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Mouse Densitometer Abstracts


SU359

Changes in Bone Density and Size Following PTH Treatment of Young GH/IGF-I Deficient and Normal Mice. L.R. Donahue, C.J. Rosen, and W.G. Beamer. The Jackson Laboratory, 600 Main St., Bar Harbor, ME.

PTH has been shown to be an effective anabolic agent when administered intermittently to older subjects. However, low bone density is often seen in young people with idiopathic or glucocorticoid induced osteoporosis, with osteopenia due to hormonal deficiencies such as growth hormone deficiency, or with renal disease. It is possible that PTH would be an effective treatment for these younger patients, particularly in an acute protocol designed to rapidly induce accretion of bone mass followed by maintenance therapy with an alternative treatment to preserve newly acquired bone. We are utilizing a mouse model of GH/IGF-I deficiency, ‘little’ (lit), that results in homozygous lit/lit mutants with reduced serum GH, low serum IGF-I and IGFBP-3, and low bone density to test this hypothesis. We have treated groups of lit/lit males and females, along with lit/+ normal littermates with 50ug/kg rat PTH(1-34) or saline for 6 weeks. Serum was assayed for IGF-I, density and size of femurs were determined by pQCT (Norland), and density of lumbar spines was assessed by PIXI DEXA (Lunar). All mice responded to PTH treatment with increased femoral cortical bone density, total lumbar density, and bone size indicated by cortical thickness, periosteal circumference and endosteal circumference. Values for all parameters approached, but did not reach mature levels, assessed at four months of age when mice reach peak bone density. Surprisingly, even the GH/IGF-I deficient lit/lit mice, whose serum IGF-I levels did not change, showed significant bone growth. Percent increases for measures that reached significance following PTH treatment compared to saline treated controls are shown below.

Measurement lit/lit male lit/lit female lit/+male lit/+female
IGF-I NS NS 9.5% 22%
Femoral cortical density 9% 10% 12% 17%
Cortical thickness 30% 30% 39% 42%
Periosteal circumference 61% 141% 61% 34%
Endosteal circumference 39% 114% 9% 20%
Lumbar density 12% 24% 14% 31%

These data show that young mice, both GH/IGF-I deficient and normal, are able to respond to PTH stimulation with increased bone density and size. Experiments are in progress to determine if increased bone density can be maintained when PTH is discontinued and if longer treatment with PTH would increase bone density and size beyond adult levels.

J Bone Miner Res, (1999) 14(Suppl. 1): S521

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SU 246

Precision and Accuracy of in Vivo Bone Mineral Measurements of Mouse Femurs Using DXA.
Tim R. Nagy, Ph.D.¹ and D. Wharton*². ¹Department of Nutritional Sciences, Univ. of Alabama at Birmingham, Birmingham, AL, ²Lunar Corp., Madison, WI.

We evaluated the precision and accuracy of dual-energy X-ray absorptiometry (DXA) on the measurement of femur bone mineral density (BMD) and bone mineral content (BMC) in mice. BMD and BMC were measured in vivo in C57BL/J6 mice using the Lunar PIXImus densitometer (software ver. 1.42.006.009). All scans were done using the total body mode with a resolution of 0.18x0.18mm. The analysis of BMD and BMC was performed on the right femur using a custom region of interest. Precision was determined by scanning six mice, five times each. The mean intra-individual coefficient of variation for BMD and BMC was 1.20 and 3.32%, respectively. Accuracy of DXA was determined by ashing the right femur of 27 mice and comparing the bone ash to BMC. DXA-derived BMC overestimated ash mass (-0.0066 ± 0.0006g; P<0.001). However, femur BMC was highly related to femur ash mass (r2=0.95; SEE=0.0006g), suggesting that DXA-derived femur BMC can be used to predict femur ash mass. These data show that DXA is a useful tool for the in vivo measurement of BMD and BMC in mice.

J Bone Miner Res, (1999) 14(Suppl. 1): S493

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SU083

The Anabolic Actions of Estrogen and PTH on the Murine Skeleton Are Additive. Abigail Samuels, Mark J. Perry, Rachel L. Gibson, Jonathan H. Tobias. Rheumatology Unit, Division of Medicine, University of Bristol, Bristol, United Knigdom.

