Effects of selective testosterone and estradiol withdrawal on skeletal sensitivity to parathyroid hormone in men

CONTEXT: Gonadal steroid withdrawal increases bone turnover and causes bone loss in men, but the underlying mechanisms have not been defined. We previously reported that gonadal steroid deprivation increases the skeletal sensitivity to the bone resorbing properties of PTH infusion in men, but it is not known whether this effect is mediated by the absence of androgens, estrogens, or both. OBJECTIVE: The objective of the study was to determine the selective effects of testosterone and estradiol withdrawal on the skeletal sensitivity to PTH infusion in healthy adult men. DESIGN AND SETTING: We randomly assigned 58 healthy men between the ages of 20 and 45 yr to receive treatment with combinations of a GnRH analog, an aromatase inhibitor, and hormone add-back therapy to produce the following treatment groups: group 1 (testosterone and estradiol deficient, n = 16); group 2 (testosterone sufficient but estradiol deficient, n = 12); group 3 (testosterone deficient but estradiol sufficient, n = 14); and group 4 (testosterone and estradiol sufficient, n = 16). Twenty-four-hour PTH infusions were performed at baseline and after 6 wk of therapy. Serum N-telopeptide (NTX), C-telopeptide (CTX), osteocalcin (OC), and amino-terminal propeptide of type I procollagen (P1NP) were measured every 6 h during the PTH infusions. RESULTS: Serum testosterone levels fell into the castrate range in groups 1 and 3, whereas estradiol levels were similarly reduced in groups 1 and 2. Gonadal steroid levels in the replaced groups were unchanged from baseline. Serum NTX levels measured before PTH infusion did not change in group 4 (+T, +E) but increased significantly in all other groups. A similar pattern was observed in serum CTX, although the increase in group 2 (+T, -E) was not significant (P = 0.12). Preinfusion concentrations of both OC and P1NP fell in most groups, but these changes were significant in group 2 (+T, -E) for both OC and P1NP and group 4 (+T, +E) for P1NP only. Serum NTX and CTX increased during PTH infusions in all groups at all time points (P < 0.001). In the eugonadal group (group 4 +T+E), the increase in NTX was the same at wk 0 and 6, whereas in all the other groups, the PTH-induced increase in serum NTX was significantly greater at wk, 6 compared with wk 0. The same pattern emerged for CTX, although the difference in group 3 (-T,+E) was not significant (P = 0.12). Serum OC and P1NP levels fell during PTH infusions in all groups and at all time points (P < 0.001), but no significant differences were observed between wk 0 and 6 in any group. CONCLUSIONS: These results demonstrate that the selective suppression of testosterone, estradiol, or both hormones increases the skeletal responsiveness to the bone-resorbing effects of PTH in men. These findings underscore the importance of both androgens and estrogens in male skeletal homeostasis and suggest that changes in skeletal sensitivity to PTH may play an important role in the pathogenesis of hypogonadal bone loss in men.     

Effects of gonadal suppression on the regulation of parathyroid hormone and 1,25-dihydroxyvitamin D secretion in women.

Although a causal association between estrogen deficiency and bone loss has been established for many years, the mechanism by which estrogen deficiency leads to bone loss is unclear. Estrogen deficiency could induce bone loss either by a direct effect on bone cells to modify the production of bone-resorbing cytokines or by altering the production or response to calcium regulatory hormones such as PTH and 1,25-dihydroxyvitamin D. To assess the effects of ovarian hormones on calcium regulatory hormones, we evaluated the ability of calcium to suppress PTH secretion and the ability of PTH to increase serum 1,25-dihydroxyvitamin D and whole blood ionic calcium levels in women before and after GnRH analog-induced ovarian suppression. Sixteen women with endometriosis underwent i.v. infusion of calcium (1.1 mg calcium gluconate/cc in 5% dextrose) at a rate of 4 cc/kg x h (n = 7) or human PTH-(1-34) (Parathar) at a dose of 0.55 U/kg x h (n = 9) before and after 6 months of suppression of ovarian function with the GnRH analog nafarelin acetate (200 microg, intranasally, twice daily). Initial infusions were performed between days 6-10 of the menstrual cycle. Serum PTH and whole blood ionic calcium levels were measured at -20, -10, and 0 min and then every 10 min for 2 h during i.v. calcium infusions. Whole blood ionic calcium and 1,25-dihydroxyvitamin D levels were measured every 6 h for 24 h during i.v. human PTH-(1-34) infusions. Serum estradiol levels were markedly suppressed by nafarelin therapy in both groups of women. The relationship between whole blood ionic calcium and serum PTH levels was similar before and during nafarelin-induced ovarian suppression. The net change and rate of rise in serum 1,25-dihydroxyvitamin D levels in response to PTH infusion were similar before and during nafarelin therapy. Peak whole blood ionic calcium and incremental increases in ionic calcium in response to PTH were similar before and during nafarelin therapy. Our data suggest that ovarian suppression does not alter the regulation of PTH secretion in response to calcium, the ability of PTH to stimulate 1,25-dihydroxyvitamin D formation, or the skeletal sensitivity to PTH. These findings suggest that alterations in calcium regulatory hormones by estrogen deficiency are unlikely to play a major role in the pathogenesis of estrogen deficiency bone loss.     

Use of vitamin K2 (menatetrenone) and 1,25-dihydroxyvitamin D3 in the prevention of bone loss induced by leuprolide.

