High Turnover Bone Loss Research Articles

A Combination of Subtherapeutic Doses of Chemically Modified Doxycycline (CMT-8) and a Bisphosphonate (Clodronate) Inhibits Bone Loss in the Ovariectomized Rat A Dynamic Histomorphometric and Gene Expression Study

Recent studies have demonstrated that tetracyclines can reduce bone loss in the ovariectomized (OVX) rat model of osteoporosis. In the current study, a non-antimicrobial, chemically modified doxycycline (CMT-8), alone or in combination with a bisphosphonate (Clodronate), was evaluated in this model. Forty-two, 6month old, female rats were randomly assigned to the following groups, (6/ group): a) sham/vehicle, b) OVX/vehicle; c) OVX/1 mg/day CMT-8; d) OVX/2 mg/day CMT-8, e) OVX/1 mg/week Clodronate; and f) OVX/1 mg/day CMT-8 + 1 mg/week Clodronate, CMT-8 was administered by oral gavage, Clodronate injected S/C. Following sham surgery or OVX, the rats were treated for 90 days with CMT-8 or vehicle alone, injected at three different times with fluorochrome labels, the rats were sacrificed, and the tibiae excised for analysis by dynamic bone histomorphometry. Femurs were aseptically removed and analyzed for collagen, collagenase and osteopontin mRNAs by Northern and dot blot analysis. As expected, OVX decreased trabecular bone volume (BV/TV by 73.8% vs. sham p<.01), and also reduced trabecular thickness, numbers, and increased spacing. Bone loss in the OVX animals was partially prevented with either 2 mg/day CMT-8 or 1 mg/wk Clodronate (p<.01), while the 1 mg/day CMT-8 had no effect. Interestingly, the efficacy of the combination therapy of CMT-8 and Clodronate was significantly better than either treatment by itself, maintaining bone mass and structural indices at levels identical to sham values. OVX rats mRNA for collagen, collagenase and osteopontin were elevated indicating high-turnover bone loss. Only COMBO therapy significantly reduced the collagenase and osteopontin mRNA. In summary, CMT-8 mono-therapy (2 mg) alone partially inhibited bone loss in this animal model of osteoporosis. However, 1 mg/day (CMT-8) monotherapy had no effect on bone loss or bone mRNA levels and when combined with Clodronate, interacted to increase efficacy. Thus, a combination of a suboptimal dose of CMT-8 and a bisphosphonate appears to increase the amount of bone by suppressing resorption in a model of osteoporosis.

Pamidronate prevents bone loss associated with carrageenan arthritis by reducing resorptive activity but not recruitment of osteoclasts.

Carrageenan arthritis is associated with high-turnover bone loss. We sought to determine whether the bisphosphonate pamidronate can modify this effect of inflammatory arthritis. Sixty mature, New Zealand White rabbits were randomly assigned to five groups: normal; normal with a therapeutic dose of pamidronate (300 microg/kg/day, administered subcutaneously); arthritis (induced in the right tibiofemoral joint with 10 intraarticular carrageenan injections); arthritis with a therapeutic dose of pamidronate (300 microg/kg/day, subcutaneous); and arthritis with a low dose of pamidronate (7.5 microg/kg/day, subcutaneous). All animals received the fluorochrome calcein green (0.5 g/l/day) in drinking water ad libitum from days 21 to 49. Undecalcified, transverse sections of the distal femur were photographed or imaged to determine bone volume; new bone volume; resting, eroded, osteoid, and osteoblast perimeters; and osteoclast number. Results were evaluated with analysis of variance and pairwise Bonferroni's tests. In trabecular bone adjacent to the joint affected by carrageenan arthritis, resting perimeter was substantially reduced compared with normal joints, and primary indices of osteoblast and osteoclast activity were abnormally high (p < 0.001). Daily treatment with a therapeutic dose of pamidronate (300 microg/kg/day, subcutaneous) during the induction of arthritis significantly decreased new bone volume, osteoid perimeter, and osteoblast perimeter compared with the untreated arthritis group (p < 0.001). Osteoclast number and eroded perimeter remained abnormally high despite treatment with pamidronate. The concomitant increase of bone volume and these osteoclast indices show that pamidronate prevents bone loss in this model of experimental inflammatory arthritis by inhibiting the resorptive activity, but not the formation or recruitment, of osteoclasts. These findings are relevant to the use of bisphosphonates in the treatment of rheumatoid arthritis.

