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At least 109 records · Page 6

Effects of Growth Hormone/IGF-I and Exercise on Unloaded Bones

Growth hormone (GH) and insulin-like growth factor-I (IGF-I) in combination with exercise prevent muscle atrophy induced by unloading in the tail-suspension rat model for space flight (Gosselink et al, FASEB J 1994). This study evaluated the effects of these treatments on bone. Hypophysectomized rats were suspended (S) and treated with 1mg/kg/day CH plus IGF-I (H) or vehicle (Sal) daily by injection and exercised (Ex) by 3 climbs up a 1m ladder carrying a load equal to 30% the initial body weight (BW) 3x/day for 10 days. Tibial epiphysis (Epi) widths were measured by micrometry and femoral Bone Mineral Content (fBMC) in excised femurs by DEXA (Lunar DPX-L). Serum calcium (Ca) and phosphorus (Pi) were measured by COBAS Autoanalyzer (Roche Diag.). Ambulatory (Amb)-H treated rats showed growth rates of 6.6+-0.9 g/day, similar to S-H-Ex and higher than S-H (3.210.6, p less than 0.05) and S-Sal (-0.711.0, p less than 0.05). Epi widths were 10% lower in S-Sal, and S-Sal-Ex, and increased 100% in all H groups. fBMC was less in S than Amb, only when all S groups are compared to both Amb groups (p less than 0.03). H treatment increased fBMC (p less than 0.05) but reduced fBMC/100g BW in all H groups (p less than 0.001). The reduced density of H bone cannot be attributed to low circulating Ca. and Pi since they were higher in H than Sal (p less than 0.001). H treatment for 10 days in doses sufficient to support normal growth in BW failed to produce normal Epi widths or fBMC, even when combined with exercise. The suspension effect observed in Epi widths was not corrected by H or Ex alone, but was improved by H plus a This regimen. although effective in preventing muscle atrophy, failed to return bone measures, Epi widths and fBMC, to normal.

Harper, J. S.↗

Skeletal Phenotype of Transgenic Mice Expressing the Beta1 Integrin Cytoplasmic Tail In Osteoblasts

To define the physiologic role of beta1 integrin in bone formation and mechanical loading, transgenic mice were generated by expressing the cytoplasmic tall and transmembrane domain of Beta1 integrin under the control of the osteocalcin promoter. In cultured cells, this truncated fragment of Beta1 can act as a dominant negative. Previously, the matrix of calvariae was shown to be abnormal in transgenic (TG) compared to wildtype (WT) mice. In this study, we analyzed appendicular bone in TG and WT, male and female mice at 14, 35, 63, 90 and 365 days old (n=8-12/gp). To assess beta1 integrin function in mechanical loading, a pilot study using hindlimb unloading by tail suspension was performed. 35d old TG and WT females were hindlimb unloaded for 4 wks (n=3-5). Body mass, bone mineral content, histomorphometric (distal femur) and biomechanical parameters were analyzed. Statistical significance (P less than.05) was defined by ANOVA using the Tukey-Kramer post-hoc test. We confirmed transgene expression by immunoprecipitating then immunoblotting bone lysates using an antibody against the beta1 tail. Body masses of TG mice at 63, 90 and 365d old were greater (16-25%) than WT. Some TG female mice at 365d appeared obese; mean abdominal fat mass was 415% greater in TG than WT mice. Tibiae were longer (5-7%) in TG than WT mice at 63 and 90d. Tibial mineral mass of 35d males was 7% lower in TG than WT mice, but at 63d was 21% higher. The % osteoblast surface in 35d TG mice was 20% higher than WT, and at 63d was 17% lower, while % osteoclast surface did not differ. In 365d mice, cancellous bone volume (125%) and endocortical mineral apposition rate (40%) were greater in TG than WT males but not females. In WT mice, hindlimb unloading caused a reduction in mineral mass of tibiae (-20%) and lumbar vertebrae (-22%) relative to normally loaded controls. Surprisingly, hindlimb unloading also caused a relative reduction (-13%) in humerus mass. The effects of hindlimb unloading on tibia and humerus mass were less obvious in TG than in WT mice. Since hindlimb unloading caused skeletal changes in both loaded and unloaded bones, systemic changes may contribute to bone responses observed using this animal model. In conclusion, transgene expression resulted in marked metabolic changes during growth and in the aged female. Our results demonstrate that expression of the Beta1 integrin cytoplasmic tail in vivo causes gender- and age-specific changes in select morphometric parameters, bone length, and bone mass.