The purpose of this study is to evaluate the efficacy of vitamin K2 and 1,25-dihydroxyvitamin D3 [1,25-(OH)2D3] in preventing bone loss induced by estrogen deficiency during therapy with the GnRH agonist (GnRH-a) leuprolide. One hundred ten women (mean age, 46.2+/-0.5 yr), receiving leuprolide therapy for estrogen-dependent diseases (such as endometriosis and uterine leiomyomas), were randomly allocated into four groups (group A, leuprolide only; group B, leuprolide with vitamin K2; group C, leuprolide with 1,25-(OH)2D3; and group D, leuprolide with vitamin K2 and 1,25-(OH)2D3). Bone mineral density of the lumbar spine was measured by dual-energy x-ray absorptiometry before and after 6 months of treatment. Bone formation and resorption markers were also measured before and after 6 months of treatment. There were no significant differences in the background parameters among the four groups. Bone mineral density was reduced in all four groups, but the percent changes varied slightly, at - 5.25% (group A), -3.72% (P < 0.05 vs. group A) (group B), -4.13% (group C), and -3.59% (P < 0.01 vs. group A) (group D), respectively. Bone formation markers were significantly increased in all four groups, and the percent changes of bone formation markers were highest in group B. Bone resorption markers also increased significantly in all four groups after treatment of 6 months. Group B tended to have the highest percent changes of bone resorption markers among the four groups, but these increases were not significantly different between any of the groups. Vitamin K2, especially when combined with 1,25-(OH)2D3, can partially prevent bone loss caused by estrogen deficiency. However, because this effect is attributable mainly to the activation of bone formation, it is not sufficient to eliminate bone loss induced by GnRH-a therapy.     

The effect of micronized estradiol on bone turnover and calciotropic hormones in older men receiving hormonal suppression therapy for prostate cancer

To examine the effect of estradiol (E(2)) without the confounding effect of hypothalamic-pituitary feedback, we studied men with prostate cancer in whom gonadotropin secretion was suppressed by LH-releasing hormone agonists (LHRH-A). Fourteen men over 65 yr of age and receiving established LHRH-A treatment (EST group) without bony metastases and 12 men who received LHRH-A as neoadjuvant therapy for locally advanced prostate cancer (NEO group) were randomized (double blind) to receive either 1 mg/d micronized E(2) (n = 12) or placebo (PL; n = 13) for 9 wk. E(2), estrone, testosterone, SHBG, PTH, and 25-hydroxy- and 1,25-dihydroxyvitamin D levels as well as markers of bone resorption [N- and C-telopeptide cross-links (NTX and CTX) and deoxypyridinoline] and bone formation (bone-specific alkaline phosphatase, osteocalcin, and N-terminal type I collagen) were measured before LHRH-A in the NEO group, before [baseline (BL)] and after 9 wk of E(2) or PL in all patients, and 6 wk after E(2) treatment in the EST group. In the NEO group, hormone levels fell 3 wk after the initial LHRH-A injection, and deoxypyridinoline increased significantly (P = 0.006). At BL, the EST group had higher bone turnover due to the longer duration of LHRH-A treatment. With E(2) treatment, E(2) levels rose into the normal male range, and two resorption markers decreased significantly from BL by 33% for NTX (P < 0.001) and 28% for CTX (P = 0.009). Bone formation markers did not change. PTH increased by 43% from BL (P < 0.01) in the E(2) group and decreased 16% from BL in the PL group (P < 0.01). Ionized calcium did not change in the E(2) group, but increased in the PL group by 2.3% (P < 0.01). NTX and CTX increased 6 wk after E(2) withdrawal in the EST group. We conclude that E(2) inhibits bone resorption in hypogonadal men through a direct skeletal effect that is independent of PTH. Low dose estrogen may be an option for the prevention and/or treatment of bone loss in this population.     

Changes in parameters of bone and mineral metabolism during therapy for hyperthyroidism

