Tibial subchondral bone mineral density: sources of variability and reproducibility.

OBJECTIVES: It has been shown that subchondral bone mineral density (sBMD) measurement may be a relevant parameter of osteoarthritis (OA) progression. However, factors implicating the reproducibility and contributing to the variability of the measurement have not been fully described. Thus, the aim of this study was to explore the reproducibility of sBMD by Dual energy X-ray Absorptiometry (DXA) and to further examine its sources of variability. METHODS: In this study, short-term, intra and inter-observer reproducibility of sBMD was examined on knee images obtained on DXA scans. The influence of software (lumbar spine and forearm modes), knee positioning (flexion or extension), site and size of regions of interest (ROI) and use of rice, on both lateral and medial tibial sBMD, were assessed. Root mean square coefficient of variation (RMS CV) and least significant changes (LSC) were calculated. RESULTS: The short-term precision of sBMD ranged between 2.24% and 5.12% for RMS CV and between 0.053 and 0.135 g/cm(2) for LSC. Good intra-observer precision was found for knee flexion conditions whatever the software used (RMS CV ranging from 0.43 to 1.41%). The reproducibility was dependant from the ROI size (the ROI including joint space exhibiting better precision results than ROI including solely the subchondral plate). For a constant size of the ROI, the precision results were site-dependant. Inter-observer RMS CV results ranged from 0.59 to 5.01% according to ROI and software used. For the specific task of monitoring medial sBMD in the ROI including solely subchondral plate, forearm flexion condition produced the highest intra-observer and short-term precision (respectively RMS CV: 0.45% and 2.77%; LSC: 0.013 and 0.080 g/cm(2)). CONCLUSION: Taking account into the excellent precision of the sBMD measurements expressed as RMS CV with the protocol proposed in the present study, clinical application of these measurements might be envisaged.

Exercise and zoledronic acid on lipid profile and bone remodeling in ovariectomized rats: a paradoxical negative association?

Exercise (EXE) and amino-bisphosphonates (BP) are both considered as useful strategies in the prevention of post-menopausal bone loss. Exercise reduces lipid levels, and BP may induce increase in high-density lipoprotein cholesterol (HDL-C). We hypothesized that combined effects of BP and exercise would produce a better improvement of lipid profile. We studied the specific and combined effects of zoledronic acid (Z) and EXE on lipid profile and bone remodeling in mature ovariectomized (OVX) rats. Six-month old female rats were randomly assigned to either a sham-ovx group (n = 12) or one of four OVX groups (n = 12): vehicle-treated sedentary (OVX); OVX + EXE (OVX-E, running on a treadmill for 12 weeks); OVX + Z (20 microg/kg, i.v.), (OVX-Z); OVX + Z+EXE (OVX-ZE). Total cholesterol (TC), HDL-C and bone remodeling markers were measured at baseline and at the end of the study. We demonstrated that both Z and EXE prevented the increase in bone resorption resulting from OVX, and individually improved the atherosclerotic risk index. Therapy with Z resulted in significant increase (39.00 +/- 0.03 vs. 53.6 +/- 0.01 mg/dl; +37.4%, P < 0.05) in serum concentration of HDL-C and a non significant decrease in TC (135.30 +/- 0.03 vs. 144.80 +/- 0.05 mg/dl; -5.8%) in the OVX-Z group compared to the OVX group. Post-menopausal women have elevated risk of CVD and bone resorption, hence, these data ultimately demonstrate (except for the elevated ratio in the combined group) that exercise and zoledronic acid are useful in minimizing the impact of these two processes in women and the combination of the two may be clinically relevant.

Does exercise modify the effects of zoledronic acid on bone mass, microarchitecture, biomechanics, and turnover in ovariectomized rats?

