society for Bone and Mineral Research     

Cortical Bone Loss Following Gastric Bypass Surgery Is Not Primarily Endocortical

Roux-en Y gastric bypass (RYGB) surgery is an effective treatment for obesity; however, it may negatively impact skeletal health by increasing fracture risk. This increase may be the result not only of decreased bone mineral density but also of changes in bone microstructure, for example, increased cortical porosity. Increased tibial and radial cortical porosity of patients undergoing RYGB surgery has been observed as early as 6 months postoperatively; however, local microstructural changes and associated biological mechanisms driving this increase remain unclear. To provide insight, we studied the spatial distribution of cortical porosity in 42 women and men (aged 46 ± 12 years) after RYGB surgery. Distal tibias and radii were evaluated with high-resolution peripheral quantitative computed tomography (HR-pQCT) preoperatively and at 12 months postoperatively. Laminar analysis was used to determine cortical pore number and size within the endosteal, midcortical, and periosteal layers of the cortex. Paired t tests were used to compare baseline versus follow-up porosity parameters in each layer. Mixed models were used to compare longitudinal changes in laminar analysis outcomes between layers. We found that the midcortical (0.927 ± 0.607 mm⁻² to 1.069 ± 0.654 mm⁻², p = 0.004; 0.439 ± 0.293 mm⁻² to 0.509 ± 0.343 mm⁻², p = 0.03) and periosteal (0.642 ± 0.412 mm⁻² to 0.843 ± 0.452 mm⁻², p < 0.0001; 0.171 ± 0.101 mm⁻² to 0.230 ± 0.160 mm⁻², p = 0.003) layers underwent the greatest increases in porosity over the 12-month period at the distal tibia and radius, respectively. The endosteal layer, which had the greatest porosity at baseline, did not undergo significant porosity increase over the same period (1.234 ± 0.402 mm⁻² to 1.259 ± 0.413 mm⁻², p = 0.49; 0.584 ± 0.290 mm⁻² to 0.620 ± 0.299 mm⁻², p = 0.35) at the distal tibia and radius, respectively. An alternative baseline-mapping approach for endosteal boundary definition confirmed that cortical bone loss was not primarily endosteal. These findings indicate that increases in cortical porosity happen in regions distant from the endosteal surface, suggesting that the underlying mechanism driving the increase in cortical porosity is not merely endosteal trabecularization. © 2022 American Society for Bone and Mineral Research (ASBMR).     

Trabecular and Cortical Microstructure and Fragility of the Distal Radius in Women

Fragility fractures commonly involve metaphyses. The distal radius is assembled with a thin cortex formed by fusion (corticalization) of trabeculae arising from the periphery of the growth plate. Centrally positioned trabeculae reinforce the thin cortex and transfer loads from the joint to the proximal thicker cortical bone. We hypothesized that growth‐ and age‐related deficits in trabecular bone disrupt this frugally assembled microarchitecture, producing bone fragility. The microarchitecture of the distal radius was measured using high‐resolution peripheral quantitative computed tomography in 135 females with distal radial fractures, including 32 girls (aged 7 to 18 years), 35 premenopausal women (aged 18 to 44 years), and 68 postmenopausal women (aged 50 to 76 years). We also studied 240 fracture‐free controls of comparable age and 47 healthy fracture‐free premenopausal mother‐daughter pairs (aged 30 to 55 and 7 to 20 years, respectively). In fracture‐free girls and pre‐ and postmenopausal women, fewer or thinner trabeculae were associated with a smaller and more porous cortical area (r = 0.25 to 0.71 after age, height, and weight adjustment, all p < 0.05). Fewer and thinner trabeculae in daughters were associated with higher cortical porosity in their mothers (r = 0.30 to 0.47, all p < 0.05). Girls and premenopausal and postmenopausal women with forearm fractures had 0.3 to 0.7 standard deviations (SD) fewer or thinner trabeculae and higher cortical porosity than controls in one or more compartment; one SD trait difference conferred odds ratio (95% confidence interval) for fracture ranging from 1.56 (1.01–2.44) to 4.76 (2.86–7.69). Impaired trabecular corticalization during growth, and cortical and trabecular fragmentation during aging, may contribute to the fragility of the distal radius. © 2014 American Society for Bone and Mineral Research.     