Hyperthyroid patients have high bone turnover and negative calcium and phosphorus balance often associated with mild osteopenia. Early during antithyroid treatment bone turnover decreases, the mineral balance is converted to positive, and sometimes hypocalcemia occurs. The aim of this investigation was to study the mechanisms of the changes in some parameters of bone and mineral metabolism after treatment of thyrotoxicosis. Thirteen newly diagnosed patients with Graves' disease (seven postmenopausal women, four premenopausal women, and two men) were studied longitudinally, every 6 weeks, for 1 yr after commencing antithyroid treatment with methimazole. Mean serum calcium and phosphorus were both slightly above the normal mean at week 0 and decreased significantly (by 10% and 24%, respectively) during treatment. Fasting urinary calcium was 236 +/- 4 (mean +/- SEM) mg/g creatinine, and the fractional excretion of Ca was 2.0 +/- 0.33% before treatment; both fell significantly to minimums of 61 +/- 20 mg/g and 0.6 +/- 0.16%, respectively. Urinary phosphorus was 282 +/- 60 mg/g creatinine, and the fractional excretion of phosphorus was 3.3 +/- 0.6% before treatment; both increased significantly to 452 +/- 40 mg/g and 8.4 +/- 1.0%, respectively, during treatment. The z-scores were calculated from the mean and SD ofthe respective control groups. The z-score of urinary N-telopeptides of type I collagen (U.NTx) was 9.3 +/- 1.3 at week 0 and declined exponentially, but failed to normalize after 1 yr of antithyroid treatment. The serum alkaline phosphatase (ALP) z-score was initially 2.2 +/- 0.2, increased to 6.0 +/- 1.0 at week 6, and declined slowly there after to 1.0 +/- 1.1 at week 54. The serum osteocalcin (OC) z-score showed a temporal pattern similar to that of ALP. It was initially 2.2 +/- 0.2, increased to 4.0 +/- 0.6 at week 6, and later declined slowly to 0.7 +/- 0.5 at week 54. The failure of the markers of bone turnover to normalize after 1 yr of therapy indicates an on-going high rate of bone turnover despite the attained euthyroidism. The uncoupling index (UI = z-score of U.NTx minus z-score of OC) was 7.1 +/- 1.2 before treatment, indicating unbalanced bone turnover in favor of bone resorption, and fell close to zero at week 30 of treatment. Pretreatment plasma PTH was suppressed slightly to 2.17 +/- 0.47 pmol/L and rose significantly during treatment, reaching a plateau of 5.27 +/- 0.78 at week 12. In all postmenopausal women PTH increased above the upper limit of normal (6.84 pmol/L). Pretreatment serum 25-hydroxyvitamin D was normal and remained unchanged during treatment, whereas 1,25-dihydroxyvitamin D was initially subnormal and rose to normal level after treatment. There was a significant positive linear correlation between PTH and U.NTx after week 12. PTH was also significantly correlated with ALP, but not with OC. ALP and OC were significantly correlated. A significant positive correlation was found between T3 and U.NTx, and a negative correlation was found between T3 and each of the formation markers (ALP and OC) over the 0- to 12-week interval. The latter correlations and the very high pretreatment UI indicate some inhibitory effect of the high thyroid hormone levels on the osteoblasts. The marked and sustained elevation of PTH, more pronounced in the postmenopausal women, during the first year of treatment of hyperthyroidism seems to play a pivotal role in maintaining a relatively high rate of bone turnover despite euthyroidism, and in the conservation of calcium by reducing renal calcium excretion and increasing calcium absorption (via 1,25-dihydroxyvitamin D). It may also account in part for the additional rise of the bone formation markers by an anabolic effect on the osteoblasts. Endogenous PTH may be important in the restoration of bone mineral density of treated hyperthyroid patients.     

Alendronate in primary hyperparathyroidism: a double-blind, randomized, placebo-controlled trial

Primary hyperparathyroidism (PHPT) is often associated with reduced bone mineral density (BMD). A randomized, double-blind, placebo-controlled trial was conducted to determine whether alendronate (ALN), 10 mg daily, maintains or improves BMD in patients with PHPT. Eligible patients had asymptomatic PHPT and did not meet surgical guidelines or refused surgery. Forty-four patients randomized to placebo or active treatment arms were stratified for gender. At 12 months, patients taking placebo crossed over to active treatment. All patients were on active treatment in yr 2. The primary outcome index, BMD, at the lumbar spine (LS), femoral neck, total hip, and distal one third radius was measured every 6 months by dual-energy x-ray absorptiometry. Calcium, phosphorous, PTH, bone-specific alkaline phosphatase (BSAP) activity, urinary calcium, and urinary N-telopeptide (NTX) excretion were monitored every 3 months. Treatment with alendronate over 2 yr was associated with a significant (6.85%; micro(d) = 0.052; +/-0.94% se; P < 0.001) increase in LS BMD in comparison with baseline. Total hip BMD increased significantly at 12 months with alendronate by 4.01% (micro(d) = 0.027; +/-0.77% se; P < 0.001) from baseline and remained stable over the next 12 months of therapy. BMD at the one third radius site did not show any statistically significant change in the alendronate-treated group at 12 or 24 months of therapy. At 24 months, the alendronate-treated group showed a 3.67% (micro(d) = 0.022; +/-1.63% se; P = 0.038) gain in bone density at the femoral neck site in comparison with baseline. The placebo group, when crossed over to alendronate at 12 months, showed a significant change of 4.1% (micro(d) = 0.034; +/-1.12% se; P = 0.003) in the LS BMD and 1.7% (micro(d) = 0.012; +/-0.81% se; P = 0.009) at the total hip site in comparison with baseline. There was no statistically significant change seen in the placebo group at 12 months at any BMD site and no significant change at 24 months for the distal one third radius or femoral neck sites. Alendronate was associated with marked reductions in bone turnover markers with rapid decreases in urinary NTX excretion by 66% (micro(d) = -60.27; +/-13.5% se; P < 0.001) at 3 months and decreases in BSAP by 49% at 6 months (micro(d) = -15.98; +/-6.32% se; P < 0.001) and by 53% at 9 and 12 months (micro(d) = -17.11; +/-7.85% se; P < 0.001; micro(d) = -17.36; +/-6.96% se; P < 0.001, respectively) of therapy. In the placebo group, NTX and BSAP levels remained elevated. Serum calcium (total and ionized), PTH, and urine calcium did not change with alendronate therapy. In PHPT, alendronate significantly increases BMD at the LS at 12 and 24 months from baseline values. Significant reductions in bone turnover occur with stable serum calcium and PTH levels. Alendronate may be a useful alternative to parathyroidectomy in asymptomatic PHPT among those with low BMD.     

