Relative impact of androgen and estrogen receptor activation in the effects of androgens on trabecular and cortical bone in growing male mice: a study in the androgen receptor knockout mouse model.

The relative importance of AR and ER activation has been studied in pubertal male AR knockout and WT mice after orchidectomy and androgen replacement therapy, either with or without an aromatase inhibitor. AR activation dominates normal trabecular bone development and cortical bone modeling in male mice. Moreover, optimal periosteal bone expansion is only observed in the presence of both AR and ER activation. INTRODUCTION: Androgen receptor (AR)-mediated androgen action has traditionally been considered a key determinant of male skeletal growth. Increasing evidence, however, suggests that estrogens are also essential for normal male bone growth. Therefore, the relative importance of AR-mediated and estrogen receptor (ER)-mediated androgen action after aromatization remains to be clarified. MATERIALS AND METHODS: Trabecular and cortical bone was studied in intact or orchidectomized pubertal AR knockout (ARKO) and male wildtype (WT) mice, with or without replacement therapy (3-8 weeks of age). Nonaromatizable (dihydrotestosterone [DHT]) and aromatizable (testosterone [T]) androgens and T plus an aromatase inhibitor (anastrazole) were administered to orchidectomized ARKO and WT mice. Trabecular and cortical bone modeling were evaluated by static and dynamic histomorphometry, respectively. RESULTS: AR inactivation or orchidectomy induced a similar degree of trabecular bone loss (-68% and -71%, respectively). Both DHT and T prevented orchidectomy-induced bone loss in WT mice but not in ARKO mice. Administration of an aromatase inhibitor did not affect T action on trabecular bone. AR inactivation and orchidectomy had similar negative effects on cortical thickness (-13% and -8%, respectively) and periosteal bone formation (-50% and -26%, respectively). In orchidectomized WT mice, both DHT and T were found to stimulate periosteal bone formation and, as a result, to increase cortical thickness. In contrast, the periosteum of ARKO mice remained unresponsive to either DHT or T. Interestingly, administration of an aromatase inhibitor partly reduced T action on periosteal bone formation in orchidectomized WT mice (-34% versus orchidectomized WT mice on T), but not in ARKO mice. This effect was associated with a significant decrease in serum IGF-I (-21% versus orchidectomized WT mice on T). CONCLUSIONS: These findings suggest a major role for AR activation in normal development of trabecular bone and periosteal bone growth in male mice. Moreover, optimal stimulation of periosteal growth is only obtained in the presence of both AR and ER activation.     

Androgens have antiresorptive effects on trabecular disuse osteopenia independent from muscle atrophy

Aging hypogonadal men are at increased risk of osteoporosis and sarcopenia. Testosterone is a potentially appealing strategy to prevent simultaneous bone and muscle loss. The androgen receptor (AR) mediates antiresorptive effects on trabecular bone via osteoblast-lineage cells, as well as muscle-anabolic actions. Sex steroids also modify the skeletal response to mechanical loading. However, it is unclear whether the effects of androgens on bone remain effective independent of mechanical stimulation or rather require indirect androgen effects via muscle. This study aims to characterize the effects and underlying mechanisms of androgens on disuse osteosarcopenia. Adult male mice received a unilateral botulinum toxin (BTx) injection, and underwent sham surgery or orchidectomy (ORX) without or with testosterone (ORX + T) or dihydrotestosterone (ORX + DHT) replacement. Compared to the contralateral internal control hindlimb, acute trabecular number and bone volume loss was increased by ORX and partially prevented DHT. T was more efficient and increased BV/TV in both hindlimbs over sham values, although it did not reduce the detrimental effect of BTx. Both androgens and BTx regulated trabecular osteoclast surface as well as tartrate-resistant acid phosphatase expression. Androgens also prevented BTx-induced body weight loss but did not significantly influence paralysis or muscle atrophy. BTx and ORX both reduced cortical thickness via endosteal expansion, which was prevented by T but not DHT. In long-term follow-up, the residual trabecular bone volume deficit in sham-BTx hindlimbs was prevented by DHT but T restored it more efficiently to pre-treatment levels. Conditional AR deletion in late osteoblasts and osteocytes or in the satellite cell lineage increased age-related trabecular bone loss in both hindlimbs without influencing the effect of BTx on trabecular osteopenia. We conclude that androgens have antiresorptive effects on trabecular disuse osteopenia which do not require AR actions on bone via muscle or via osteocytes.     