Regular activity has effects on bone size, shape, and density, resulting in an increase in mechanical strength. The mechanism of action that underlies this improvement in bone strength is mainly linked to an increase in bone formation. Zoledronic acid (Z), in contrast, may prevent bone strength changes in ovariectomized (OVX) rodents by its potent antiresorptive effects. Based on these assumptions we hypothesized that combined effects of exercise (E) and Z may produce higher benefits on bone changes resulting from estrogen deficiency than either intervention alone. At 6 months of age, 60 female Wistar rats were OVX or sham operated (SH) and divided into five groups: SH, OVX, OVX-E, OVX-Z, and OVX-ZE. OVX rats were treated with a single IV injection of Z (20 microg/kg) or vehicle and submitted or not to treadmill exercise (15 m/min, 60 min/day, 5 days/week) for 12 weeks. Whole-body BMD and bone turnover markers were analyzed longitudinally. At sacrifice, femurs were removed. BMD by DXA, three-point bending test, and microCT were performed to study biomechanical and trabecular structure parameters, respectively. After 12 weeks, bone volume fraction decreased in OVX rats, whereas bone turnover rate, trabecular spacing, and structure model index increased compared with those in the SH group (P < 0.05). Zoledronic acid prevented the ovariectomy-induced trabecular bone loss and its subsequent trabecular microarchitectural deterioration. Treadmill exercise running was shown to preserve the bone strength and to induce bone turnover changes in favor of bone formation. However, the combined effects of zoledronic acid and running exercise applied simultaneously did not produce any synergetic or additive effects.

Assessment of trabecular bone microarchitecture by two different x-ray microcomputed tomographs: a comparative study of the rat distal tibia using Skyscan and Scanco devices.

Important aspects of modern skeletal research depend on the phenotypical characterization of trabecular bone microarchitecture as assessed by microcomputed tomography (micro-CT). Until now, however, there have been no studies which compare the two most commonly utilized micro-CT devices, namely, Skyscan and Scanco. The purpose of the current study was to examine the reproducibility and accuracy of these two micro-CT devices in comparison to traditional histomorphometry in ovariectomized rats treated with either propranolol, salbutamol, or vehicle. 6 month old female Wistar rats were ovariectomized (n = 48) or sham operated (n = 12). OVX rats were divided into four groups and then subcutaneously injected with propranolol 0.1 mg/kg/day, propranolol 20 mg/kg/day, salbutamol 3 mg/kg/day, or vehicle for 10 weeks. At sacrifice, the left tibial trabecular bone microarchitecture was analyzed using both the micro-CT Skyscan 1072 (ex vivo) and Scanco vivaCT40 (in vivo). Histomorphometric analysis was performed on the right proximal tibia. Comparisons between the different methods were performed using regression analysis, Bland-Altman, Passing-Bablock, and Mountain plots. Correlations were highly significant for all parameters measured between the two micro-CT instruments and were less significant between histomorphometry and micro-CT measurements taken from either the Skyscan or Scanco apparatus. Micro-CT overestimated bone volume compared to histomorphometry. In the ovariectomized rat model, the two micro-CT instruments revealed the same difference between groups whereas histomorphometry revealed only the difference which displayed the largest disparity between groups. In regards to the comparison between the two micro-CT devices, Mountain plot methods indicated that BV/TV, BS/BV, and TbSp were equivalent, whereas a systematic bias was observed for TbN and TbTh. The authors were also able to describe the routine method used to determine the threshold between the two micro-CT devices, which may help explain these differences. While some minor differences in the absolute values of the morphometry parameters exist between the micro-CT measurements from the Skyscan and Scanco instruments, both of these devices display a high degree of accuracy and reproducibility.

Doping dose of salbutamol and exercise training: impact on the skeleton of ovariectomized rats.

Previous studies in healthy rats have demonstrated a deleterious bone impact of beta-agonist treatment. The purpose of this study was to examine the trabecular and cortical effects of beta(2)-agonists at doping dose on treadmill exercising rats with estrogen deficiency. Adult female rats were ovariectomized (OVX; n = 44) or sham operated (n = 12). Then, OVX rats received a subcutaneous injection of salbutamol (SAB) or vehicle with (EXE) or without treadmill exercise for 10 wk. Bone mineral density (BMD) was analyzed by densitometry. Microcomputed tomography and histomorphometric analysis were performed to study trabecular bone structure and bone cell activities. After 10 wk, SAB rats presented a much more marked decrease of BMD and trabecular parameters. Exercise did not change the high level of bone resorption in OVX EXE SAB compared with OVX SAB group (both on COOH-terminal collagen cross-links and osteoclast number). These results confirm the deleterious effect of beta(2)-agonists on bone quantity (femoral BMD gain: OVX EXE, +6.8%, vs. OVX EXE SAB, -1.8%; P < 0.01) and quality (-8.0% of femoral trabecular thickness in OVX EXE SAB vs. OVX EXE), indicating that SAB suppresses the effect of EXE in OVX rats. Furthermore, we notice that the slight beneficial effect of exercise was mainly localized in the tibia. These findings indicate the presence of a bone alteration threshold below which there is no more alteration in structural bone quantity and quality. The negative effects of SAB on bone observed in this study in trained rats may indicate potential complications in doping female athletes with exercise-induced amenorrhea.