Ketoconazole-induced reduction in serum 1,25-dihydroxyvitamin D

The antimycotic agent ketoconazole is known to inhibit several cytochrome P450-dependent enzymes involved in the biosynthesis of steroid hormones from cholesterol. Since 1,25-dihydroxyvitamin D is also a sterol synthesized by cytochrome P450-dependent enzymes, we assessed whether ketoconazole would lower serum 1,25-dihydroxyvitamin D levels. In nine normal men, administration of ketoconazole for 1 week in doses of 300-1200 mg/day led to a dose-dependent reduction in serum 1,25-dihydroxyvitamin D levels (r = -0.64; P less than 0.001). At the highest dose taken by each man (1200 mg/day in six, 900 mg/day in one, and 600 mg/day in two), serum levels of 1,25-dihydroxyvitamin D fell significantly compared to baseline [14 +/- 1 (+/- SEM) vs. 39 +/- 3 pg/ml; P less than 0.001), but there was no change in serum levels of 25-hydroxyvitamin D, PTH, calcium, phosphate, or alkaline phosphatase. Ketoconazole may be potentially useful in exploring the pathogenetic role of 1,25-dihydroxyvitamin D in disorders of calcium metabolism and in treatment of patients with hypercalcemic disorders or renal stone disease.     

Hormonal regulation of osteocalcin plasma production and clearance in sheep

The plasma osteocalcin (OC) concentration correlates with histological parameters of bone formation and has been used as an index of osteoblast activity. Although the rate of OC synthesis is likely to be a major determinant of the plasma OC level, the contribution of other processes, for example, OC plasma clearance, should be evaluated if changes in the plasma OC concentration are to be interpreted meaningfully. We have treated oophorectomized sheep with 1,25-dihydroxyvitamin D3 (1,25-(OH)2D3) and glucocorticoids to alter the plasma OC concentration and used an 125I-OC infusion method to measure changes in the OC plasma clearance (PCR) and to calculate production (PPR) rates. 1,25-(OH)2D3 (8 micrograms/day by iv injection) increased plasma OC levels from 15.8 +/- 2.1 to 52.7 +/- 5.3 ng/ml (n = 10; P less than 0.0005) and the OC PPR from 0.9 +/- 0.08 to 2.6 +/- 0.17 mg/day (n = 5, P less than 0.0005). There was no effect on the OC PCR. Both triamcinolone acetonide (TA) and cortisol, administered by iv infusion, decreased the plasma OC concentration to less than 4 ng/ml and the OC PPR to less than 0.4 mg/day when infused in the doses 0.05-1.0 mg/h (TA) and 2.0 mg/h (cortisol). TA (0.05 mg/h) decreased plasma OC to an undetectable level within 24 h. The effect of cortisol infusion on the plasma OC level and the OC PPR was dose dependent in the range 0.2-2.0 mg/h. TA, infused at the rates of 0.25 and 1.0 mg/h, increased the OC PCR from 3.0 +/- 0.10 to 4.0 liters/h (P less than 0.005; n = 4) and from 2.6 +/- 0.10 to 3.9 +/- 0.10 liters/h (P less than 0.01; n = 3), respectively. It is concluded that the OC PPR and the plasma OC concentration in sheep are responsive to treatment with 1,25-(OH)2D3 and glucocorticoids. Although the changes in plasma OC concentration were attributable largely to effects on OC production, high infusion rates of the glucocorticoid TA led to a highly significant increase in OC plasma clearance. These findings suggest that alterations in OC clearance may contribute to the changes in plasma OC levels seen in some disease states, for example, glucocorticoid excess.     

1,25-Dihydroxyvitamin D maintains bone cell activity, and parathyroid hormone modulates bone cell number in dogs

The singular and combined effects of 1,25-dihydroxyvitamin D [1,25-(OH)2D] and PTH on bone were evaluated in a long term in vivo study in dogs. Dogs were rendered deficient in 1,25-(OH)2D and PTH by five sixths nephrectomy and parathyroidectomy. A control group was sham operated. Various combinations in status of 1,25-(OH)2D and PTH were produced by daily sc injections of 1,25-(OH)2D (1.25) and/or continuous infusion of 1-34 bovine PTH. These were 1.25+/PTH+, 1.25-/PTH-, 1.25+/PTH-, 1.25-/PTH+. Serum calcium levels were kept in the normal range by the administration of one or two of the hormones or by oral supplementation of calcium lactate. Histomorphometric evaluation of static and dynamic parameters of bone after 8 months of experimental observation revealed that deficiency in 1,25-(OH)2D and PTH resulted in decreased number and activity of bone-forming and resorbing cells. Administration of 1,25-(OH)2D increased the activity but not the number of bone cells. In contrast, administration of PTH increased the number but not the activity of bone cells. Tissue level activity was decreased when one or both hormones were deficient, and normal tissue level activity was found only when both hormones were given. These data are relevant for understanding and management of diseases with perturbations in vitamin D and/or PTH.     

The effect of infliximab on bone metabolism markers in patients with rheumatoid arthritis