Globus, R. K.↗

Is Animal Age a Factor In the Response of Bone to Spaceflight?

The rodent bone response to spaceflight may be influenced by a multitude of actors including flight duration, strain, and housing. Review of bone formation rates during spaceflight suggests that age may also play a role in the response. Weanling rats show fewer bone changes than older rats. To determine if the long bones of weanling rats were insensitive to weight-bearing, a hindlimb unloading experiment was conducted simultaneously with a 9d shuttle flight in 34d old group-housed male rats. All animals were injected with bone markers 7d and 1d before flight and euthanized at landing, 24hr, and 72hr following recovery. If no differences in body weight, bone length, or bone formation at the tibiofibular junction were noted at the different time points, data were combined for each group. No significant differences in body weight were found at any time period among the groups. The humerus, tibia, and femur elongated significantly during the flight period with no difference in lengths between groups at the end of the flight period. The group-housed flight rats showed no change in cortical bone formation rate compared to preflight values, flight controls, or vivarium controls. However, the hindlimb unloading group showed a significant 30% decrease in bone formation rate compared to all other groups. Individually-housed 38d old animals flown for 14d showed approx. 10% suppression of cortical growth. We speculate that the mechanical threshold required for cross-sectional bone growth is reached in group-house weanling rats during spaceflight, perhaps, through physical interactions, and that the weanling animals are sensitive to loading. However, the threshold is not fully reached in either singly-housed flight or hindlimb unloaded weanling rats. Older singly-housed flight animals appear to show equal or greater bone changes compared to hindlimb unloaded rats. We conclude that age, flight duration, strain, and housing have important roles in rodent skeletal responses to spaceflight.

Morey-Holton, E. R.↗

Hindlimb unloading has a greater effect on cortical compared with cancellous bone in mature female rats

This study was designed to determine the effects of 28 days of hindlimb unloading (HU) on the mature female rat skeleton. In vivo proximal tibia bone mineral density and geometry of HU and cage control (CC) rats were measured with peripheral quantitative computed tomography (pQCT) on days 0 and 28. Postmortem pQCT, histomorphometry, and mechanical testing were performed on tibiae and femora. After 28 days, HU animals had significantly higher daily food consumption (+39%) and lower serum estradiol levels (-49%, P = 0.079) compared with CC. Proximal tibia bone mineral content and cortical bone area significantly declined over 28 days in HU animals (-4.0 and 4.8%, respectively), whereas total and cancellous bone mineral densities were unchanged. HU animals had lower cortical bone formation rates and mineralizing surface at tibial midshaft, whereas differences in similar properties were not detected in cancellous bone of the distal femur. These results suggest that cortical bone, rather than cancellous bone, is more prominently affected by unloading in skeletally mature retired breeder female rats.

NASA Discipline Musculoskeletal↗

Insulin-like growth factor I is required for the anabolic actions of parathyroid hormone on mouse bone

Parathyroid hormone (PTH) is a potent anabolic agent for bone, but the mechanism(s) by which it works remains imperfectly understood. Previous studies have indicated that PTH stimulates insulin-like growth factor (IGF) I production, but it remains uncertain whether IGF-I mediates some or all of the skeletal actions of PTH. To address this question, we examined the skeletal response to PTH in IGF-I-deficient (knockout [k/o]) mice. These mice and their normal littermates (NLMs) were given daily injections of PTH (80 microg/kg) or vehicle for 2 weeks after which their tibias were examined for fat-free weight (FFW), bone mineral content, bone structure, and bone formation rate (BFR), and their femurs were assessed for mRNA levels of osteoblast differentiation markers. In wild-type mice, PTH increased FFW, periosteal BFR, and cortical thickness (C.Th) of the proximal tibia while reducing trabecular bone volume (BV); these responses were not seen in the k/o mice. The k/o mice had normal mRNA levels of the PTH receptor and increased mRNA levels of the IGF-I receptor but markedly reduced basal mRNA levels of the osteoblast markers. Surprisingly, these mRNAs in the k/o bones increased several-fold more in response to PTH than the mRNAs in the bones from their wild-type littermates. These results indicate that IGF-I is required for the anabolic actions of PTH on bone formation, but the defect lies distal to the initial response of the osteoblast to PTH.