Doping dose of salbutamol and exercise: deleterious effect on cancellous and cortical bones in adult rats.

Animal studies suggest that bone remodeling is under beta-adrenergic control via the sympathetic nervous system. To our knowledge, the impact of beta-agonist substances, at doping doses, has not been studied in adult rats. The purpose of this study was to examine the effects of salbutamol injections with or without treadmill exercise on trabecular and cortical bone in adult rats. Adult (36 wk of age) female Wistar rats (n = 56) were treated with salbutamol (3 mg.kg(-1).day(-1) sc, 5 days/wk) or vehicle (sham) with or without subsequent treadmill exercise (13 m/min, 60 min/day, 5 days/wk) for 10 wk. Tibial and femoral bone mineral density was analyzed by dual-energy X-ray absorptiometry. Metaphysic trabecular bone structure was analyzed by micro-CT at the time of the animals' death. Bone cell activities were assessed histomorphometrically. After 10 wk, the increase in bone mineral density was less in salbutamol-treated than in sham rats (+3.3% vs. +12.4%, P < 0.05), and trabecular parameters were altered and bone resorption was increased in salbutamol-treated rats compared with controls. The negative effect on bone architecture in salbutamol-treated rats persisted, even with treadmill exercise. These results confirm the deleterious effect of beta(2)-agonists on bone mass during chronic treatment and describe its effects on bone mechanical properties in adult rats. Bone loss occurred independently of a salbutamol-induced anabolic effect on muscle mass and was equally severe in sedentary and exercising rats, despite a beneficial effect of exercise on the extrinsic and intrinsic energy to ultimate strain. These bone effects may have important consequences in athletes who use salbutamol as a doping substance.

Dose effects of propranolol on cancellous and cortical bone in ovariectomized adult rats.

Animal studies suggest that bone remodeling is under beta-adrenergic control via the sympathetic nervous system. The purpose of this study was to examine the preventive effect of different doses of nonspecific beta-blockers (propranolol) on trabecular and cortical bone envelopes in ovariectomized rats. Six-month-old female Wistar rats were ovariectomized (OVX, n = 60) or sham-operated (n = 15). Then, OVX rats were subcutaneously injected with 0.1 (n = 15), 5 (n = 15), or 20 (n = 15) mg/kg propranolol or vehicle (n = 15) for 10 weeks. Tibial and femoral bone mineral density (BMD) were analyzed longitudinally by dual-energy X-ray absorptiometry. At death, the left tibial metaphysis and L(4) vertebrae were removed, and microcomputed tomography (Skyscan 1072; Skyscan, Aartselaar, Belgium) was performed for trabecular bone structure investigation. Histomorphometry analysis was performed on the right proximal tibia to assess bone cell activities. After 10 weeks, OVX rats had decreased BMD and trabecular parameters and increased bone turnover, as well as cortical porosity compared with the sham group (p < 0.001). Bone architecture alteration was preserved by 0.1 mg/kg propranolol due to higher trabecular number and thickness (+50.35 and +6.81%, respectively, than OVX; p < 0.001) and lower cortical pore number (-52.38% than OVX; p < 0.001). Animals treated by 0.1 mg/kg propranolol had a lower osteoclast surface and a higher osteoblast activity compared with OVX. Animals treated by 20 mg of propranolol did not significantly differ from OVX rats. Animals treated by 5 mg of propranolol have been partially preserved from the ovariectomy. These results showed a dose effect of beta-blockers. The lower the dose of propranolol breeding, the better the preventive effect against ovariectomy.