Paratyhroid hormone (PTH) has previously been found to stimulate bone formation in several species. However, it is unclear whether a similar action also occurs in the mouse, and if so, whether this is additive with respect to estrogen’s anabolic action in this species. To address this question, we examined the effect of treatment with PTH and/or 17ß -estradiol (E2) by daily s.c. injection for 28 days, on skeletal indices of long bones in ten week old intact female mice. Our preliminary dose-response studies had suggested that PTH stimulates bone formation in this species in a biphasic manner, with maximal effect at 10µ g/kg/day, and this dose was used in the present study. Based on previous dose-response studies, we selected E2 40µ g/kg/day as a dose of estrogen associated with a partial anabolic response, to enable us to detect a possible additive effect of PTH. BMD (mg/cm2) was assessed in the distal femoral metaphysis by Lunar PIXI using dedicated small animal software. Trabecular width (Tb.W m m), cancellous mineralising surface (dLS/BS %), and bone formation rate (BFR/TV mm3/mm3) were measured on longitudinal sections of the proximal tibial metaphysis. Results show mean ± SEM.

Vehicle

PTH

E2

PTH +E2

BMD

120.4 ± 1.6

136.3 ± 2.0 **

147.9 ± 2.5**

158.0 ± 3.3 +

Tb.W

12.6 ± 2.1

20.9 ± 2.2 *

23.26 ± 2.0*

28.8 ± 3.6**

dLS/BS

21.6 ± 7.7

26.6 ± 4.1

30.1 ± 4.73

40.7 ± 4.2*

BFR/TV

0.41 ± 0.18

1.46 ± 0.42

1.99 ± 0.52

2.33 ± 0.39*

*p<0.05 vs Veh, **p<0.001 vs Veh, +p<0.05 vs E2 by one way ANOVA

Our results are consistent with a modest anabolic effect of PTH and E2 at the doses used in this study. In mice treated with both of these agents in combination, stimulation of bone formation appeared to be more marked than after treatment with either of these agents alone, suggesting that the anabolic action of these two agents may be additive. These findings raise the possibility that the success of combination therapy with estrogen and PTH in the treatment of postmenopausal osteoporosis may in part reflect an additive effect of the stimulatory action of these two agents on cancellous bone formation.

J Bone Miner Res, (1999) 14(Suppl. 1): S452

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SU240

Comparison of the Peripheral X-ray Imager with Standard DXA Measurement of Bone Mass in Rats and Rhesus Monkeys. R.J. Colman,¹ D. Settergren,*² A.K. Garber,*³ N. Binkley1,4. ¹Wisconsin Regional Primate Research Center, University of Wisconsin, Madison, WI, ²Lunar Corporation, Madison, WI, ³Department of Nutritional Sciences, University of Wisconsin, 4Institute on Aging, university of Wisconsin, Madison, WI.

Guidelines for preclinical evaluation of osteoporosis treatment agents require bone mass measurement in small and large animals, classically rats and non-human primates. In these animals, bone mineral content and density (BMC/BMD) can be determined using standard dual-energy x-ray absorptiometry (DXA) instruments with small animal software. Recently, DXA equipment developed to quantitate human bone mass (the Peripheral Instantaneous X-ray Imager [PIXI], Lunar Corp) has been adapted for use in small animals. We compared PIXI with standard DXA (DPX-L, Lunar Corp.) for the in vivo assessment of bone mass in the rat femur (n = 20) and monkey forearm (n = 9). DPX-L measurements were performed using small animal software version 1.0d, appendicular mode. PIXI determinations were performed using software version 1.42.7, whole body mode. PIXI scan time (approximately five minutes for femur or forearm) was shorter than DPX; (approximately 10 minutes for femur and 20 for radius). All scan analyses required manual region of interest (ROI) placement. Identical ROI’s were used with both instruments. In rats, the ROI included the proximal half of the femoral shaft. For the monkey radius, 2mm ultradistal and 5mm central ROIs were placed 10 and 47.5mm, respectively, from the most distal aspect of the styloid process. Precision (CV%) was determined by scanning five animals of both species five times each with repositioning between scans. DPX-L BMC and BMD precision was 2.4 and 1.6, respectively, for rat femur; 2.4 and 1.9, respectively, for the ultradistal radius; and 1.3 and 1.8, respectively, for the proximal radius. PIXI precision for BMC and BMD was 1.7 and 1.2, respectively, for rat femur, 1.1 and 0.8, respectively, for the ultradistal radius, and 0.4 and 0.8, respectively, for the proximal radius. DPX-L and PIXI results correlated for all sites analyzed (femur BMC r=0.82, BMD r=0.69; radius ultradistal BMC r=0.95, ultradistal BMD r=0.93, proximal BMC r=0.99, proximal BMD r=0.92). PIXI is highly precise, quick and well correlated with traditional DXA technology for in vivo bone mass assessment of the rat femur and monkey radius.