Non-NASA Center↗

Calcium metabolism and cardiovascular function after spaceflight

To determine the influence of dietary calcium on spaceflight-induced alterations in calcium metabolism and blood pressure (BP), 9-wk-old spontaneously hypertensive rats, fed either high- (2%) or low-calcium (0.02%) diets, were flown on an 18-day shuttle flight. On landing, flight animals had increased ionized calcium (P < 0.001), elevated parathyroid hormone levels (P < 0.001), reduced calcitonin levels (P < 0.05), unchanged 1,25(OH)(2)D(3) levels, and elevated skull (P < 0.01) and reduced femur bone mineral density. Basal and thrombin-stimulated platelet free calcium (intracellular calcium concentration) were also reduced (P < 0.05). There was a tendency for indirect systolic BP to be reduced in conscious flight animals (P = 0.057). However, mean arterial pressure was elevated (P < 0.001) after anesthesia. Dietary calcium altered all aspects of calcium metabolism (P < 0.001), as well as BP (P < 0.001), but the only interaction with flight was a relatively greater increase in ionized calcium in flight animals fed low- compared with high-calcium diets (P < 0.05). The results indicate that 1) flight-induced disruptions of calcium metabolism are relatively impervious to dietary calcium in the short term, 2) increased ionized calcium did not normalize low-calcium-induced elevations of BP, and 3) parathyroid hormone was paradoxically increased in the high-calcium-fed flight animals after landing.

NASA Discipline Cardiopulmonary↗

Calcium bioavailability of vegetarian diets in rats: potential application in a bioregenerative life-support system

Calcium bioavailability of vegetarian diets containing various proportions of candidate crops for a controlled ecological life-support system (CELSS) was determined by femur 45Ca uptake. Three vegetarian diets and a control diet were labeled extrinsically with 45Ca and fed to 5-wk old male rats. A fifth group of rats fed an unlabeled control diet received an intraperitoneal (IP) injection of 45Ca. There was no significant difference in mean calcium absorption of vegetarian diets (90.80 +/- 5.23%) and control diet (87.85 +/- 5.25%) when calculated as the percent of an IP dose. The amounts of phytate, oxalate, and dietary fiber in the diets did not affect calcium absorption.

NASA Discipline Life Support Systems↗

Effect of oral calcium and calcium + fluoride treatments on mouse bone properties during suspension

The bone effects of oral dosages of calcium chloride with or without supplementary sodium fluoride were assessed in antiorthostatically suspended mice. Two calcium dosages were used to replace half (3.1 mM) or all(6.3 mM) of the dietary calcium lost due to reduced food intake by the suspended mice. Two groups of 6.3 mM CaCl2-treated mice were additionally treated with 0.25 or 2.5 mM NaF. The results indicate that supplementation of the mouse drinking water with calcium salts prevents bone changes induced by short-term suspension, while calcium salts in combination with fluoride are less effective as fluoride dosage increases. However, the calcium supplements change the relationship between the femur mechanical properties and the mineral composition of the bone. Because of this, it appears that oral calcium supplements are effective through a mechanism other than simple dietary supplementation and may indicate a dependence of bone consistency on systemic and local fluid conditions.

Non-NASA Center↗

Fractal dimension analysis of weight-bearing bones of rats during skeletal unloading