Effects of cyclosporin A on sex hormone and estrogen receptor in male rat with special reference to cyclosporin A-induced osteoporosis.

The mechanisms of high turnover bone loss induced by Cyclosporin A (CsA) are not clearly understood. Deficiencies in sex hormones result in high turnover osteoporosis, and not only androgen but also estrogen plays an important role in maintaining bone mass in men. To study whether or not there are any changes in the levels of sex hormones, aromatization, and the expression of estrogen receptors in CsA-induced osteoporosis, we treated 39 rats with vehicle, low-dose CsA (5 mg/kg) and high dose CsA (15 mg/kg) for 28 days, and measured sex hormone levels by radioimmunoassay. Aromatase activities in ROS cells and 3T3-L1 cells were determined by measuring the conversion rate of 3H-androstenedione into 3H-estrone. ER and ER mRNA were measured by competitive RT-PCR in collected marrow cells and ROS cells. The levels of free testosterone in the serum in low-dose CsA-treated rats were unchanged, but the levels were significantly decreased in those treated with high-dose CsA as previously reported. The levels of total estradiol in the serum were significantly increased in the low-dose CsA-treated group (5 mg/kg) and were comparable to levels of the control group in the high-dose CsA-treated group (15 mg/kg). CsA increased the conversion of 3H-androstenedione to 3H-estrone in ROS cells, but not in 3T3-L1 cells. Meanwhile, CsA treatment did not change the rates of ER or ER mRNA expression in ROS cells or in collected bone marrow cells. In conclusion, CsA treatment decreased the level of free testosterone in the serum, but did not decrease the level of serum estradiol by enhancing aromatization. High-turnover osteoporosis induced by clinical dosage CsA treatment may not be caused by lowering the levels of circulating estrogen or by decreasing the expression of estrogen receptors.

High turnover bone disease following lung transplantation

Recipients of lung transplants are at very high risk for significant bone loss. Nevertheless, data on bone disease after lung transplantation are still limited. We, therefore, retrospectively evaluated the data of 33 patients surviving at least 1 year after lung transplantation (LTx) who were seen in our outpatient clinic for osteologic evaluation. Results of clinical evaluations, radiographs, and dual-energy X-ray absorptiometry (DXA) were related to each other, to clinical variables, and to serum levels of osteocalcin, parathyroid hormone (PTH), and 25-hydroxyvitamin D: 14 of 33 patients (42%) had vertebral fractures, 9 of whom were diagnosed within 2 years after transplantation. Bone mineral density values (DXA) were markedly decreased and predictive of compression fractures. 25-Hydroxyvitamin D levels were low in 13 patients (39%) and PTH was elevated in 7 (21%). Despite corticosteroids and low 25-hydroxyvitamin D, serum osteocalcin was elevated in 12 patients (36%). This was only partially explained by hyperparathyroidism, low sex hormones, and impaired renal function, and may partly be caused by cyclosporin A. We thus conclude that severe symptomatic bone disease is common in lung transplant recipients and due to a complex situation including high turnover bone loss and hypovitaminosis D. DXA can be used to estimate fracture risk for individual patients.

T lymphocytes play a critical role in the development of cyclosporin A-induced osteopenia.

The T lymphocyte suppressor, cyclosporin A, has been shown to cause high turnover osteoporosis. We postulated that cyclosporin A may exert its effects via the T cell rather than direct activity on bone. In this study we administered cyclosporin A (15 mg/kg.day by gavage) to 11 10-week-old Rowett athymic nude rats and to 12 age-matched immunocompetent Sprague-Dawley rats. Placebo was administered to control groups (n = 12 for both). After 28 days of treatment, the Sprague-Dawley rats displayed high turnover bone loss, but the nude rats were largely unaffected by the drug. Sprague-Dawley treated rats had less than half the percent trabecular area of their controls as measured at the secondary spongiosa of the proximal tibial metaphysis (P < 0.001; strain by treatment, P = 0.007). The same pattern was evident for trabecular number, separation, and thickness (strain by treatment, P = 0.034, P = 0.001, and P = 0.021, respectively). Only the Sprague-Dawley rats had an elevated percent eroded perimeter and an elevated bone area referent bone formation rate (strain by treatment, P = 0.002 and P = 0.0003, respectively). Mass, glucose, ionized calcium, PTH, osteocalcin, 1,25-dihydroxyvitamin D, and creatinine all responded similarly to cyclosporin A regardless of strain. T Lymphocytes thus appear to be a prerequisite for the development of cyclosporin A-induced osteopenia.