The Glucocorticoid Receptor in Osterix-Expressing Cells Regulates Bone Mass,Bone Marrow Adipose Tissue,and Systemic Metabolism in Female Mice During Aging

Hallmarks of aging-associated osteoporosis include bone loss, bone marrow adipose tissue (BMAT) expansion, and impaired osteoblast function. Endogenous glucocorticoid levels increase with age, and elevated glucocorticoid signaling, associated with chronic stress and dysregulated metabolism, can have a deleterious effect on bone mass. Canonical glucocorticoid signaling through the glucocorticoid receptor (GR) was recently investigated as a mediator of osteoporosis during the stress of chronic caloric restriction. To address the role of the GR in an aging-associated osteoporotic phenotype, the current study utilized female GR conditional knockout (GR-CKO; GR^fl/fl:Osx-Cre+) mice and control littermates on the C57BL/6 background aged to 21 months and studied in comparison to young (3- and 6-month-old) mice. GR deficiency in Osx-expressing cells led to low bone mass and BMAT accumulation that persisted with aging. Surprisingly, however, GR-CKO mice also exhibited alterations in muscle mass (reduced % lean mass and soleus fiber size), accompanied by reduced voluntary physical activity, and also exhibited higher whole-body metabolic rate and elevated blood pressure. Moreover, increased lipid storage was observed in GR-CKO osteoblastic cultures in a glucocorticoid-dependent fashion despite genetic deletion of the GR, and could be reversed via pharmacological inhibition of the mineralocorticoid receptor (MR). These findings provide evidence of a role for the GR (and possibly the MR) in facilitating healthy bone maintenance with aging in females. The effects of GR-deficient bone on whole-body physiology also demonstrate the importance of bone as an endocrine organ and suggest evidence for compensatory mechanisms that facilitate glucocorticoid signaling in the absence of osteoblastic GR function; these represent new avenues of research that may improve understanding of glucocorticoid signaling in bone toward the development of novel osteogenic agents. © 2021 American Society for Bone and Mineral Research (ASBMR).     

Brain-Derived Acetylcholine Maintains Peak Bone Mass in Adult Female Mice

Preclinical and clinical data support a role of the sympathetic nervous system in the regulation of bone remodeling, but the contribution of parasympathetic arm of the autonomic nervous system to bone homeostasis remains less studied. In this study, we sought to determine whether acetylcholine (ACh) contributes to the regulation of bone remodeling after peak bone mass acquisition. We show that reduced central ACh synthesis in mice heterozygous for the choline transporter (ChT) leads to a decrease in bone mass in young female mice, thus independently confirming the previously reported beneficial effect of ACh signaling on bone mass accrual. Increasing brain ACh levels through the use of the blood brain barrier (BBB)-permeable acetylcholinesterase inhibitor (AChEI) galantamine increased trabecular bone mass in adult female mice, whereas a peripheral increase in ACh levels induced by the BBB-impermeable AChEI pyridostigmine caused trabecular bone loss. AChEIs did not alter skeletal norepinephrine level, and induced an overall increase in osteoblast and osteoclast densities, two findings that do not support a reduction in sympathetic outflow as the mechanism involved in the pro-anabolic effect of galantamine on the skeleton. In addition, we did not detect changes in the commitment of skeletal progenitor cells to the osteoblast lineage in vivo in AChEI-treated mice, nor a direct impact of these drugs in vitro on the survival and differentiation of osteoblast and osteoclast progenitors. Last, ChT heterozygosity and galantamine treatment triggered bone changes in female mice only, thus revealing the existence of a gender-specific skeletal response to brain ACh level. In conclusion, this study supports the stimulatory effect of central ACh on bone mass accrual, shows that it also promotes peak bone mass maintenance in adult mice, and suggests that central ACh regulates bone mass via different mechanisms in growing versus sexually mature mice. © 2020 American Society for Bone and Mineral Research.     