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.     

Role of the androgen receptor in skeletal homeostasis: the androgen-resistant testicular feminized male mouse model.

The role of androgen receptor-mediated androgen action on bone was investigated in testicular feminized male (Tfm) mice. Cortical bone was found to be unresponsive to testosterone (T) in orchidectomized Tfm mice, whereas cortical thickness as well as trabecular BMD and structure were fully maintained by T in the corresponding Tabby control mice. These data show an essential role for androgen receptor-mediated androgen action in periosteal bone formation. INTRODUCTION: Androgens can affect the male skeleton both directly-through activation of the androgen receptor (AR)-and indirectly-through stimulation of estrogen receptors after aromatization. We assessed the importance of AR-mediated androgen action on bone in a mouse model of androgen resistance. MATERIALS AND METHODS: Eight-week-old androgen-resistant testicular feminized male (Tfm) and Tabby control mice were orchidectomized (ORX) and treated for 4 weeks with a slow-release testosterone (T) pellet (delivering 167 microg/day) or a placebo pellet. A comprehensive analysis of the skeletal effects of androgen deficiency and replacement was performed using histomorphometry, QCT, and biochemical assessment of bone turnover. RESULTS: As expected, T increased trabecular BMD, volume, number, and width in ORX Tabby mice. In ORX Tfm mice, however, T had less effect on trabecular BMD and no effect on trabecular bone structure. T action on trabecular bone was associated with opposite changes in bone turnover: trabecular and endocortical bone turnover and serum levels of osteocalcin were all reduced by T in ORX Tabby mice, but not in ORX Tfm mice. T also increased cortical thickness (+16%), area, and density in ORX Tabby mice, but not in Tfm mice, resulting in greater bone strength in the Tabby control strain. The positive effects of T on cortical bone reflected a stimulatory effect on periosteal bone formation (+137%), which was again absent in Tfm mice. CONCLUSIONS: These data show that, in male mice, AR-mediated T action is essential for periosteal bone formation and contributes to trabecular bone maintenance.     

Sexual dimorphism in cortical bone size and strength but not density is determined by independent and time‐specific actions of sex steroids and IGF‐1: Evidence from pubertal mouse models

Although it is well established that males acquire more bone mass than females, the underlying mechanism and timing of this sex difference remain controversial. The aim of this study was to assess the relative contribution of sex steroid versus growth hormone–insulin‐like growth factor 1 (GH–IGF‐1) action to pubertal bone mass acquisition longitudinally in pubertal mice. Radial bone expansion peaked during early puberty (3 to 5 weeks of age) in male and female mice, with significantly more expansion in males than in females (+40%). Concomitantly, in 5 week old male versus female mice, periosteal and endocortical bone formation was higher (+70%) and lower (?47%), respectively, along with higher serum IGF‐1 levels during early puberty in male mice. In female mice, ovariectomy increased radial bone expansion during early puberty as well as the endocortical perimeter. In male mice, orchidectomy reduced radial bone expansion only during late puberty (5 to 8 weeks of age), whereas combined androgen and estrogen deficiency modestly decreased radial bone expansion during early puberty, accompanied by lower IGF‐1 levels. GHRKO mice with very low IGF‐1 levels, on the other hand, showed limited radial bone expansion and no skeletal dimorphism. From these data we conclude that skeletal sexual dimorphism is established during early puberty and depends primarily on GH–IGF‐1 action. In males, androgens and estrogens have stimulatory effects on bone size during late and early puberty, respectively. In females, estrogens limit bone size during early puberty. These longitudinal findings in mice provide strong evidence that skeletal dimorphism is determined by independent and time‐specific effects of sex steroids and IGF‐1. © 2010 American Society for Bone and Mineral Research     

The Role of GH/IGF-I-Mediated Mechanisms in Sex Differences in Cortical Bone Size in Mice

Cortical bone dimensions are important determinants of bone strength. Gender differences in cortical bone size caused by greater periosteal expansion in males than in females during the pubertal growth spurt are well established both in humans and in experimental animal models. However, the mechanism by which gender influences cortical bone size is still a matter of investigation. The role of androgens and estrogen in pubertal bone growth has been examined in human disorders as well as animal models, such as gonadectomized or sex steroid receptor knockout mice. Based on the findings that growth hormone (GH) and insulin-like growth factor I (IGF-I) are major regulators of postnatal skeletal growth, we and others have predicted that sex hormones interact with the GH/IGF-I axis to regulate cortical bone size. However, studies conflict as to whether estrogen and androgens impact cortical bone size through the canonical pathway, through GH without IGF-I mediation, through IGF-I without GH stimulation, or independent of GH/IGF-I. We review recent data on the impact of sex steroids and components of the GH/IGF axis on sexual dimorphism in bone size. While the GH/IGF-I axis is a major player in regulating peak bone size, the relative contribution of GH/IGF-dependent mechanisms to sex differences in cortical bone size remains to be established.     

