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The Journal of Nutrition Vol. 128 No. 10 October 1998,
pp. 1811-1814
* Institute of Dentistry, University of Oulu, 90220 Oulu, Finland and Oral and Maxillofacial Department, Oulu University Hospital, 90220 Oulu, Finland
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ABSTRACT |
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Abstract
Introduction
Methods
Discussion
References
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The effects of dietary xylitol (xyl) on bone biomechanical properties in ovariectomized rats (ovx) were studied. Forty-two 3-mo-old female Wistar rats were divided into three groups of 14. Rats in two groups were ovariectomized, while those in the control group underwent a sham operation. All rats received a basal diet, and half of the ovx were given an additional 10 g/100 g dietary xyl supplementation. Three months later the rats were killed and their tibias, femurs and humeri were prepared. The tibias were used for analyses of bone density and trabecular bone volume (BV/TV) and for the three-point bending test. The femurs were used for the torsion test and for the loading test of the femoral neck. The humeri were used for analyses of bone ash weight and bone concentrations of calcium and phosphorus. Dietary xyl gave a significant protection against ovariectomy-caused decline of tibial stress in the three-point bending test, of femoral shear stress in the torsion test, and of stress of the femoral neck, without affecting bone elasticity values. Xyl restricted the ovariectomy-caused reduction in bone density, in bone ash weight and in concentrations of bone calcium and phosphorus. Furthermore, trabecular bone loss in ovx was significantly suppressed by dietary xyl. These results indicate that a 10% dietary supplementation of xyl in ovx has a protective effect against the weakening of bone biomechanical properties. This is related to greater BV/TV and maintained bone mineral content.
KEY WORDS: · bone biomechanics · dietary xylitol · rats · experimental osteoporosis · trabecular bone volume
Xylitol (xyl)4 is a 1,2,3,4,5-pentahydroxypentane, classified as a polyol and as a sugar alcohol. Xyl occurs widely in nature, especially in plants and berries (Mäkinen and Söderling 1980 Supplementing rat diet with xyl increases the calcium and phosphorus levels of bone (Knuuttila et al. 1989 On the basis of the results of our previous studies, it can be hypothesized that dietary xyl also could protect against the weakening of bone biomechanical properties following ovariectomy. The bending and torsional strength of long bones, both of which are decreased after ovariectomy, have been used as indicators of cortical bone strength (Danielsen et al. 1992 Laboratory animals.
Forty-two 3-mo-old female Wistar rats (Laboratory Animal Center, University of Oulu, Oulu, Finland) weighing 200 ± 10 g were divided into three groups of 14. Rats in two groups were bilaterally ovariectomized by a dorsal approach while control rats underwent a sham operation under Hypnorm-Dormicum anesthesia [Hypnorm® (Janssen Pharmaceutica, Beerse, Belgium) and Dormicum® (Roche, Basel, Switzerland), 1:1:2 water 0.2-0.4 mL/100 g ip]. The rats were fed a basal powder diet, RM1 (Special Diet Services, Witham, Essex, UK). Of this diet 1 kg contains 885 g cereal products (wheat, barley and wheatfeed), 60 g vegetable proteins, 25 g animal protein (whey powder), 5 g soybean oil, 7.1 g calcium, 2.9 g phosphorus and 15 µg cholecalciferol. The diet of 14 ovariectomized rats was supplemented with 10 g/100 g xylitol (Xyrofin, Kotka, Finland). The diets were isocaloric. The rats had free access to the food and to tap water.
Biochemical analyses.
Whole bone wet weight (with the marrow) of right tibia was determined, followed by a pycnometric measurement of bone density. The right humeri were used for the other biochemical analyses. After epiphyses and bone marrow of the humeri had been carefully removed and the samples dried at 60°C for 24 h, the bone samples were pulverized with a micromill Mixer Type III 695 (Retsch, Haan, Germany). Bone calcium and phosphorus content were determined spectrophotometrically as presented earlier (Knuuttila et al. 1989
Mechanical testing procedures.
Left tibias and both femurs were stored at Histology.
The proximal part of the right tibia was selected for the measurement of trabecular bone volume (BV/TV). Proximal tibias were cut sagittally in two equal halves with a diamond saw, dehydrated with ethanol (40%) and embedded in methylmethacrylate. Undecalcified sections of 5 µm were cut with a Polycut S heavy-duty microtome (Reichert-Jung, Leica Instruments GmbH, Nussloch, Germany) and stained according to the method of von Kossa (Dickson, 1984 Statistical methods.
Statistically significant differences among the groups were determined by using one-way analysis of variance. When the F-statistic was significant, further comparisons were made using Fishers Protected Least Significant Difference. The statistical computer program used was StatView II for Macintosh (Abacus Concepts, Berkeley, CA). Values in the text are means ± SD.
The weight gain of ovx, 80 ± 21 g, significantly exceeded that of sham-operated controls (sham), 36 ± 13 g, during the 3-mo experimental period (P < 0.01). However, the weight gain of ovx fed a diet supplemented with 10% xyl (ovx + xyl), 61 ± 19 g, was 24% less than that of ovx without dietary xyl supplementation (P < 0.05). The mean food intake of ovx rats, 18.0 ± 3.3 g/d exceeded the food intake of both sham rats, 16.5 ± 3.5 g/d, and ovx ± xyl rats, 16.7 ± 3.4 g/d (P < 0.05).
Signs of estrogen deficiency-caused osteopenia, including a decline in bone biomechanical properties and a decrease in bone mineral content, were clearly detected in this study. The results concerning bone strength properties were in accordance with previous studies performed either with the same testing machine (Lepola et al. 1993
INTRODUCTION
Abstract
Introduction
Methods
Discussion
References
). It is also an intermediate of mammalian carbohydrate metabolism (Touster 1974). Xyl is used as a source of energy in intravenous nutrition as well as a caries-preventive sugar substitute (Mäkinen 1978).