J Bone Miner Res, (1999) 14(Suppl. 1): S491

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PRECISION AND ACCURACY OF IN VIVO BONE MINERAL MEASUREMENTS OF MICE USING DUAL-ENERGY X-RAY ABSORPTIOMETRY. . T.R. Nagy, D. Wharton, M. Blaylock, and S. Powell. Univ. of Alabama at Birmingham, Birmingham, AL 35294 and Lunar Corp., Madison, WI 53713.

We evaluated the precision and accuracy of dual-energy X-ray absorptiometry (DXA) on the measurement of bone mineral content (BMC) and bone mineral density (BMD) in mice. BMC and BMD were measured using the Lunar PIXI Small Animal Bone Densitometer, software ver. 1.42.3. All scans were done using the total body mode (0.18x0.18mm) and the analysis of BMD and BMC was performed with a subcranial ROI. Precision was determined using 10 C57BL/J6 mice (21–35g), scanned 5 times each. Three mice were scanned without, and 7 were scanned with repositioning between scans. The mean intra-individual coefficient of variation for BMD and BMC was 1.20 and 2.21% without repositioning and 1.50 and 1.70% with repositioning. In the second experiment, accuracy was tested in 25 C57BL/J6 mice (10–25g) by comparing BMC to ash weight (body ashed at 600oC for 8 hrs). DXA-derived values underestimated ash weight (0.098 ± 0.005g; P<0.001). However, BMC was highly related with ash (r2 = 0.982; SEE = 0.015g), suggesting that DXA-derived values can be used to predict body ash weight. These data suggest that DXA may be a useful tool for the measurement of BMD and BMC in vivo. (Supported by NIH R01-DK54918 and Lunar Corp.)

FASEB Journal (1999) 13(5):A912

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RESTING METABOLIC RATE DOES NOT DECLINE WITH AGE IN LONG-LIVED NAKED MOLE-RATS. T.P. O'Connor and R. Buffenstein. City College of New York, CUNY. New York, NY 10031.

Metabolic rate is widely regarded as an important component and baro-meter of aging processes. Specifically, conventional mammalian models of aging such as rats and mice tend to show a progressive decline in resting metabolic rate (RMR) with advancing age. The decline in RMR is often associated with increased proportions of body fat and decreased lean mass. In this study, we examined age-related changes in RMR in a remarkable new model of mam-malian aging, the naked mole-rate (Heterocephalus glaber). Although they are the same size as mice, naked mole-rates are rodents that often live beyond 20 years of age in captivity. We measured RMR as oxygen consumption rate in 1, 10, and 20-year-old naked mole-rats. Body mass increased slightly, but not significantly (p 0.06) with age, and most of this increase took place between the ages of 1 and 10 y. Age did not have a significant effect on either total RMR (in units of ml oxygen consumed per hour) nor mass-specific RMR (p 0.7 in both cases). Preliminary evidence from a body composition study showed that, unlike other mammalian models, naked mole-rats do not show an increased percentage of body fat with advancing age. We have previously reported that the decline in age-related intestinal function is much more modest in mole-rats than in other rodents. Given their extreme longevity and novel physiology, naked mole-rats merit further study as a model of mammalian aging.