Fractal analysis was used to quantify changes in trabecular bone induced through the use of a rat tail-suspension model to simulate microgravity-induced osteopenia. Fractal dimensions were estimated from digitized radiographs obtained from tail-suspended and ambulatory rats. Fifty 4-month-old male Sprague-Dawley rats were divided into groups of 24 ambulatory (control) and 26 suspended (test) animals. Rats of both groups were killed after periods of 1, 4, and 8 weeks. Femurs and tibiae were removed and radiographed with standard intraoral films and digitized using a flatbed scanner. Square regions of interest were cropped at proximal, middle, and distal areas of each bone. Fractal dimensions were estimated from slopes of regression lines fitted to circularly averaged plots of log power vs. log spatial frequency. The results showed that the computed fractal dimensions were significantly greater for images of trabecular bones from tail-suspended groups than for ambulatory groups (p < 0.01) at 1 week. Periods between 1 and 4 weeks likewise yielded significantly different estimates (p < 0.05), consistent with an increase in bone loss. In the tibiae, the proximal regions of the suspended group produced significantly greater fractal dimensions than other regions (p < 0.05), which suggests they were more susceptible to unloading. The data are consistent with other studies demonstrating osteopenia in microgravity environments and the regional response to skeletal unloading. Thus, fractal analysis could be a useful technique to evaluate the structural changes of bone.

Non-NASA Center↗

Skeletal unloading causes resistance of osteoprogenitor cells to parathyroid hormone and to insulin-like growth factor-I

Skeletal unloading decreases bone formation and osteoblast number in vivo and decreases the number and proliferation of bone marrow osteoprogenitor (BMOp) cells in vitro. We tested the ability of parathyroid hormone (PTH) to stimulate BMOp cells in vivo by treating Sprague Dawley rats (n = 32) with intermittent PTH(1-34) (1 h/day at 8 microg/100 g of body weight), or with vehicle via osmotic minipumps during 7 days of normal weight bearing or hind limb unloading. Marrow cells were flushed from the femur and cultured at the same initial density for up to 21 days. PTH treatment of normally loaded rats caused a 2.5-fold increase in the number of BMOp cells, with similar increases in alkaline phosphatase (ALP) activity and mineralization, compared with cultures from vehicle-treated rats. PTH treatment of hind limb unloaded rats failed to stimulate BMOp cell number, ALP activity, or mineralization. Hind limb unloading had no significant effect on PTH receptor mRNA or protein levels in the tibia. Direct in vitro PTH challenge of BMOp cells isolated from normally loaded bone failed to stimulate their proliferation and inhibited their differentiation, suggesting that the in vivo anabolic effect of intermittent PTH on BMOp cells was mediated indirectly by a PTH-induced factor. We hypothesize that this factor is insulin-like growth factor-I (IGF-I), which stimulated the in vitro proliferation and differentiation of BMOp cells isolated from normally loaded bone, but not from unloaded bone. These results suggest that IGF-I mediates the ability of PTH to stimulate BMOp cell proliferation in normally loaded bone, and that BMOp cells in unloaded bone are resistant to the anabolic effect of intermittent PTH therapy due to their resistance to IGF-I.

NASA Discipline Musculoskeletal↗

[Clinical and physiological evaluation of bone changes among astronauts after long-term space flights]

Results of the joint Russian/US studies of the effect of microgravity on bone tissues in 18 cosmonauts on return from 4.5- to 14.5-month long missions are presented. Dual-energy x-ray gamma-absorbtiometry (QDR-1000 W, Hologic, USA) was used to measure bone mineral density (BMD, g/cm2) and mineral content (BMC, g) in the whole body, the scalp including cervical vertebra, arms, ribs, sternal and lumbar regions of the spinal column, pelvis and legs. A clearly defined dependence of topography of changes upon the position of a skeletal segment in the gravity vector was established. The greatest BMD losses have been observed in the skeleton of the lower body, i.e. in pelvic bones (-11.99 +/- 1.22%), lumbar vertebra (-5.63 +/- 0.817%), and in proximal femur, particularly in the femoral neck (-8.17 +/- 1.24%). Bones of the upper skeleton were either unchanged (insignificant) or showed a positive trend. Overall changes in bone mass of the whole skeleton of male cosmonauts during the period of about 6 months on mission made up -1.41 +/- 0.406% and suggest the mean balance of calcium over flight equal to -227 +/- 62.8 mg/day. Reasoning is given to qualify these states of cosmonauts' bone tissues as local osteopenia. On the literature and results of authors' clinical evidence, discussed is availability of the densitometric data for predicting risk of trauma. A biological nature of the changes under observation is hypothesized.