The Estrogen Receptor: A Logical Target for the Prevention of Breast Cancer with Antiestrogens

A strategy for the prevention of breast cancerhas been refined over the last century beginning withthe first observation that oophorectomy caused diseaseregression in some patients, to the identification of the estrogen receptor some 60 years later,and finally to the synthesis of the first nonsteroidalantiestrogen. Tamoxifen was the first clinically usefulantiestrogen and has been used for the treatment of breast cancer for the last twenty-one yearsin the United States. It is therefore a logicalprogression that antiestrogens are now recognized asuseful agents for the prevention of breast cancer. We will discuss the estrogen receptor as a targetfor the treatment and now the prevention of breastcancer. Data from the National Surgical and BowelProject (NSABP)⁴ tamoxifen prevention trial will be discussed with the preliminary resultsof two other European studies. The status of breastcancer prevention to date involves the comparison of thecurrent standard of prevention, tamoxifen, with the osteoporosis prevention drug, raloxifene inan ongoing trial called Study of Tamoxifen andRaloxifene (STAR).     

Bone Volume Fraction and Fabric Anisotropy Are Better Determinants of Trabecular Bone Stiffness Than Other Morphological Variables

As our population ages, more individuals suffer from osteoporosis. This disease leads to impaired trabecular architecture and increased fracture risk. It is essential to understand how morphological and mechanical properties of the cancellous bone are related. Morphology‐elasticity relationships based on bone volume fraction (BV/TV) and fabric anisotropy explain up to 98% of the variation in elastic properties. Yet, other morphological variables such as individual trabeculae segmentation (ITS) and trabecular bone score (TBS) could improve the stiffness predictions. A total of 743 micro–computed tomography (μCT) reconstructions of cubic trabecular bone samples extracted from femur, radius, vertebrae, and iliac crest were analyzed. Their morphology was assessed via 25 variables and their stiffness tensor () was computed from six independent load cases using micro finite element (μFE) analyses. Variance inflation factors were calculated to evaluate collinearity between morphological variables and decide upon their inclusion in morphology‐elasticity relationships. The statistically admissible morphological variables were included in a multiple linear regression model of the dependent variable . The contribution of each independent variable was evaluated (ANOVA). Our results show that BV/TV is the best determinant of (r²_adj = 0.889), especially in combination with fabric anisotropy (r²_adj = 0.968). Including the other independent predictors hardly affected the amount of variance explained by the model (r²_adj = 0.975). Across all anatomical sites, BV/TV explained 87% of the variance of the bone elastic properties. Fabric anisotropy further described 10% of the bone stiffness, but the improvement in variance explanation by adding other independent factors was marginal (<1%). These findings confirm that BV/TV and fabric anisotropy are the best determinants of trabecular bone stiffness and show, against common belief, that other morphological variables do not bring any further contribution. These overall conclusions remain to be confirmed for specific bone diseases and postelastic properties. © 2015 American Society for Bone and Mineral Research.     

Use of the Autopsy to Study Ontogeny and Expression of the Estrogen Receptor Gene in Human Breast

Historically, autopsies have been a valuable resource for morphologic studies of the human breast, and have been used in conjunction with radiologic evaluation and epidemiology to provide information about population prevalence of pre-malignant and malignant disease. More recently, well-preserved post-mortem breast tissue has also been used to evaluate the status of genes and their expression. Using molecular techniques and immunohistochemistry, quantitation of gene expression and localization of proteins of hormone receptors, growth factor receptors, growth factors, cell-proliferation related antigens and proto-oncogenes can be evaluated in autopsy-derived breast tissue. Expression of the estrogen receptor (ER)³ gene at the mRNA and protein product levels has been evaluate d in breast tissue from infants, children, adolescent girls, and adult women in various phases of the menstrual cycle, pregnancy and post-menopausal. The results of these studies support a role for the ER gene in early as well as pubescent breast development, and also in normal cyclical and abnormal cell proliferation in the terminal-duct-lobular-units (TDLU) of adult women.     