Comparison of tamoxifen and testosterone propionate in male rats: differential prevention of orchidectomy effects on sex organs, bone mass, growth, and the growth hormone-IGF-I axis

Testis dysfunction can weaken bone and reduce muscle mass as well as impair sexual function. Testosterone (T) therapy has useful effects on sex organs, bone, and muscle in T-deficient males, but prostate concerns can preclude T use in some men. Although estrogens or other drugs can protect bone in men, gynecomastia makes estrogens unappealing, and other drugs may also be undesirable in some cases. Selective estrogen receptor modulators (SERMs) inhibit estrogen-evoked sex organ growth but mimic estrogen effects on bone and cholesterol and are advantageous for some women. SERMs may also be useful in men who must avoid androgens. As a preclinical test of this idea, tamoxifen (a SERM) and testosterone propionate (TP, a classic androgen) were compared for their efficacy in preventing varied effects of orchidectomy (ORX) in adult male rats. ORX led to ventral prostate and seminal vesicle atrophy and decreases in somatic growth, proximal tibia bone mineral density (BMD), and serum growth hormone (GH) and insulin-like growth factor I (IGF-I). ORX also increased anterior pituitary glandular kallikrein, serum cholesterol, and body temperature. Pituitary prolactin (PRL) content was unaltered. ORX effects on sex organs, somatic growth, IGF-I, cholesterol, body temperature, and pituitary kallikrein were prevented by TP at 1 mg/kg (3 doses per week), but BMD and GH were unresponsive. ORX effects on BMD and GH were prevented by TP at 10 mg/kg, but this dose evoked supraphysiologic increases in sex organs and PRL, failed to restore somatic growth, and further reduced IGF-I. Tamoxifen (1 mg/kg daily) prevented ORX effects on BMD, GH, and cholesterol without altering basal or TP-induced sex organ growth and further reduced IGF-I and somatic growth. Tamoxifen did not alter basal PRL but blocked increases caused by TP at 10 mg/kg. In summary, tamoxifen prevented ORX effects on bone and cholesterol in male rats without affecting sex organs or PRL and might be useful for men who must avoid androgens. Unexpectedly, a TP dose that replicated testis effects on sex organs and other targets had no effect on BMD or GH, and a larger TP dose that restored BMD and GH was worse at replicating normal male physiology. In addition, correlation/regression results suggested that the GH-IGF-I axis contributes to changes in BMD.     

Androgen and bone mass in men

Androgens have multiple actions on the skeleton throughout life. Androgens promote skeletal growth and accumulation of minerals during puberty and adolescence and stimulate osteoblast but suppress osteoclast function,activity and lifespan through complex mechanisms. Also androgens increase periosteal bone apposition, resulting in larger bone size and thicker cortical bone in men. There is convincing evidence to show that aromatization to estrogens was an important pathway for mediating the action of testosterone on bone physiology. Estrogen is probably the dominant sex steroid regulating bone resorption in men, but both testosterone and estrogen are important in maintaining bone formation. ( Asian J Androl 2003 Jun; 5: 148-154)     

Differential effects of androgens on cortical bone histomorphometry in gonadectomized male and female rats

The physiological role of gonadal androgens in regulating bone metabolism is not established. To determine if androgens antagonize the changes in cortical bone after gonadectomy, we treated orchiectomized (ORX) rats with testosterone (T) and 5 alpha-dihydrotestosterone (DHT), and ovariectomized (OVX) rats with the afore-mentioned androgens, as well as the synthetic androgen fluoxymesterone (Fl) and the nonsteroidal estrogen diethylstilbestrol (DES). OVX resulted in a rapid, sustained increase in periosteal bone formation at the tibial diaphysis, whereas ORX resulted in decreased bone formation. Androgen treatment stimulated bone formation in ORX rats and suppressed bone formation in OVX rats. A large dose of DES suppressed bone formation in OVX rats to values below the intact controls. The results of these studies demonstrate that androgens counteract the changes in cortical bone formation after gonadectomy in females as well as males, and thereby reestablish the sex difference observed in intact rats.     