OBJECTIVE: The aim of this study was to evaluate urinary excretion of N-telopeptide of type I collagen (NTX) and deoxypyridinoline (DPD), markers of bone resorption, and serum bone alkaline phosphatase (BAP) level, a marker of bone formation and an early marker of osteoblast differentiation, in patients with rheumatoid arthritis (RA) treated with infliximab. METHODS: Seventeen male and female patients (age 60.7+/-2.53 yr; mean disease duration 12.9+/-3.01 yr; Steinbrocker's class II-IV) with RA, diagnosed according to the criteria of the American College of Rheumatology (ACR), took part in the study between March 2003 and January 2005. None of the patients had a history of oestrogen replacement therapy. All patients were treated with infliximab combined with methotrexate. Infliximab was infused intravenously at 3 mg/kg at baseline, 2 and 6 weeks, then every 8 weeks. To evaluate disease activity, ESR, CRP, the numbers of swollen and tender joints, modified Stanford Health Assessment Questionnaire (mHAQ) score and ACR score were measured. Levels of NTX and DPD in urine and BAP in serum were measured in all patients. RESULTS: ESR, CRP, the number of swollen joints and tender joints, and mHAQ score had decreased significantly 6 weeks after initial treatment and were still low 6 months after initial treatment. NTX levels had decreased significantly 6 weeks after the initial treatment and were still low 6 months after initial treatment. DPD levels had decreased 6 months after initial infusion. Mean serum BAP level did not differ significantly among the three time points. NTX levels were statistically corresponding with the number of swollen joints and mHAQ scores. DPD levels were statistically lower corresponding with ESR. CONCLUSION: Infliximab therapy may inhibit generalized bone loss in patients with RA. NTX is a more sensitive marker than DPD.     

Mineral metabolism in Turner's syndrome: evidence for impaired renal vitamin D metabolism and normal osteoblast function.

We examined intact PTH and 1,25-dihydroxyvitamin D [1,25-(OH)2D] in both baseline and dynamic conditions (low calcium diet) in 14 patients with Turner's syndrome (mean age, 12.6 +/- 5.9 yr; range, 4.2-21.0 yr) and bone demineralization as well as in a control group of 15 healthy girls (mean age, 12.8 +/- 5.6 yr; range, 3.8-22.7 yr). In both groups we also measured osteocalcin serum levels in response to oral 1,25-(OH)2D3 administration (1.8 micrograms/m2/daily for 6 days) to assess osteoblast function. The low calcium diet decreased ionized calcium (Ca2+) levels and elevated PTH values to the same extent in both patients (Ca2+, -8.40 +/- 3.78%; intact PTH, +47.88 +/- 13.24%) and controls (Ca2+, -9.09 +/- 3.25%; intact PTH, +52.77 +/- 10.52%; P = NS vs. patients). While controls showed an increment in their serum 1,25-(OH)2D levels (+52.15 +/- 8.95%), patients did not (+10.93 +/- 4.71%; P = NS vs. baseline; P < 0.001 vs. controls). 1,25-(OH)2D3 administration caused a rise in the serum osteocalcin levels in a similar fashion in both groups (peak values: patients, +35.38 +/- 7.20%; controls, +34.09 +/- 7.98%; P = NS). We conclude that in patients with Turner's syndrome there is an altered renal vitamin D metabolism in response to physiological stimulus, while osteoblast function in response to 1,25-(OH)2D3 administration is not affected.     

Serum bone gla protein and the vitamin D endocrine system in the oophorectomized rat

Because of the importance of estrogen in osteoporosis, the effects of decreased estrogen production using sensitive measurements of bone mineral metabolism were studied in oophorectomized rats. Serum levels of ionized calcium, bone gla protein (BGP), vitamin D metabolites (25-hydroxyvitamin D and 1,25-dihydroxyvitamin D), and estradiol were measured before and serially for 6 weeks after oophorectomy in the rat. In addition, static and dynamic indices of bone histomorphometry were determined after double tetracycline labeling. Fifty Sprague-Dawley female rats (approximately 250 g) were studied. Twenty-five rats underwent oophorectomy (O), while the remaining rats were sham operated. Estrogen deficiency was noted in the O group within a week after surgery (estradiol, 2.45 +/- 0.78 vs. 27.9 +/- 4.15 pg/ml; P less than 0.05). Serum ionized calcium levels, 25-hydroxyvitamin D, 1,25-dihydroxyvitamin D, and PTH levels did not differ between the two groups during the length of the study. Serum BGP levels were the same in both groups until the second week postoophorectomy, after which BGP remained significantly elevated in the O animals (121.7 +/- 5.95 vs. 76.7 +/- 3.87; P less than 0.001). Bone histomorphometry revealed increased osteoid volume (4.4 +/- 0.9% vs. 2.3 +/- 0.7%), osteoblast surface (26.5 +/- 2.4% vs. 3.2 +/- 1.2%), tetracycline surface (18.9 +/- 4.1% vs. 6.8 +/- 2.2%), as well as osteoclast surface (8.2 +/- 1.4% vs. 2.5 +/- 2%) in all O animals compared with those in the sham-operated group. These data indicate that oophorectomy and decreased estrogen result in increased bone turnover with elevated BGP levels. The marked BGP elevation within 2 weeks postoophorectomy suggests that estrogen withdrawal results in rapid altered bone mineral metabolism. The lack of concomitant increase in circulating PTH levels suggests that other factors may be mediating the bone loss following surgical oophorectomy.     