FASEB Journal (2000) Vol. 14, No. 4:A440

Lunar note: PIXImus was used to determine the body composition of these unique animals even though not credited here. Data from the PIXImus was displayed during the poster session. Interestingly, mole-rats are larger than the image area of the PIXImus, so they were imaged in two sections.

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EVALUATION OF A NEW DUAL-ENERGY X-RAY ABSORPTIOMETRY TECHNIQUE FOR IN VIVO BODY COMPOSITION MEASUREMENTS IN MICE. M. Punyanitya, R.L. Leibel, S.B. Heymsfield and C.N. Boozer. Obesity Research Center, St. Luke's-Roosevelt Hospital, Columbia University, NY, NY 10025 & 1Division of Molecular Genetics and Naomi Berrie Diabetes Center, College of Physicians and Surgeons, Columbia University, NY, NY 10032

Precise in vivo assessment of body composition in small animals is an important methodological goal. The present study evaluated the accuracy and precision of a new dual energy X-ray absorptiometry (DXA) technique for the measurement of lean tissue mass (LM) in mice. LM was measured using the Lunar PIXImus Small Animal Bone Densitometer (Madison, WI), software version 1.43. All scans were performed using the total body mode (0.18 x 0.18 mm) and the analysis of LM was performed using a total body Region of Interest. The within-day precision was assessed using 5 mice (mean ± std dev; 26.58 ± 5.03 g), scanned 10 times each, without repositioning between scans. The within-day coefficient of variation (CV) for LM in grams ranged from 0.95 to 2.59% (1.59 ± 0.62%). The between-day precision was assessed using the same 5 mice, scanned once each day, for 5 consecutive days, without repositioning between scans. The between-day CV for LM in grams ranged from 0.55 to 1.83% (1.05 ± 0.52%). For the same 5 mice, total body weight, as measured by an electronic balance, was compared to total DXA weight, the sum of bone mineral content, fat mass, and lean mass. Total measured weight and total DXA weight were highly and significantly correlated (r = 0.999, p 0.001, SEE = 0.093 g), with a slight offset of the intercept. These data suggest that the Lunar PIXImus DXA may be a reliable tool for the in vivo measurement of LM in mice.

FASEB (2000) Vol. 14, No. 4:A4

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PRECISION AND ACCURACY OF IN VIVO BONE MINERAL MEASUREMENTS OF MICE USING DUAL-ENERGY X-RAY ABSORPTIOMETRY. T.R. Nagy, D. Wharton, M. Blaylock, and S. Powell. Univ. of Alabama at Birmingham, Birmingham, AL 35294 and Lunar Corp., Madison, WI 53713.

We evaluated the precision and accuracy of dual-energy X-ray absorptiometry (DXA) on the measurement of bone mineral content (BMC) and bone mineral density (BMD) in mice. BMC and BMD were measured using the Lunar PIXI Small Animal Bone Densitometer, software ver. 1.42.3. All scans were done using the total body mode (0.18x0.18mm) and the analysis of BMD and BMC was performed with a subcranial ROI. Precision was determined using 10 C57BL/J6 mice (21-35g), scanned 5 times each. Three mice were scanned without, and 7 were scanned with repositioning between scans. The mean intra-individual coefficient of variation for BMD and BMC was 1.20 and 2.21% without repositioning and 1.50 and 1.70% with repositioning. In the second experiment, accuracy was tested in 25 C57BL/J6 mice (10-25g) by comparing BMC to ash weight (body ashed at 600oC for 8 hrs). DXA-derived values underestimated ash weight (0.098 ± 0.005g; P 0.001). However, BMC was highly related with ash (r2 = 0.982; SEE = 0.015g), suggesting that DXA-derived values can be used to predict body ash weight. These data suggest that DXA may be a useful tool for the measurement of BMD and BMC in vivo. (Supported by NIH R01-DK54918 and Lunar Corp.)