Evidence from the aged orchidectomized male rat model that 17beta-estradiol is a more effective bone-sparing and anabolic agent than 5alpha-dihydrotestosterone.

This study was designed to evaluate the impact of estrogen versus androgen action on orchidectomy (ORX)-induced bone loss and associated changes in body composition. During an experimental period of 4 months, aged (12-month-old) ORX rats were treated with 17beta-estradiol (E2; 0.75 microg/day) or different doses of the nonaromatizable androgen 5alpha-dihydrotestosterone (DHT; 45, 75, and 150 microg/day, respectively), via subcutaneous (sc) silastic implants. Low doses of DHT and E2 inhibited the ORX-induced rise of bone turnover markers (serum osteocalcin and urinary deoxypyridinoline [DPD]) to a similar extent. High-dose DHT prevented the ORX-induced decrease of trabecular bone density but had no significant effect on cortical thinning as assessed by peripheral quantitative computed tomography (pQCT). This bone-sparing action of DHT occurred at the expense of hypertrophy of the ventral prostate and seminal vesicles. On the other hand, E2 restored both trabecular bone density and cortical thickness in ORX rats and even prevented age-related bone loss. In contrast to DHT, E2 increased lean body mass and inhibited the ORX-associated increase of fat mass, as measured by DXA. Administration of E2 was associated with increased serum concentrations of insulin-like growth factor (IGF) I and decreased circulating levels of leptin. We conclude that, in the aged ORX rat model, E2 is more effective in preventing ORX-induced bone loss than DHT. Additionally, E2 has anabolic effects on muscle tissue and prevents the ORX-related increase of fat mass. Overall, these data suggest that androgen action on bone and body composition is dependent on stimulation of both androgen receptors (ARs) and estrogen receptors (ERs).     

Postnatal growth and bone mass in mice with IGF-I haploinsufficiency

We examined the influence of IGF-I haploinsufficiency on growth, bone mass and osteoblast differentiation in Igf1 heterozygous knockout (HET) mice. Cohorts of male and female wild type (WT) and HET mice in the outbred CD-1 background were analyzed at 1, 2, 4, 8, 12, 15 and 18 months of age for body weight, serum IGF-I and bone morphometry. Compared to WT mice, HET mice had 20–30% lower serum IGF-I levels in both genders and in all age groups. Female HET mice showed significant reductions in body weight (10–20%), femur length (4–6%) and femoral bone mineral density (BMD) (7–12%) before 15 months of age. Male HET mice showed significant differences in all parameters at 2 months and thereafter. At 8 and 12 months, WT mice also showed a significant gender effect: despite their lower body weight, female mice had higher femoral BMD and femur length compared to males. Microcomputed tomography showed a significant reduction in cortical bone area (7–20%) and periosteal circumference (5–13%) with no consistent pattern of change in trabecular bone measurements in 2- and 8-month old HET mice in both genders. HET primary osteoblast cultures showed a 40% reduction in IGF-I protein expression and a 50% decrease in IGF-I mRNA expression. Cell growth and proliferation were decreased in HET cultures. Thus, IGF-I haploinsufficiency in outbred male and female mice resulted in reduced body weight, femur length and areal BMD at most ages. Serum IGF-I levels showed a high level of positive correlation with body weight and skeletal morphometry. These studies show that IGF-I is a determinant of bone size and mass in postnatal life. We speculate that impaired osteoblast proliferation may contribute to the skeletal phenotype of mice with IGF-I haploinsufficiency.     

Estrogen and androgen play distinct roles in bone turnover in male mice before and after reaching sexual maturity