Analysis of relationships between sex hormone dynamics and bone metabolism and changes in bone mass in surgically induced menopause.

A 3-year follow-up study was performed of bone metabolism and bone changes induced by surgical menopause as a consequence of hysterectomy and oophorectomy (OVX) is 52 nonmenopausal women. We investigated 22 bone parameters and determined seven bone indices as indicators of bone mineral content by dual energy X-ray absorptiometry (DXA), quantitative computed tomography (QCT), and microdensitometry (MD). The significant correlations between levels of sex hormones and/or bone parameters and bone indices demonstrated that marked sex-steroid deficiency after surgical menopause induced bone uncoupling during high bone turnover and subsequent rapid bone loss on the early period after OVX. Principal component analysis using correlation coefficients suggested a seven-loading-factor matrix composed of bone parameters and a two-loading-factor matrix composed of bone indices. Two groups of parameters--estradiol and estriol, and androstenedione together, and luteinizing hormone and follicle stimulating hormone together--indicated that the rate of bone loss was greater in the trabecular bone than in the cortical bone. Three other groups of parameters--urine calcium, urine hydroxyproline, and serum bone Gla-protein together; serum alkaline phosphatase, serum calcium, and 1,25-dihydroxycholecalciferol [1,25(OH)2D] together; and plasma tartrate-resistant acid phosphatase--indicated that bone uncoupling, with a prevalence of resorption over formation of bone, was greater in trabecular bone than is cortical bone and also that magnitude and rate of bone loss in the axial vertebrae surpassed those in the appendicular metacarpals after OVX. Two other groups of parameters, namely, trabecular bone mineral density (Dd) and bone mineral content (Dc), both measured by DXA, and bone mineral density (L2, L3), measured by QCT, together; and the cortical thickness index (MCI), cortical bone mineral density (sigmaGS/D), and the ratio of GSmin/max, measured by MD, indicated that the relative rates of bone reduction at the 3-year follow-up were greater in the axial vertebrae than in the appendicular metacarpals. Thus, bone change in the trabecular bone was associated with rapid loss during the early phase after OVX, whereas that in the cortical bone was slow during the late phase.

Attenuation of postmenopausal high turnover bone loss in patients with hypoparathyroidism

Attenuation of postmenopausal high turnover bone loss in patients with hypoparathyroidism

AzaThioprine alone is bone sparing and does not alter cyclosporin A-induced osteopenia in the rat

The immunosuppressant agent cyclosporin A (CsA) induces a high turnover osteopenic state, while the effect on bone of the antimetabolite azathioprine, a drug often used in conjunction with CsA in transplant patients, is less clear. This study was therefore designed to investigate the outcome of azathioprine administration, with reference to CsA, on bone mineral metabolism using the rat model. Four groups of 10-week-old male Sprague-Dawley rats (12 per group) were randomly allocated to receive by daily gavage for a 28-day period: (1) no treatment (control group); (2) azathioprine 1.5 mg/kg bw; (3) CsA 15 mg/kg bw; and (4) a combination of azathioprine and CsA, as described above. Rats were weighed and blood assayed serially for osteocalcin, ionized calcium, 1,25-dihydroxyvitamin D (1,25(OH)2VitD), and parathyroid hormone (PTH). Tibiae were removed following sacrifice on day 28 after double calcein labeling for histomorphometric analysis. Immunosuppressant groups were compared with nontreated control. We confirmed our previous findings that CsA induces a state of high turnover bone loss which is accompanied by a diminished gain in body weight (p < 0.01) and elevated serum osteocalcin (p < 0.001) and 1,25(OH)2VitD levels (p < 0.001). Azathioprine treatment alone did not alter ionized calcium, 1,25(OH)2VitD, or PTH levels. However, there was biochemical evidence of impaired osteoblastic activity as seen by decreased osteocalcin values on days 14 and 28 (p < 0.001). Azathioprine caused no loss of bone volume nor any deviation from the norm in mineral apposition rate, bone formation rate, or longitudinal bone growth. All three treatment groups showed an increased recruitment of osteoclasts to the bone surface.(ABSTRACT TRUNCATED AT 250 WORDS)

Thyroxine suppressive therapy decreases bone mineral density in post‐menopausal women