Validation of the Surrogate Threshold Effect for Change in Bone Mineral Density as a Surrogate Endpoint for Fracture Outcomes: The FNIH-ASBMR SABRE Project

The surrogate threshold effect (STE) is defined as the minimum treatment effect on a surrogate that is reliably predictive of a treatment effect on the clinical outcome. It provides a framework for implementing a clinical trial with a surrogate endpoint. The aim of this study was to update our previous analysis by validating the STE for change in total hip (TH) BMD as a surrogate for fracture risk reduction; the novelty of this study was this validation. To do so, we used individual patient data from 61,415 participants in 16 RCTs that evaluated bisphosphonates (nine trials), selective estrogen receptor modulators (four trials), denosumab (one trial), odanacatib (one trial), and teriparatide (one trial) to estimate trial-specific treatment effects on TH BMD and all, vertebral, hip, and nonvertebral fractures. We then conducted a random effects meta-regression of the log relative fracture risk reduction against 24-month change in TH BMD, and computed the STE as the intersection of the upper 95% prediction limit of this regression with the line of no fracture reduction. We validated the STE by checking whether the number of fractures in each trial provided 80% power and determining what proportion of trials with BMD changes ≥ STE reported significant reductions in fracture risk. We applied this analysis to (i) the trials on which we estimated the STE; and (ii) trials on which we did not estimate the STE. We found that the STEs for all, vertebral, hip, and nonvertebral fractures were 1.83%, 1.42%, 3.18%, and 2.13%, respectively. Among trials used to estimate STE, 27 of 28 were adequately powered, showed BMD effects exceeding the STE, and showed significant reductions in fracture risk. Among the validation set of 11 trials, 10 met these criteria. Thus STE differs by fracture type and has been validated in trials not used to develop the approach. © 2021 The Authors. Journal of Bone and Mineral Research published by Wiley Periodicals LLC on behalf of American Society for Bone and Mineral Research (ASBMR).     

The role of estrogen receptor α in growth plate cartilage for longitudinal bone growth

Estrogens enhance skeletal growth during early sexual maturation, whereas high estradiol levels during late puberty result in growth plate fusion in humans. Although the growth plates do not fuse directly after sexual maturation in rodents, a reduction in growth plate height is seen by treatment with a high dose of estradiol. It is unknown whether the effects of estrogens on skeletal growth are mediated directly via estrogen receptors (ERs) in growth plate cartilage and/or indirectly via other mechanisms such as the growth hormone/insulin‐like growth factor 1 (GH/IGF‐1) axis. To determine the role of ERα in growth plate cartilage for skeletal growth, we developed a mouse model with cartilage‐specific inactivation of ERα. Although mice with total ERα inactivation displayed affected longitudinal bone growth associated with alterations in the GH/IGF‐1 axis, the skeletal growth was normal during sexual maturation in mice with cartilage‐specific ERα inactivation. High‐dose estradiol treatment of adult mice reduced the growth plate height as a consequence of attenuated proliferation of growth plate chondrocytes in control mice but not in cartilage‐specific ERα^?/? mice. Adult cartilage‐specific ERα^?/? mice continued to grow after 4 months of age, whereas growth was limited in control mice, resulting in increased femur length in 1‐year‐old cartilage‐specific ERα^?/? mice compared with control mice. We conclude that during early sexual maturation, ERα in growth plate cartilage is not important for skeletal growth. In contrast, it is essential for high‐dose estradiol to reduce the growth plate height in adult mice and for reduction of longitudinal bone growth in elderly mice. © 2010 American Society for Bone and Mineral Research.     