Changes in biochemical markers of osteoblastic activity during the menstrual cycle

Serum levels of osteocalcin [OC; bone Gla protein (BGP)] and bone alkaline phosphatase (B-AP) are both correlated to osteoblastic activity, which may be regulated by several hormones, including estrogen, 1,25-dihydroxyvitamin D3 [1,25-(OH)2D3], and PTH. Estrogen shows reproducible variations during the menstrual cycle, while available data on variations in serum 1,25-(OH)2D3 and serum immunoreactive PTH show midcyclic increases or no changes. In the present study we evaluated osteoblastic activity by measuring serum OC and B-AP during the menstrual cycle in eight healthy women, aged 20-47 yr. The cycles were synchronized by LH peaks, and follicular and luteal periods were normalized by lengths. Repeated measures analysis of variance showed that serum OC varied significantly (P less than 0.05), with highest levels during the luteal period. Although the same pattern was seen for serum B-AP, the variation just failed to reach significance (P less than 0.10), but the mean level was significantly higher during the luteal than during the follicular period (P less than 0.05). Gonadotropins and ovarian sex hormones showed significant variations. There were no significant changes in serum vitamin D-binding protein, serum total and free 1,25-(OH)2D3 index, or serum immunoreactive PTH-(1-84), but serum levels of somatomedin-C showed a significant variation, with the highest level during the luteal period (P less than 0.05). Blood levels and urinary excretion of minerals exhibited no significant variations. Cross-correlation studies between OC and estradiol showed the highest correlation coefficient, when OC was lagged about 7 days after estradiol (r = 0.69; P less than 0.05). Moreover, a high correlation was found between OC and somatomedin-C when matched at concurrent time points (r = 0.76; P less than 0.01). No significant correlations were found between the other calcium-regulating hormones and OC when matched at concurrent time points. In conclusion, we found a significant effect of the menstrual cycle on the serum levels of two osteoblastic bone markers, OC and B-AP. The changes indicated that osteoblastic activity is higher during the luteal period. However, whether the changes are caused by direct or indirect effects of the fluctuations in calciotropic hormones is still unresolved.     

Effect of experimental human magnesium depletion on parathyroid hormone secretion and 1,25-dihydroxyvitamin D metabolism

Magnesium (Mg) deficiency in man may result in hypocalcemia, impaired PTH secretion, and low serum concentrations of 1,25-dihydroxyvitamin D [1,25-(OH)2D]. To determine whether these changes are due to selective Mg depletion, we studied 26 normal subjects before and after a 3-week low Mg (less than 1 meq/day) diet. This diet induced Mg deficiency, as demonstrated by a fall in pre- to postdiet serum Mg levels from 0.80 +/- 0.01 to 0.61 +/- 0.02 mmol/L (P less than 0.001), an increase in Mg retention from 11 +/- 4% to 62 +/- 4% (P less than 0.001), and a fall in red blood cell free Mg2+ from 205 +/- 10 to 162 +/- 7 microM (P less than 0.001). Serum calcium (Ca) fell significantly from 2.36 +/- 0.02 to 2.31 +/- 0.03 mmol/L (P less than 0.05), and serum 1,25-(OH)2D fell from 55 +/- 4 to 43 +/- 3 pmol/L (P less than 0.05). PTH secretion was impaired, as demonstrated by a fall or no change in serum PTH in 20 of 26 subjects despite a fall in the serum Ca and Mg. In addition, an iv injection of Mg in eight subjects after the diet resulted in a significant rise in PTH from 15 +/- 2 to 19 +/- 2 ng/L (P less than 0.01), whereas a similar injection given to six of the subjects before the diet resulted in a significant fall from 28 +/- 5 to 13 +/- 3 ng/L (P less than 0.001). The fall in serum 1,25-(OH)2D may be due to both the decrease in PTH secretion and a renal resistance to PTH. PTH resistance was suggested, as no increase in serum 1,25-(OH)2D was observed in the six subjects in which the PTH concentration rose by mean of 68% after the diet. Also, the rise in serum 1,25-(OH)2D after a 6-h human PTH-(1-34) infusion was significantly less after Mg deprivation. The results demonstrate that mild Mg depletion can impair mineral homeostasis and may be implicated as risk factor for osteoporosis in disorders such as chronic alcoholism and diabetes mellitus, in which Mg deficiency and osteoporosis are both common.     

Evaluation of bone and mineral metabolism in patients recently diagnosed with leprosy

This study was conducted to evaluate patients recently diagnosed with the tuberculoid and lepromatous forms of leprosy for bone mass, bone remodeling, and hormones related to mineral control. Eleven normal control individuals (CG) and 12 patients with leprosy (LG) matched for physical characteristics were submitted to evaluation of bone mass density (BMD) and to the determination of serum levels of PTH, 25-hydroxyvitamin D [25(OH)D], testosterone, LH, FSH, osteocalcin (OC), and urinary levels of deoxypyridinoline (DPD). The T score of lumbar spine and total radius (mean +/- SD) were significantly lower in leprosy patients (L1-L4: CG = -0.7 +/- 1.5 vs LG = -1.8 +/- 1.0 SD, P < 0.04, and total radius: CG = -1.43 +/- 0.6 vs LG = -2.1 +/- 0.8 SD, P <0.02), whereas no significant differences were observed in total hip or femoral neck T score. However, at all sites, the rate of low bone mass (T score < -1.0) was higher in LG (femoral neck: CG = 18% vs LG = 50%, total hip: CG = 27% vs LG = 42%). There was a significant difference in albumin and PTH levels between groups but not in serum 25(OH)D and OC levels or urinary DPD levels. The present results indicate that bone mass loss is an early event in leprosy patients and frequently is already present at diagnosis. Its etiopathogenesis is multifactorial, and further studies are needed to determine the most efficient way to prevent fractures in this condition. The data obtained in the present study need confirmation by the evaluation of a larger sample.     

