E Van Herck

Intestinal calcium transporter genes are upregulated by estrogens and the reproductive cycle through vitamin D receptor-independent mechanisms.

1α,25(OH)2‐vitamin D strongly regulates the expression of the epithelial calcium channel CaT1. CaT1 expression is reduced in ERKOα mice and induced by estrogen treatment, pregnancy, or lactation in VDR WT and KO mice. Estrogens and vitamin D are thus independent potent regulators of the expression of this calcium influx mechanism, which is involved in active intestinal calcium absorption.

Aromatization of androgens is important for skeletal maintenance of aged male rats

A nonsteroidal aromatase inhibitor vorozole (VOR) was administered to aged (12 months old) male Wistar rats and its effect was compared with the effect of androgen deficiency. The rats were either sham-operated (SHAM) or orchidectomized (ORCH) and treated with or without VOR. Thus, four experimental groups were created (SHAM, ORCH, SHAM + VOR, ORCH + VOR). The follow-up period was 4 months. At the end of the experimental period, bone mineral density (BMD) of the first four lumbar vertebrae and right femur was measured ex vivo with dualenergy X-ray absorptiometry, bone formation was evaluated by serum osteocalcin, and bone resorption by urinary excretion of (deoxy)pyridinoline. Orchidectomy increased bone resorption 2-to 3-fold whereas bone formation was only slightly increased. Treatment of intact male rats with VOR also increased bone resorption (+30% increase) whereas bone formation was not increased in this SHAM + VOR group. Their BMD was 7% lower in the femur (P < 0.01) and 6% lower in the lumbar vertebrae (P < 0.01) compared with the SHAM group that had not received VOR. Moreover, this decrease of bone mineral density was not significantly different from the expected decrease of bone density observed in the ORCH groups (6–10%). This was also reflected by a decrease of calcium content of the first four lumbar vertebrae of 15% (P < 0.001) in the SHAM + VOR group and 9–14% (P < 0.05) in the ORCH groups compared with the SHAM group, respectively. These data therefore suggest that inhibition of aromatization of androgens into estrogens increases bone resorption and bone loss similar to that observed after complete removal of androgens. Aromatization of androgens into estrogens may therefore, at least partly, explain the effects of androgens on skeletal maintenance.

Skeletal effects of estrogen deficiency as induced by an aromatase inhibitor in an aged male rat model.

Aromatization of androgens into estrogens may be important for maintenance of the male skeleton. To address this hypothesis, we evaluated the skeletal effects of selective estrogen deficiency as induced by the aromatase inhibitor vorozole (Vor), with or without 17beta-estradiol (E(2)) administration (1.35 microg/day), in aged (12-month-old) male rats. A baseline group was killed at the start of the experiment (Base). The control group (Control), the group treated with vorozole alone (Vor), the group treated with E(2) alone (E(2)), or the group with a combination of both (Vor + E(2)) were killed 15 weeks later. Vorozole significantly increased serum testosterone (T) and reduced serum E(2) compared with Control. Body weight gain and serum insulin-like growth factor-I (IGF-I) were also lower in Vor, whereas significant weight loss and decrease of serum IGF-I occurred as a result of E(2) administration. Bone formation as assessed by serum osteocalcin was unaffected but osteoid surface in the proximal metaphysis of the tibia was increased in Vor-treated rats. Bone resorption as evaluated by urinary deoxypyridinoline excretion was increased in Vor. Biochemical parameters of bone turnover were reduced significantly in all E(2) treated rats. Premature closure of the growth plates and decreased osteoid and mineralizing surfaces were also observed in E(2) and Vor + E(2). Apparent bone density of lumbar vertebrae and femur, as measured by dual-energy X-ray absorptiometry (DXA), was significantly reduced in Vor. Vorozole decreased femoral bone density mainly in the distal femur (trabecular and cortical region). This decrease of bone density was not present in E(2) and Vor + E(2). Similar findings were observed when bone density was assessed by peripheral quantitative computed tomography (pQCT); that is, trabecular density of the distal femur, the proximal tibia, and the distal lumbar vertebra were all lower in Vor. This decrease in density was not observed in all E(2)-treated animals. In conclusion, administration of the aromatase inhibitor, vorozole, to aged male rats induces net trabecular bone loss in both the appendicular and axial skeleton, despite a concomitant increase in serum testosterone. E(2) administration is able to prevent this trabecular bone loss in vorozole-treated animals.

Evidence From the Aged Orchidectomized Male Rat Model That 17β‐Estradiol Is a More Effective Bone‐Sparing and Anabolic Agent Than 5α‐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 17β‐estradiol (E2; 0.75 μg/day) or different doses of the nonaromatizable androgen 5α‐dihydrotestosterone (DHT; 45, 75, and 150 μg/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).