Aromatase is the sole enzyme which converts androgen into estrogen. We have reported that aromatase-knockout (ArKO) mice showed bone loss by increased bone resorption not only in female but also in male mice, suggesting essential roles of estrogen in bone metabolism in both sexes. However, loss of testicular androgen by orchidectomy (ORX) could induce bone loss in male mice. To clarify the relationship between estrogen and androgen in bone metabolism in male mice, 7-week-old ArKO mice were orchidectomized (ORX) to induce a double deficiency of estrogen and androgen. Bone loss in ORX/ArKO mice was more severe than that in ORX/wild-type and sham/ArKO mice because of advanced bone resorption, indicating that androgen and estrogen individually regulate bone mass by suppressing bone resorption in male mice after reaching sexual maturity. Cortical bone formation was elevated in sham/ArKO mice, but ORX did not influence cortical bone formation in the adult male mice. To examine the influence of androgen deficiency in weaning stage, 3-week-old wild-type mice were orchidectomized. Four weeks after operation, periosteal bone formation in the femur was markedly reduced in ORX mice. Since cortical bone in the same age of ArKO mice was normal, testicular androgen is indispensable for cortical bone formation especially at puberty in male mice. Therefore, estrogen and androgen may play distinct roles in bone turnover of male mice before and after reaching sexual maturity.     

Androgen prevents hypogonadal bone loss via inhibition of resorption mediated by mature osteoblasts/osteocytes

Androgen receptor (AR) is expressed throughout the osteoblast lineage. Two different AR transgenic families (AR2.3-tg and AR3.6-tg mice) demonstrating overlapping and distinct expression profiles were employed to assess the effects of enhanced androgen sensitivity to ameliorate hypogonadal loss. Two different paradigms of steroid replacement following orchidectomy (ORX) were used as either preventative or therapeutic therapy. ORX was performed in male wild-type (WT), AR2.3-tg and AR3.6-tg mice at 5months with immediate DHT replacement (prevention, higher turnover) or at 3months with DHT treatment delayed for 2months (therapeutic, lower turnover), both with treatment for the last 6weeks. Dual energy X-ray absorptiometry (DXA), micro-computed tomography (μCT), and histomorphometry were performed. In the prevention model, ORX significantly reduced BMD and BMC in all genotypes compared to sham and DHT was effective at prevention of osteopenia. In the therapeutic model, all genotypes became osteopenic compared to sham, but after a prolonged hypogonadal period, delayed DHT treatment provided little benefit. μCT analysis of mid-shaft total bone in all genotypes generally showed reductions after ORX. Delayed DHT was ineffective at restoring bone volume in any genotype whereas immediate treatment prevented loss only in AR transgenic mice. Cortical thickness also decreased with ORX but immediate DHT treatment was effective to increase thickness only in WT mice, likely due to expansion of marrow volume in both AR-tg lines. In metabolically highly active cancellous bone, ORX resulted in lower bone volume/tissue volume (BV/TV) in all genotypes, consistent among 3 sites measured. Again with delayed treatment, there was little effect of DHT to restore BV/TV, but when administered at the time of ORX, DHT completely prevented the decrease in cancellous bone in all genotypes. Improvement in cancellous bone architecture was seen with immediate DHT replacement that was enhanced in AR transgenic lines compared to WT. In contrast, there were only modest changes in all genotypes using the delayed treatment paradigm. With ORX in both paradigms, trabecular number was decreased while spacing increased. Thus, androgen therapy is effective for the prevention of endosteal and cancellous osteopenia primarily through its anti-resorptive properties, but shows little anabolic action as a therapeutic strategy to restore bone. Given the similarity in response to androgen treatment in both AR transgenic lines, overlapping expression profiles suggest that the target cells mediating androgen action in vivo are mature osteoblast/osteocytes. Combined, these results demonstrate that in the adult mouse, androgen treatment can reduce bone resorption but has little overall anabolic activity.     

Expression of osteotropic growth factors and growth hormone receptor in a canine distraction osteogenesis model

Osteotropic growth factors play an important role in bone metabolism. Nevertheless, knowledge about their expression in relation to distraction osteogenesis remains limited. The aim of the present study was to determine the expression of growth hormone (GH), growth hormone receptor (GHR), insulin-like growth factor I (IGF-I), insulin-like growth factor II (IGF-II), and bone morphogenetic protein 2 (BMP-2) in distraction-induced bone regeneration. Expression of these factors was assessed during the consolidation phase, comparing distraction osteogenesis with osteotomy-induced bone formation. Real-time PCR was performed as a semiquantitative measurement of mRNA, and the relative expression levels of these factors were determined. In addition, plasma GH profiles and plasma concentrations of IGF-I, IGF-II, and insulin-like growth factor-binding protein 4 and -6 (IGFBP-4 and -6) were measured to assess their potential systemic role during bone formation. Expression of GHR, IGF-I, and BMP-2 had significantly increased in comparison with the expression of these factors in mature bone. Expression of GHR was significantly higher in distraction-induced bone regenerate than in osteotomy-induced bone. No significant differences were found for the expression of IGF-I and BMP-2 between distraction and osteotomy. Plasma concentrations of GH, IGF-I, IGF-II, IGFBP-4, and IGFBP-6 did not demonstrate any significant differences between treatment groups and controls. Upregulation of GHR expression in distraction osteogenesis may enhance sensitivity to endogenous systemic GH and thus promote consolidation of the regenerated bone. Changes in the systemic osteotropic growth factors GH, IGF-I, IGF-II, IGFBP-4, and IGFBP-6 do not seem to be of importance during distraction osteogenesis.     