Hyperthyroidism is associated with increased bone turnover and decreased bone mass. This study aimed to evaluate the bone mineral density (BMD) of post-menopausal women on long-term thyroxine suppressive therapy. An age and sex-matched cross-sectional study. Thirty-four post-menopausal women with carcinoma of thyroid, post total thyroidectomy and 131I ablation, on L-T4 for 12.2 +/- 6.6 years (mean +/- SD). Controls were 34 age-matched healthy Southern Chinese women. Total body and regional BMDs were determined by dual-energy X-ray absorptiometry. Bone turnover was assessed by biochemical markers. In the thyroxine treated group, total body mineral content was significantly lower than the controls (1652 +/- 356 vs 1994 +/- 270 g mean +/- SD, P < 0.005). They also had lower BMDs in the lumbar spine, femoral neck, trochanter and Ward's triangle (0.75 +/- 0.15 vs 0.92 +/- 0.16 g/cm2, P < 0.005; 0.62 +/- 0.12 vs 0.70 +/- 0.12 g/cm2, P < 0.01; 0.55 +/- 0.14 vs 0.63 +/- 0.15 g/cm2, P < 0.001; 0.55 +/- 0.14 vs 0.63 +/- 0.14 g/cm2, P < 0.005 respectively.) The thyroxine treated group also had higher serum alkaline phosphatase and osteocalcin levels as well as urinary hydroxyproline excretion, suggesting that they had high turnover bone loss. However, the Z-scores of the various regional BMDs were correlated only with the serum osteocalcin level and showed no correlation with the serum thyroxine level or with the dosage or duration of thyroxine treatment. Long-term thyroxine suppressive therapy was associated with bone loss and preventive therapy may be indicated in these post-menopausal women at risk of osteoporosis.

The aminobisphosphonate alendronate inhibits bone loss induced by thyroid hormone in the rat comparison between effects on tibiae and vertebrae

The aims of this study were to develop a rat model of hyperthyroidism and to determine the efficacy of alendronate in the prevention of thyroid hormone-induced bone loss. Ten week-old Sprague-Dawley rats injected with thyroxine 250 μg/kg/day (+T4) or vehicle (−T4) were treated with alendronate (+ALN) or vehicle (−ALN) orally 0.5 mg/kg/day. After 3 weeks of treatment histomorphometric parameters of cancellous bone remodeling were assessed in the proximal tibia and in the first lumbar vertebra. In the secondary spongiosa of the tibia T4 treatment caused significant bone loss, associated with increased bone turnover; trabecu lar bone volume, trabecular thickness and trabecular number were significantly decreased. Osteoid and osteoclast surfaces increased in +T4/−ALN as compared to control. Alendronate prevented the increase in bone turnover and increased bone volume above control values without interfering with the recruitment of osteoclasts. These changes were not apparent in the vertebra. It is concluded that excess thyroid hormone in the rat induces high turnover bone loss in the tibia which can be prevented by alendronate through an inhibition of osteoclastic activity. The lack of effects of thyroid hormone on the vertebra may be ascribed to a lower rate of basal bone turnover at that site.

Salmon Calcitonin Prevents Cyclosporin-A-Induced High Turnover Bone Loss*

Cyclosporin-A (CsA) has greatly influenced the outcome of organ transplantation and has also been effective in the treatment of many autoimmune diseases. Unfortunately, it has deleterious effects on bone remodelling, causing a high turnover bone loss, with bone resorption exceeding bone formation. Salmon calcitonin (SCtn) has been shown to inhibit bone resorption in high turnover states such as Paget's disease and postmenopausal osteoporosis. In an attempt to attenuate the high turnover bone remodelling caused by CsA alone, we studied the bone mineral effects of CsA in combination with SCtn in male Sprague-Dawley rats. Group A (n = 20) received vehicle as control, group B (n = 20) received CsA (15 mg/kg BW) by daily gavage and SCtn vehicle sc, group C (n = 20) received SCtn (1.3 IU/kg BW) daily sc and CsA vehicle, and group D (n = 20) received a combination of CsA and Ctn daily, as described above. Rats were bled weekly for determination of circulating biochemical bone parameters. Eight rats from each group were killed on day 14 (short term), and the remaining rats were killed on day 28 (long term). Tibiae were removed for bone histomorphometry after death, which revealed a reduction of trabecular bone volume and an increase in osteoclast number induced by CsA alone. These changes were significantly attenuated by the combination of CsA and SCtn to resemble the histomorphometry of the control group. The inhibition of osteoclast number by SCtn is the most plausible mechanism by which the combination therapy attenuates the high turnover bone loss induced by CsA alone.