Time-related increase of biochemical markers of bone turnover in androgen-deficient male rats

Bone loss during androgen deficiency has been associated with accelerated bone turnover and imbalance between bone formation and resorption but the relative increase of both phenomena is not well described. Serum osteocalcin as marker of bone formation and urinary excretion of pyridinoline (PYD) and deoxypyridinoline (DPD) as markers of bone resorption were measured in both orchidectomized (ORCH, n = 8) and sham-operated (SHAM, n = 8) aged (12-month-old) male rats from 2 days before until 66 days after surgery. PYD and DPD were significantly higher in the ORCH group compared to the SHAM group starting from 21 days after surgery until the end of the experiment. Serum osteocalcin was only significantly increased in the ORCH group at 30 and 40 days. The maximal increase of serum osteocalcin was also smaller than the increase in PYD and DPD (30% versus 74% and 112%, respectively). The two markers of bone resorption were correlated with osteocalcin (r = 0.63 for PYD and r = 0.71 for DPD). Based on these results, we conclude that (1) bone resorption, as measured by PYD and DPD, increased during androgen deficiency; (2) moreover, the increase of bone resorption, as measured by DPD and PYD, was followed by a more moderate increase in bone formation as measured by serum osteocalcin, supporting the hypothesis that androgen deficiency causes accelerated bone turnover and imbalance between bone resorption and bone formation.

Effects of exercise and disuse on bone remodeling, bone mass, and biomechanical competence in spontaneously diabetic female rats

Diabetes is associated with low bone formation. In this study we investigate the effect of additional or reduced mechanical loading on indices of bone formation and resorption, bone mass, and biomechanical properties in spontaneously diabetic BB rats. Female diabetic (mean age 13 weeks) and age-matched control rats were each allocated to three experimental groups: no-intervention; supervised running exercise program (Ex); and unloading induced by unilateral sciatic neurectomy (USN). The study period was 8 weeks. We measured biochemical parameters of bone formation (plasma osteocalcin) and resorption (urinary deoxypyridinoline [Dpd]); bone mineral density (BMD) by dual-energy X-ray absorptiometry (DXA) at middiaphyseal and metaphyseal regions of the femur; histomorphometry of the proximal tibial metaphysis (PTM); and biomechanical properties of the femur (neck, diaphysis, and metaphysis) and lumbar vertebra (L-5). In nondiabetic rats, Ex did not affect parameters of bone formation/resorption and BMD, and had little effect on biomechanical properties. USN increased Dpd excretion, whereas there was a decreased trabecular bone formation rate (BFR) on morphometry of PTM in both paralyzed and intact limbs. Compared with intact limbs, paralyzed limbs of USN rats showed decreased trabecular bone volume at the PTM, and decreased BMD and biomechanical properties at the distal femoral metaphysis (DFM) and, to a lesser extent, femoral neck. Diabetic rats of the three experimental groups had low plasma osteocalcin levels and Dpd excretion, as well as low BFR on morphometry. The BMD and biomechanical properties of both femur and L-5 were unchanged in diabetic rats. Diabetic Ex rats, however, showed a lower maximum load and stress at DFM than control Ex rats. Diabetic USN rats showed no increase in Dpd excretion; their paralyzed limbs showed decreased maximum load at DFM, but there was no significant decrease in trabecular bone volume at PTM or BMD at DFM. Thus, the running exercise does not affect low bone formation in diabetic rats; however, trabecular bone loss caused by disuse is less pronounced in diabetic rats, probably as a result of low bone resorption.

Androgen resistance and deficiency have different effects on the growing skeleton of the rat

Mature male, female, and androgen-resistant testicular feminized (Tfm) male rats of the same strain were sacrificed at the age of 120 days. Young male and Tfm rats were orchidectomized (orch) at 1 month of age and sacrificed at 120 days. The right femora were dissected, cleaned, defatted, and scanned with the Hologic QDR-1000. Orch and Tfm rats had similar body weights that were intermediate between body weights of their normal male and female littermates. Serum IGF-I concentrations were lowest in Tfm rats; IGF-1 concentrations in orch rats were not lower than in males. Dual-energy X-ray absorptiometry yielded the following results: Total femoral mass and area were lower in female, Tfm rats and in both orch groups compared with intact male rats. Femoral bone density was, however, only decreased in orch rats. Bone density measured in an area containing only cortical bone was not different between groups. However, the density was lower in orch rats in an area containing both cancellous and cortical bone. This finding is consistent with a ±50% decrease of cancellous bone volume in orch rats compared with all other groups at the proximal tibial metaphysis (an area containing mainly cancellous bone). These data show that Tfm rats, despite having lower IGF-I levels in serum, low body weight, and decreased femoral areas, manage—in contrast with orchidectomized rats—to maintain similar trabecular bone densities and volumes during growth. We conclude that trabecular bone densities can be preserved in androgen-resistant male rats independent of bone or body growth velocity or IGF-I secretion. We postulate that the modest increase of estrogen concentration in this animals and/or in situ aromatization may be responsible for the maintenance of the cancellous bone.