FASEB Journal (1999) 13(5):A912

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Bone Mineral Density in a Rat Model of Type 2 Diabetes. R.P. HAYES*, T.R. NAGY, F. RAHEMTULLA, P. BOUNELIS, C.W. PRINCE. University of Alabama at Birmingham, Birmingham, AL.

Altered bone homeostasis, often resulting in osteoporosis, is a significant problem in diabetes, which is an exclusion criterion for placement of dental implants. The cause for the disruption of bone homeostasis in diabetes is poorly understood. To investigate bone metabolism in diabetes, we did a pilot study using a novel rat model of Type 2 (non-insulin dependent) diabetes, the ZDF-GMI diabetic rat. Groups of two ZDF-GMI rats and one ZDF-lean control rat were sacrificed after an overnight fast at ages 11, 20 and 29 weeks. Blood glucose was measured and both tibias excised and analyzed for bone mineral density (BMD) and bone mineral content (BMC) by dual energy x-ray absorptiometry (DXA) using the Lunar PIXImusä densitometer (software version 1.43.036.007). BMD and BMC values from right and left tibias of the same rat were averaged. Data (body weight, blood glucose, BMD, BMC) were analyzed by 2-way ANOVA (age and genotype). Genotype had no effect on body weight but, as expected, did on blood glucose (diabetics 2-3X higher than controls, p0.05). Genotype had significant effects on BMD (p0.01) and BMC (p0.05). At 29 weeks of age, values for BMD and BMC for diabetic and control rats, respectively, were 0.134 vs. 0.169 g/cm2 and 0.259 vs. 0.396 g. These data suggest that prolonged elevated blood glucose leads to impaired mineralization of bone. This study also demonstrates the utility of the Lunar PIXImusä densitometer for making rapid (-5 min.) assessments of bone mineral content and density. Future studies will investigate glucose-induced alterations in bone extracellular matrix molecules as a potential cause for the reduction in bone mass seen in this model of diabetes. Supported by NIM grant P50 DE08328.

J Dent Res 79 (IADR Abstracts) 2000:565

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Validation of Body Composition Measurements of Mice Using DXA. Tim R. Nagy and Anne-Laure Clair. Dept. of Nutrition Sciences. Dept. of Nutrition Sciences, Univ. of Alabama at Birmingham.

Objective: To evaluate the precision and accuracy of dual-energy X-ray absorptiometry (DXA) for the measurement of bone mineral density (BMD), bone mineral content (BMC), fat, and soft-lean tissue (SLT) in mice. Methods: Twenty-five male C57BL/6J mice (6-11wks; 19-29g) were anesthetized and scanned three times (with repositioning between scans) using a Lunar Piximus densitometer (software ver. 1.42.006.010). Gravimetric and chemical extraction techniques (Soxhlet) were used as the criterion method for the determination of body composition and body ash was determined by burning at 600oC for 8 hrs.

Results: The mean intra-individual coefficient of variation (CV) for the repeated DXA analyses were: BMD, 0.85%; BMC, 1.61%; fat, 2.21%, and SLT, 0.85%. Accuracy was determined by comparing the DXA-derived data from the first scan with the chemical analysis data. DXA underestimated ash mass (0.24±0.01g, p 0.001) and SLT (0.59±0.05g, p 0.001), and overestimated fat mass (2.19±0.06g, p 0.001). DXA-derived values were used to predict chemical values. Ash and fat mass were predicted using DXA-derived values of BMC, SLT, and fat (ash, r2=0.93, SEE=0.11g; fat, r2=0.86, SEE=1.1g). Soft-lean tissue mass was predicted by DXA SLT (r2=0.99, SEE=0.46g).

Conclusions: These data show that DXA has excellent precision for measuring BMD, BMC, fat, and SLT. DXA-derived values were significantly different than chemical-derived values with DXA underestimating BMC and SLT, and overestimating SLT. However, because DXA values were strongly related to chemical values, prediction equations can be used.

Supported by NIH DK54918 and an unrestricted grant from Bristol-Myers-Squibb.

Obesity Research, Vol. 7 Suppl. 1, Nov 1999, 27S

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