Troglitazone Treatment Increases Bone Marrow Adipose Tissue Volume but Does not Affect Trabecular Bone Volume in Mice

Aging is associated with decreased trabecular bone mass and increased adipocyte formation in bone marrow. As osteoblasts and adipocytes share common precursor cells present in the bone marrow stroma, it has been proposed that an inverse relationship exists between adipocyte and osteoblast differentiation. In order to test this hypothesis, we studied mice treated with troglitazone (n = 9) given as a 0.2% of food admixture (2.0 g troglitazone per kg food) for 10 months and control mice (n = 9). Troglitazone is a potent stimulator of adipogenesis acting at the nuclear receptor: peroxisome proliferator activated receptor-gamma (PPARgamma). Histomorphometric analysis of proximal tibia was performed in order to quantitate the amount of trabecular bone volume per total volume (BV/TV %), adipose tissue volume per total volume (AV/TV %), and hematopoietic marrow volume per total volume (HV/TV %) using the point-counting technique. Bone size did not differ between the two groups. In troglitazone-treated mice, AV/TV was significantly higher than in control mice (4.7+/-2.1% vs. 0.2+/-0.3%, respectively, mean +/- SD, P < 0.001). BV/TV was similar in the two groups (16.9+/-5.6% for troglitazone-treated group vs. 14.9+/-4.7% for control group) as well as ash weight of the vertebrae. HV/TV was reduced in troglitazone-treated mice compared with control mice (78.4+/-6.8% vs. 84.9+/-4.7%, respectively, P < 0.05) and the presence of vascular sinusoids was reduced (7.3+/-1.7% vs. 16.1+/-5.6%, respectively, P < 0.05). Our data demonstrate that adipogenesis and osteogenesis can be regulated independently. Troglitazone-induced adipogenesis in the bone marrow may be caused by changes in the bone marrow vascularity.     

The role of activation functions 1 and 2 of estrogen receptor‐α for the effects of estradiol and selective estrogen receptor modulators in male mice

Estradiol (E2) is important for male skeletal health and the effect of E2 is mediated via estrogen receptor (ER)‐α. This was demonstrated by the findings that men with an inactivating mutation in aromatase or a nonfunctional ERα had osteopenia and continued longitudinal growth after sexual maturation. The aim of the present study was to evaluate the role of different domains of ERα for the effects of E2 and selective estrogen receptor modulators (SERMs) on bone mass in males. Three mouse models lacking either ERαAF‐1 (ERαAF‐1⁰), ERαAF‐2 (ERαAF‐2⁰), or the total ERα (ERα^?/?) were orchidectomized (orx) and treated with E2 or placebo. E2 treatment increased the trabecular and cortical bone mass and bone strength, whereas it reduced the thymus weight and bone marrow cellularity in orx wild type (WT) mice. These parameters did not respond to E2 treatment in orx ERα^?/? or ERαAF‐2⁰ mirx ERαAF‐1⁰ mice were tissue‐dependent, with a clear response in cortical bone parameters and bone marrow cellularity, but no response in trabecular bone. To determine the role of ERαAF‐1 for the effects of SERMs, we treated orx WT and ERαAF‐1⁰ mice with raloxifene (Ral), lasofoxifene (Las), bazedoxifene (Bza), or vehicle. These SERMs increased total body areal bone mineral density (BMD) and trabecular volumetric BMD to a similar extent in orx WT mice. Furthermore, only Las increased cortical thickness significantly and only Bza increased bone strength significantly. However, all SERMs showed a tendency toward increased cortical bone parameters. Importantly, all SERM effects were absent in the orx ERαAF‐1⁰ mice. In conclusion, ERαAF‐2 is required for the estrogenic effects on all evaluated parameters, whereas the role of ERαAF‐1 is tissue‐specific. All evaluated effects of Ral, Las and Bza are dependent on a functional ERαAF‐1. Our findings might contribute to the development of bone‐specific SERMs in males. © 2013 American Society for Bone and Mineral Research.     