Effects of long-term diabetes and/or high-dose 17β-estradiol on bone formation, bone mineral density, and strenght in ovariectomized rats

Abstract Long-term diabetes in female rats preserves the bone mineral density (BMD) but impairs the strength of the femur. In this study, we have compared the effects of diabetes and high-dose 17β-estradiol (E 2 ), two conditions of low bone formation, in ovariectomized (ovx) rats. Spontaneously diabetic BB rats were ovx 0–3 days after onset, and nondiabetic ovx littermates were used as controls; the rats were either untreated or treated with E 2 (30 μg/day), subcutaenously), for 6 or 12 weeks (n = 9 in each of the eight groups). Analysis included: plasma 1,25-dihydroxyvitamin D 3 , insulin-like growth factor-I (IGF-I), and osteocalcin concentrations; histomorphometry of the proximal tibial metaphysis (PTM); and DXA and biomechanical testing of the femur. Both E 2 treatment and diabetes markedly lowered plasma IGF-I and osteocalcin concentrations, as well as dynamic morphometric parameters of bone formation in the PTM. Plasma IGF-I and osteocalcin were correlated ( R 2 = 0.55; p 2 treatment in both control and diabetic ovx rats increased the trabecular bone volume in the PTM and the BMD in the metaphysis of the distal femur; there was no difference between control and diabetic rats, however. The diaphyseal area and BMC were decreased in E 2 -treated or/and diabetic ovx rats, but the diaphyseal BMD remained unchanged compared with untreated ovx rats. The biomechanical properties of the whole femur (strength, angular deformation, and stiffness) were decreased in E 2 -treated and diabetic E 2 -treated ovx rats after 12 weeks. The data indicate that in situations of chronic low bone formation, whole bone strenght does not reflect total BMD but correlates better with bone size and bone mineral content measurements.

The Estrogen Receptor Ligand ICI 182,780 Does Not Impair the Bone-Sparing Effects of Testosterone in the Young Orchidectomized Rat Model

Testosterone (T) can affect bone metabolism not only directly, but also via its metabolites, estrogen or dihydrotestosterone, produced by enzymes present in bone. Therefore, the aim of this study was to investigate whether the high-affinity estrogen receptor ligand ICI 182,780 (ICI) impaired the bone-protective action of T in 3-month-old orchidectomized (Orch) rats, studied during an experimental period of 3 months. As expected, Orch significantly decreased trabecular bone volume in the proximal tibial metaphysis (-52%), as measured by histomorphometry, and had a similar negative effect on volumetric bone mineral density (BMD) in the distal femoral metaphysis (-53%), as assessed by peripheral quantitative computed tomography (pQCT). The loss of bone induced by Orch was completely prevented by T administration. Moreover, the Orch-associated increases of biochemical markers of bone turnover (serum osteocalcin, urinary deoxypyridinoline, and calcium excretion) did not occur when Orch rats received T. Administration of ICI in combination with T did not impair this bone-sparing effect. Cortical bone parameters (as determined by pQCT), body weight gain, and body composition (as measured by dual-energy X-ray absorptiometry) were not affected by T or ICI in combination with T. Furthermore, no differences were observed in serum concentrations of insulin-like growth factor-I or glucose homeostasis. In conclusion, ICI does not impair the long-term bone-protective effects of T in orchidectomized male rats, suggesting that testosterone can mediate its effect on the male skeleton directly via the androgen receptor. The absence of effects on body growth via the growth hormone--insulin-like growth factor-I axis may be a possible explanation for the lack of skeletal effects of this selective estrogen receptor antagonist.

Umbilical cord osteocalcin in normal pregnancies and pregnancies complicated by fetal growth retardation or diabetes mellitus.

A homologous radioimmunoassay for human osteocalcin was used to measure cord serum osteocalcin concentrations in normal pregnancies and in pregnancies complicated by intrauterine growth retardation (IUGR) and diabetes. The mean osteocalcin concentrations in term newborns (100-110 micrograms/l) were comparable with levels that we previously measured in pubertal children. There was a small increase in mean osteocalcin concentrations during the third trimester of fetal life, with the maximum at 35 weeks; between weeks 36 and 41, the osteocalcin levels dropped by 13%. Osteocalcin was 20% lower in IUGR neonates than in age- or weight-matched newborns. The newborns of diabetic mothers had markedly lower osteocalcin concentrations than the weight-matched neonates, and 16 of the 19 samples were more than 1 SD below the mean of the gestational-age osteocalcin regression curve. Cord serum osteocalcin appears to be a useful parameter in studying normal and abnormal fetal mineralization.