Administration of pharmacological amounts of 25(s),26-dihydroxyvitamin D3 reduces serum 1,25-dihydroxyvitamin D3 levels in rats

Pharmacological amounts of 25(s),26-dihydroxyvitamin D3 were administered to normal, vitamin D-replete rats in order to assess its pharmacological activity. Treatment with 25(s),26-dihydroxyvitamin D3 (20 micrograms/day for 1 week) caused a marked and significant fall in the circulating concentration of 1,25-dihydroxyvitamin D (16 +/- 5 SEM vs. 28 +/- 4 pg/ml, P = 0.02). This reduction of 1,25-dihydroxyvitamin D was dependent on the dose of 25,26-dihydroxyvitamin D3 administered since a 5 micrograms/day dosing regimen failed to alter serum 1,25-dihydroxyvitamin D levels. Despite the 25-66% reduction in circulating 1,25-dihydroxyvitamin D concentration produced by 25,26-dihydroxyvitamin D3 therapy, serum calcium and intestinal calcium absorption remained normal. These results suggested that 25,26-dihydroxyvitamin D has a weak agonist action or that a further metabolite that stimulates bone calcium resorption and/or intestinal calcium absorption is formed. Rats predosed with 25,26-dihydroxyvitamin D3 (20 micrograms/day) for 4 days and subsequently dosed with both 1,25-dihydroxyvitamin D3 (0.15 micrograms/day) and 25,26-dihydroxyvitamin D3 for an additional 3 days, demonstrated serum 1,25-dihydroxyvitamin D levels significantly higher than that found for control rats (47 +/- 5 vs. 25 +/- 4 pg/ml, P less than 0.001) but significantly reduced from the value observed for rats receiving only 1,25-dihydroxyvitamin D3 (47 +/- 5 vs. 187 +/- 38 pg/ml, P less than 0.001). These results suggest that 25,26-dihydroxyvitamin D3 has a previously unrecognized action in affecting the metabolism of 1,25-dihydroxyvitamin D3.     

A single luteinizing hormone determination 2 hours after depot leuprolide is useful for therapy monitoring of gonadotropin-dependent precocious puberty in girls

Long-acting GnRH analogs represent the standard treatment for gonadotropin-dependent precocious puberty. The aim of this study was to determine the hormonal parameters for monitoring the adequacy of depot leuprolide acetate treatment in girls with clinical and hormonal diagnosis of gonadotropin-dependent precocious puberty. Eighteen girls were treated monthly with 3.75 mg depot leuprolide acetate. Adequate hypothalamic-pituitary-gonadal axis suppression during treatment was achieved in 16 of the 18 girls according to the clinical parameters and prepubertal LH levels. In these 16 well-controlled girls, the LH peak after a classical GnRH test was compared with a single LH measurement obtained 2 h after depot leuprolide acetate administration before and during GnRH analog treatment. Before therapy, the mean +/- sd LH peak after a classical GnRH test was 18.4 +/- 11.2 IU/liter (ranging from 7-41.5 IU/liter), and it was 22.6 +/- 8.3 IU/liter 2 h after the first depot leuprolide dose (ranging from 10-35.3 IU/liter). During therapy, the mean +/- sd of LH peak after classical GnRH test was 1.4 +/- 0.6 IU/liter (ranging from <0.6 to 2.3 IU/liter), and it was 2.7 +/- 1.9 IU/liter (ranging from 0.7-6.6 IU/liter) 2 h after depot leuprolide. The LH peak after a classical GnRH test and that 2 h after depot leuprolide administration correlate significantly before and during treatment. In conclusion, we established the LH cut-off values for an adequate depot leuprolide therapy as an LH peak below 2.3 IU/liter after a classical GnRH test or below 6.6 IU/liter 2 h after depot leuprolide. The latter measurement may replace the classical GnRH test as a reliable and convenient tool for monitoring therapy in female gonadotropin-dependent precocious puberty.     

The effect of short-term treatment with micronized estradiol on bone turnover and gonadotrophins in older men

Evidence for the role of estrogen in male bone metabolism has been confirmed by studies on a man with a genetic defect in the estrogen receptor as well as men with aromatase defects. All exhibited tall stature, delayed epiphysial closure, decreased bone density and increased bone turnover. Estrogen is likely to affect bone turnover in men throughout life; therefore, we hypothesized that older men would show decreased bone resorption in response to estrogen therapy. To test our hypothesis, fourteen community-dwelling men with osteopenia of the femoral neck were treated for 9 weeks with micronized estradiol, 1 mg/d, a dose which is effective in postmenopausal women. Each subject served as his own control. Markers of bone resorption, N-terminal collagen crosslinks (NTX) and C-terminal collagen crosslinks (CTX) and markers of bone formation, osteocalcin (OC) and bone specific alkaline phosphatase (BSAP) were measured every 3 weeks during a 9-week treatment period and 9 weeks post-treatment. Sex hormones, gonadotrophins and calciotropic hormones were measured at baseline, 9 weeks on treatment and 9 weeks post- treatment. After 9 weeks of treatment, estradiol and estrone levels increased significantly by greater than 6-fold and 15-fold, respectively. SHBG levels increased significantly by 17%. Testosterone and free testosterone levels decreased significantly by 27% and 34%, respectively. Markers of bone resorption showed wide variation at baseline and while on treatment. There was no correlation between changes in bone markers and changes in estrogen levels. During treatment, 11 patients showed a decrease of NTX or CTX, but three showed an increase. These three and one other subject had high initial levels of FSH and LH, suggesting some degree of primary gonadal failure, which decreased during estrogen therapy. Thus, the change in NTX (and CTX) after 9 weeks of E2 treatment was correlated with initial FSH (r= -.66, p= .01) and LH (r= -.73, p= .003) values. In addition, the largest decrease in free testosterone at 9 weeks was correlated with the higher values for NTX, CTX and BAP (r=-0.66, -0.68, -0.70 respectively; p< or =.01 for each of the markers). Treatment was generally well tolerated. Side effects of treatment were minimal, and included breast tenderness and decreased libido which reversed after treatment. We conclude that it is feasible to give low dose estrogen to healthy older men, but that the effects on bone turnover are not consistent. Changes in central feedback and in endogenous sex hormone production may alter the response of bone turnover to exogenous estrogen in this population.     