Androgen Receptor Action in Osteoblasts in Male Mice Is Dependent on Their Stage of Maturation

Androgen action via the androgen receptor (AR) is essential for normal skeletal growth and bone maintenance post‐puberty in males; however, the molecular and cellular mechanisms by which androgens exert their actions in osteoblasts remains relatively unexplored in vivo. To identify autonomous AR actions in osteoblasts independent of AR signaling in other tissues, we compared the extent to which the bone phenotype of the Global‐ARKO mouse was restored by replacing the AR in osteoblasts commencing at either the 1) proliferative or 2) mineralization stage of their maturation. In trabecular bone, androgens stimulated trabecular bone accrual during growth via the AR in proliferating osteoblasts and maintained trabecular bone post‐puberty via the AR in mineralizing osteoblasts, with its predominant action being to inhibit bone resorption by decreasing the ratio of receptor activator of NF‐κB ligand (RANKL) to osteoprotegerin (OPG) gene expression. During growth, replacement of the AR in proliferating but not mineralizing osteoblasts of Global‐ARKOs was able to partially restore periosteal circumference, supporting the concept that androgen action in cortical bone to increase bone size during growth is mediated via the AR in proliferating osteoblasts. This study provides further significant insight into the mechanism of androgen action via the AR in osteoblasts, demonstrating that it is dependent on the stage of osteoblast maturation. © 2014 American Society for Bone and Mineral Research.     

Making rats rise to erect bipedal stance for feeding partially prevented orchidectomy-induced bone loss and added bone to intact rats.

The objectives of this study were to investigate the different effects on muscle mass and cancellous (proximal tibial metaphysis [PTM]) and cortical (tibial shaft [TX]) bone mass of sham-operated and orchidectomized (ORX) male rats by making rats rise to erect bipedal stance for feeding. Specially designed raised cages (RC) were used so that the rats had to rise to erect bipedal stance to eat and drink for 12 weeks. Dual-energy X-ray absorptiometry (DEXA) and peripheral quantitative computerized tomography (pQCT) were used to estimate the lean leg mass and bone mineral. Static and dynamic histomorphometry were performed on the triple-labeled undecalcified sections. We found that making the intact rats rise to erect bipedal stance for feeding increased muscle mass, cortical bone volume, and periosteal bone formation. Orchidectomy increased net losses of bone next to the marrow by increasing bone turnover. Making the ORX rats rise to erect bipedal stance increased muscle mass, partially prevented cancellous bone loss in the PTM, and prevented net cortical bone loss in TX induced by ORX by depressing cancellous and endocortical high bone turnover and stimulating periosteal bone formation. The bone-anabolic effects were achieved mainly in the first 4 weeks in the PTM and by 8 weeks in the TX. These findings suggested that making the rats rise to erect bipedal stance for feeding helped to increase muscle mass and cortical bone mass in the tibias of intact rats, increase muscle mass, and partially prevented cancellous and net cortical bone loss in ORX rats.     

Effects of liver-derived insulin-like growth factor I on bone metabolism in mice.

Insulin-like growth factor (IGF) I is an important regulator of both skeletal growth and adult bone metabolism. To better understand the relative importance of systemic IGF-I versus locally expressed IGF-I we have developed a transgenic mouse model with inducible specific IGF-I gene inactivation in the liver (LI-IGF-I-/-). These mice are growing normally up to 12 weeks of age but have a disturbed carbohydrate and lipid metabolism. In this study, the long-term effects of liver-specific IGF-I inactivation on skeletal growth and adult bone metabolism were investigated. The adult (week 8-55) axial skeletal growth was decreased by 24% in the LI-IGF-I-/- mice whereas no major reduction of the adult appendicular skeletal growth was seen. The cortical cross-sectional bone area, as measured in the middiaphyseal region of the long bones, was decreased in old LI-IGF-I-/- mice. This reduction in the amount of cortical bone was caused mainly by decreased periosteal circumference and was associated with a weaker bone determined by a decrease in ultimate load. In contrast, the amount of trabecular bone was not decreased in the LI-IGF-I-/- mice. DNA microarray analysis of 30-week-old LI-IGF-I-/- and control mice indicated that only four genes were regulated in bone whereas approximately 40 genes were regulated in the liver, supporting the hypothesis that liver-derived IGF-I is of minor importance for adult bone metabolism. In summary, liver-derived IGF-I exerts a small but significant effect on cortical periosteal bone growth and on adult axial skeletal growth while it is not required for the maintenance of the trabecular bone in adult mice.     