), and promotes the restoration of bone calcium content during rehabilitation following dietary calcium deficiency (Svanberg and Knuuttila 1993). Furthermore, dietary xyl retards bone resorption (Mattila et al. 1995, Svanberg and Knuuttila 1994b) and preserves bone mineral concentration after ovariectomy in rats (Svanberg and Knuuttila 1994a).
, Paavolainen 1978, Strömberg and Dalen 1976). Loading of the femoral neck gives an indication of trabecular bone strength (Hou et al. 1991, Peng et al. 1994). As is known, an increased bone mass does not always lead to better mechanical properties of bones, e.g., following fluoride therapy (Jiang et al. 1996). Consequently, the objective of the present study was to investigate whether a 10% xyl supplementation in the diet of ovariectomized rats (ovx) could preserve bone biomechanical properties as measured by three-point bending test of the tibia, by torsion test of the femur and by loading test of the femoral neck.
SUBJECTS AND METHODS
Abstract
Introduction
Methods
Discussion
References
). Bone ash weight was determined by ashing 25 mg of the pulverized bone at 900°C for 24 h.
View this table:
Table 1.
Biomechanical parameters of three-point bending test of tibia, of torsion test of femur and of loading test of femoral neck in ovariectomized rats (ovx) fed diet supplemented with 10% xylitol (ovx + yxl) and in sham-operated rats1,2
20°C until used. Before testing, the bones were thawed at room temperature and kept moist until the test was completed. Length of the bones was measured with calipers. To avoid confusion with earlier loadings, all mechanical tests were performed with different bones. The left femur was used for the torsion test, the right femur for the loading test of the femoral neck, and the left tibia for the three-point bending test.
. The point of fracture in the three-point bending test was standardized by always placing the bone similarly in the testing machine. A supporter with two loading points, 13 mm apart from each other, was used on the stage of the testing machine. The lateral surface of the tibia at the tibiofibular junction was placed upon the first point and proximal tibia upon the other. A press head compressed the middle of the tibial shaft until fracture occurred. The press head was rounded to avoid cutting into the bone when loaded. Stress, strain, and Young's modulus were derived from load-deformation curves obtained by using equations described by Turner and Burr (1993). The roundness of tibia cross-sections was within 0.92-0.94, and the cross-sectional moment of inertia was determined by the equation of round specimens. In the loading test of femoral neck, the head of the femur was loaded with a force parallel to the shaft of the femur until failure. The cross-section was measured using a standardized position of the opposite bone. This was done by cutting the opposite femoral neck perpendicularly at its narrowest point. The area of the femoral neck was measured and the ratio load/area was calculated.
with a machine constructed on the basis of those introduced by Strömberg and Dalen (1976) and Paavolainen (1978). Each end of the femur was placed concentrically in a cavity of a nut (M8, Bulten Kanthal, Hallstahammar, Sweden) by a standardized method, and fixed with dental stone (Fujirock, G-C Dental Industrial, Tokyo, Japan). The femur, complete with the nuts, was inserted into two head sleeves of the torsion machine, and torsion was performed by twisting the bone inward. The cross-section was measured using a standardized position of the opposite bone. This was done by cutting the opposite femur perpendicularly at its narrowest point. Shear stress and shear modulus of elasticity were derived from load-deformation curves obtained by using equations described by Turner and Burr (1993).
). Sections were taken near the sagittal midline of the tibia at five levels, 50 µm apart, and one microscopic field per section was evaluated. BV/TV was measured in an area of 5 mm2 at 4× objective magnification using a computer image analyzer (MCID, Model M1, Imaging Research, St. Catharines, Canada). The area situated within 1 mm from the upper surface of growth plates, as well as all trabeculae in contact with cortices, was excluded from the measurements.
RESULTS
View this table:
Table 2.
Weight, density, biochemical and biometrical data of tibia and humerus in ovariectomized rats (ovx) fed a diet supplemented with 10% xylitol (ovx + xyl), and in sham-operated rats (sham)1
DISCUSSION
Abstract
Introduction
Methods
Discussion
References
, Peng et al. 1994) or similar equipment (Paavolainen 1978, Strömberg and Dalen 1976).
). As a result, the favorable effects of a 10% dietary supplementation of xyl on bone could not fully protect against the ovariectomy-induced changes in trabecular bone.
), are in accordance with the improved biomechanical properties of the cortical bone.
). Additionally, our recent study with ovx showed no relative changes in cross-linking of bone collagen (Svanberg et al. 1997). These findings indicate that alterations in bone during osteoporosis are more prominent in the mineral than in the collagen fraction. On the other hand, these data negate the possibility that dietary xyl decreases bone elasticity.
) and leading to an increased bone calcium content (Knuuttila et al. 1989) is probably involved. Furthermore, the first steps in the metabolism of xyl produce a reduced redox state (increased NADPH/NADP and NADH/NAD ratios) (Froesch and Jakob 1974). An increased NADH concentration promotes calcium transport across the cell surface membrane (Lehninger et al. 1978), and the reduced redox state is further associated with active calcification (Shapiro et al. 1982), increased collagen synthesis and decreased collagenase activity (Hernández-Munoz et al. 1994).
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FOOTNOTES |
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Manuscript received 30 December 1997. Initial reviews completed 20 February 1998. Revision accepted 11 June 98.
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ACKNOWLEDGMENTS |
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We thank Kalervo Väänänen for valuable comments during the preparation of this manuscript. We also thank Timo Jämsä for his help concerning the biomechanical testing of the bones and Tarja Lassila for her technical assistance.
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LITERATURE CITED |
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Abstract
Introduction
Methods
Discussion
References
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