Parathyroid hormone contributes to regulating milk calcium content and modulates neonatal bone formation cooperatively with calcium

To determine whether PTH and calcium (Ca) interact in neonatal bone formation, female lactating mice either heterozygous (PTH(+/-)) or homozygous (PTH(-/-)) for targeted deletion of the pth gene were fed either a normal (1% Ca, 0.6% phosphate) or high-Ca diet (2% Ca and 0.4% phosphate). Dietary effects on milk Ca content and Ca-regulating hormones were determined in dams, and the effects of milk content were assessed on bone turnover in 3-wk-old pups. On the normal diet, milk Ca and 1,25-dihydroxyvitamin D(3) levels were lower, but milk PTH-related protein levels were higher in the PTH(-/-) dams compared with the PTH(+/-) dams. On the high-Ca diet, milk Ca levels were higher, but milk 1,25-dihydroxyvitamin D(3) and PTH-related protein levels were lower in both PTH(+/-) and PTH(-/-) dams. In pups fed by PTH(-/-) dams compared with pups fed by PTH(+/-) dams on normal diets, bone mineral density, trabecular bone volume relative to tissue volume, and the number of osteoblasts were reduced in both PTH(+/-) (32.5 +/- 1.2 vs. 39.6 +/- 1.5 mg/cm(2), P < 0.05; 23.3 +/- 1.6 vs. 29.2 +/- 2.8%, P < 0.01; and 94.2 +/- 8.2 vs. 123.5 +/- 3.5/mm(2), P < 0.01, respectively) and PTH(-/-) (20.4 +/- 0.9 vs. 27.0 +/- 1.2 mg/mm(2), P < 0.05; 16.8 +/- 1.9 vs. 19.3 +/- 2.1%, P < 0.05; and 48.6 +/- 7.9 vs. 90.5 +/- 8.6/mm(2), P < 0.01, respectively) pups but were lower in the PTH(-/-) pups compared with the PTH(+/-) pups. In contrast, in pups fed by either PTH(+/-) or PTH(-/-) dams on the high-Ca diet, bone mineral density, bone volume/tissue volume, and osteoblast numbers were significantly higher, in both PTH(+/-) (50.5 +/- 1.7 vs. 58.7 +/- 2.0 mg/mm(2), P < 0.05; 37.9 +/- 5.2 vs. 46.1 +/- 5.1, P < 0.05; and 120.5 +/- 9.2 vs. 159.3 +/- 14.7/mm(2), P < 0.01, respectively) and PTH(-/-) (33.0 +/- 1.2 vs. 47.5 +/- 2.2 mg/mm(2), P < 0.001; 23.8 +/- 3.1 vs. 35.9 +/- 2.0, P < 0.05; and 78.7 +/- 10.1 vs. 99.8 +/- 13.6/mm(2), P < 0.05, respectively), and were highest in the PTH(+/-) pups fed by the PTH(+/-) dams on the high-Ca diet. These results indicate that PTH can modulate Ca content of milk, and that PTH and Ca can each exert cooperative roles on osteoblastic bone formation in the neonate.     

1,25-dihydroxyvitamin D suppresses circulating levels of parathyroid hormone in a patient with primary hyperparathyroidism and coexistent sarcoidosis

CONTEXT: PTH is excessively secreted to develop hypercalcemia and accelerate bone turnover in patients with primary hyperparathyroidism. PTH stimulates the production of 1,25-dihydroxyvitamin D [1,25(OH)2D] that in turn suppresses the synthesis of PTH in parathyroid cells. OBJECTIVE: The objective of the study was to clarify whether 1,25(OH)2D indeed inhibits circulating levels of PTH and influences bone turnover, even in a patient with primary hyperparathyroidism. DESIGN, SETTING, AND PATIENT: We evaluated PTH levels in a patient with primary hyperparathyroidism and coexistent sarcoidosis whose serum 1,25(OH)2D levels were independent of PTH. INTERVENTIONS AND MAIN OUTCOME MEASURES: The present case was treated with prednisolone before and after surgical resection of parathyroid adenoma, and Ca-regulating hormones and bone markers were measured. RESULTS: Serum Ca and PTH levels significantly decreased after parathyroid surgery, whereas serum 1,25(OH)2D levels remained high. Prednisolone administration promptly decreased serum 1,25(OH)2D levels and reciprocally increased PTH levels despite consistent serum Ca levels either before or after surgery. PTH levels were negatively correlated with serum 1,25(OH)2D levels before and after surgery. Urine N-telopeptides, serum osteocalcin, and bone-type alkaline phosphatase all decreased to physiological ranges after parathyroid surgery. CONCLUSIONS: These results suggest that 1,25(OH)2D indeed inhibits the production of PTH not to exacerbate hypercalcemia in a patient with primary hyperparathyroidism. Furthermore, PTH but not 1,25(OH)2D may primarily be involved in the stimulation of bone turnover.