Comparative effects of orchidectomy and sciatic neurectomy on cortical and cancellous bone in young growing rats

 The purpose of the present study was to compare the effects of orchidectomy and sciatic neurectomy on cortical and cancellous bone in male rats. Fifty male Sprague-Dawley rats, 6 weeks of age, were randomized into five groups, with ten rats in each group: baseline control, age-matched intact control, orchidectomy (ORX), unilateral sciatic neurectomy (NX), and ORX + NX. After 8 weeks of feeding, the tibial shaft and proximal tibia were processed for cortical and cancellous bone histomorphometric analyses, respectively. ORX-induced reductions in maturation-related cortical and cancellous bone gains were attributable to decreased periosteal bone gain and increased trabecular bone resorption, respectively. NX- and ORX + NX-induced reductions in maturation-related cortical bone gain were attributable to decreased periosteal bone formation and increased endocortical bone turnover, while NX- and ORX + NX -induced reductions in maturation-related cancellous bone gain were attributable to increased bone resorption and decreased bone formation. NX more markedly reduced maturation-related cortical and cancellous bone gains than did ORX, and the ORX-induced reductions in maturation-related cortical and cancellous bone gains were more pronounced when combined with NX. The present study demonstrated differences in changes in cortical and cancellous bone following ORX and NX in young rats. The importance of mechanical loading, with or without testosterone deficiency, is emphasized in cortical and cancellous bone growth. Received: October 5, 2002 / Accepted: January 20, 2003 RID="*" ID="*" Offprint requests to: J. Iwamoto     

The Effects of Androgens on Murine Cortical Bone Do Not Require AR or ERα Signaling in Osteoblasts and Osteoclasts

In men, androgens are critical for the acquisition and maintenance of bone mass in both the cortical and cancellous bone compartment. Male mice with targeted deletion of the androgen receptor (AR) in mature osteoblasts or osteocytes have lower cancellous bone mass, but no cortical bone phenotype. We have investigated the possibility that the effects of androgens on the cortical compartment result from AR signaling in osteoprogenitors or cells of the osteoclast lineage; or via estrogen receptor alpha (ERα) signaling in either or both of these two cell types upon conversion of testosterone to estradiol. To this end, we generated mice with targeted deletion of an AR or an ERα allele in the mesenchymal (AR^f/y;Prx1‐Cre or ERα^f/f;Osx1‐Cre) or myeloid cell lineage (AR^f/y;LysM‐Cre or ERα^f/f;LysM‐Cre) and their descendants. Male AR^f/y;Prx1‐Cre mice exhibited decreased bone volume and trabecular number, and increased osteoclast number in the cancellous compartment. Moreover, they did not undergo the loss of cancellous bone volume and trabecular number caused by orchidectomy (ORX) in their littermate controls. In contrast, AR^f/y;LysM‐Cre, ERα^f/f;Osx1‐Cre, or ERα^f/f;LysM‐Cre mice had no cancellous bone phenotype at baseline and lost the same amount of cancellous bone as their controls following ORX. Most unexpectedly, adult males of all four models had no discernible cortical bone phenotype at baseline, and lost the same amount of cortical bone as their littermate controls after ORX. Recapitulation of the effects of ORX by AR deletion only in the AR^f/y;Prx1‐Cre mice indicates that the effects of androgens on cancellous bone result from AR signaling in osteoblasts—not on osteoclasts or via aromatization. The effects of androgens on cortical bone mass, on the other hand, do not require AR or ERα signaling in any cell type across the osteoblast or osteoclast differentiation lineage. Therefore, androgens must exert their effects indirectly by actions on some other cell type(s) or tissue(s). © 2015 American Society for Bone and Mineral Research.