Enteroendocrine K Cells Exert Complementary Effects to Control Bone Quality and Mass in Mice

The involvement of a gut-bone axis in controlling bone physiology has been long suspected, although the exact mechanisms are unclear. We explored whether glucose-dependent insulinotropic polypeptide (GIP)-producing enteroendocrine K cells were involved in this process. The bone phenotype of transgenic mouse models lacking GIP secretion (GIP-GFP-KI) or enteroendocrine K cells (GIP-DT) was investigated. Mice deficient in GIP secretion exhibited lower bone strength, trabecular bone mass, trabecular number, and cortical thickness, notably due to higher bone resorption. Alterations of microstructure, modifications of bone compositional parameters, represented by lower collagen cross-linking, were also apparent. None of these alterations were observed in GIP-DT mice lacking enteroendocrine K cells, suggesting that another K-cell secretory product acts to counteract GIP action. To assess this, stable analogues of the known K-cell peptide hormones, xenin and GIP, were administered to mature NIH Swiss male mice. Both were capable of modulating bone strength mostly by altering bone microstructure, bone gene expression, and bone compositional parameters. However, the two molecules exhibited opposite actions on bone physiology, with evidence that xenin effects are mediated indirectly, possibly via neural networks. Our data highlight a previously unknown interaction between GIP and xenin, which both moderate gut-bone connectivity. © 2020 American Society for Bone and Mineral Research.     

Targeting Bortezomib to Bone Increases Its Bone Anabolic Activity and Reduces Systemic Adverse Effects in Mice

Bortezomib (Btz) is a proteasome inhibitor approved by the FDA to treat multiple myeloma. It also increases bone volume by promoting osteoblast differentiation and inhibiting osteoclastogenesis in mice. However, Btz has severe systemic adverse effects, which would limit its use as a bone anabolic agent. Here, we designed and synthesized a bone-targeted form of Btz by conjugating it to a bisphosphonate (BP) with no antiresorptive activity. We report that BP-Btz inhibited osteoclast formation and bone resorption and stimulated osteoblast differentiation in vitro similar to Btz. In vivo, BP-Btz increased bone volume more effectively than Btz in three mouse models: untreated wild-type mice, mice with ovariectomy, and aged mice with tibial factures. Importantly, BP-Btz had significantly less systemic side effects than Btz, including less thymic cell death, sympathetic nerve damage, and thrombocytopenia, and it improved survival rates in aged mice. Thus, BP-Btz represents a novel anabolic agent to treat conditions, such as postmenopausal and age-related bone loss. Bone targeting is an attractive approach to repurpose approved drugs to treat skeletal diseases. © 2019 American Society for Bone and Mineral Research. © 2019 American Society for Bone and Mineral Research.     

PTH Treatment Increases Cortical Bone Mass More in Response to Compression than Tension in Mice

Parathyroid hormone (PTH) is an anabolic osteoporosis treatment that increases bone mass and reduces fracture risk. Clinically, the effects of PTH are site-specific, increasing bone mass more at the spine than the hip and not increasing bone mass at the radius. Differences in local loading environment between the spine, hip, and radius may help explain the variation in efficacy, as PTH and mechanical loading have been shown to synergistically increase bone mass. We hypothesized that differences in loading mode might further explain these variations. Owing to the curvature of the mouse tibia, cyclic compression of the hindlimb causes bending at the tibial midshaft, placing the anterior surface under tension and the posterior surface under compression. We investigated the combination of PTH treatment and tibial loading in an osteoblast-specific estrogen receptor-alpha knockout mouse model of low bone mass (pOC-ERαKO) and their littermate controls (LCs) and analyzed bone morphology in the tensile, compressive, and neutral regions of the tibial midshaft. We also hypothesized that pretreating wild-type C57Bl/6J (WT) mice with PTH prior to mechanical loading would enhance the synergistic anabolic effects. Compression was more anabolic than tension, and PTH enhanced the effect of loading, particularly under compression. PTH pretreatment maintained the synergistic anabolic effect for longer durations than concurrent treatment and loading alone. Together these data provide insights into more effective physical therapy and exercise regimens for patients receiving PTH treatment. © 2022 American Society for Bone and Mineral Research (ASBMR).