NASA Discipline Musculoskeletal↗

Future human bone research in space

Skylab crewmembers demonstrated negative calcium (Ca) balance reaching about -300 mg/day by flight day 84. Limited bone density (BMD) measurements documented that bone was not lost equally from all parts of the skeleton. Subsequent BMD studies during long duration Russian flights documented the regional extent of bone loss. These studies demonstrated mean losses in the spine, femur neck, trochanter, and pelvis of about 1%-1.6% with large differences between individuals as well as between bone sites in a given individual. Limited available data indicate postflight bone recovery occurred in some individuals, but may require several years for complete restoration. Long duration bedrest studies showed a similar pattern of bone loss and calcium balance (-180 mg/day) as spaceflight. During long duration bedrest, resorption markers were elevated, formation markers were unchanged, 1,25 vitamin D (VitD) and calcium absorption were decreased, and serum ionized Ca was increased. Although this information is a good beginning, additional spaceflight research is needed to assess architectural and subregional bone changes, elucidate mechanisms, and develop efficient as well as effective countermeasures. Space research poses a number of unique problems not encountered in ground-based laboratory research. Therefore, researchers contemplating human spaceflight research need to consider a number of unique problems related to spaceflight in their experimental design.

NASA Center JSC↗

Ultrasonic wave velocity measurement in small polymeric and cortical bone specimens

A system was refined for the determination of the bulk ultrasonic wave propagation velocity in small cortical bone specimens. Longitudinal and shear wave propagations were measured using ceramic, piezoelectric 20 and 5 MHz transducers, respectively. Results of the pulse transmission technique were refined via the measurement of the system delay time. The precision and accuracy of the system were quantified using small specimens of polyoxymethylene, polystyrene-butadiene, and high-density polyethylene. These polymeric materials had known acoustic properties, similarity of propagation velocities to cortical bone, and minimal sample inhomogeneity. Dependence of longitudinal and transverse specimen dimensions upon propagation times was quantified. To confirm the consistency of longitudinal wave propagation in small cortical bone specimens (< 1.0 mm), cut-down specimens were prepared from a normal rat femur. Finally, cortical samples were prepared from each of ten normal rat femora, and Young's moduli (Eii), shear moduli (Gij), and Poisson ratios (Vij) were measured. For all specimens (bone, polyoxymethylene, polystyrene-butadiene, and high-density polyethylene), strong linear correlations (R2 > 0.997) were maintained between propagation time and distance throughout the size ranges down to less than 0.4 mm. Results for polyoxymethylene, polystyrene-butadiene, and high-density polyethylene were accurate to within 5 percent of reported literature values. Measurement repeatability (precision) improved with an increase in the wave transmission distance (propagating dimension). No statistically significant effect due to the transverse dimension was detected.

Non-NASA Center↗

Correspondence between theoretical models and dual energy x-ray absorptiometry measurements of femoral cross-sectional growth during adolescence

We have developed an analytical model of long bone cross-sectional ontogeny in which appositional growth of the diaphysis is primarily driven by mechanical stimuli associated with increasing body mass during growth and development. In this study, our goal was to compare theoretical predictions of femoral diaphyseal structure from this model with measurements of femoral bone mineral and geometry by dual energy x-ray absorptiometry. Measurements of mid-diaphyseal femoral geometry and structure were made previously in 101 Caucasian adolescents and young adults 9-26 years of age. The data on measured bone mineral content and calculated section modulus were compared with the results of our analytical model of cross-sectional development of the human femur over the same age range. Both bone mineral content and section modulus showed good correspondence with experimental measurements when the relationships with age and body mass were examined. Strong linear relationships were evident for both parameters when examined as a function of body mass.

NASA Discipline Musculoskeletal↗

Estrogen regulates the rate of bone turnover but bone balance in ovariectomized rats is modulated by prevailing mechanical strain

Estrogen deficiency induced bone loss is associated with increased bone turnover in rats and humans. The respective roles of increased bone turnover and altered balance between bone formation and bone resorption in mediating estrogen deficiency-induced cancellous bone loss was investigated in ovariectomized rats. Ovariectomy resulted in increased bone turnover in the distal femur. However, cancellous bone was preferentially lost in the metaphysis, a site that normally experiences low strain energy. No bone loss was observed in the epiphysis, a site experiencing higher strain energy. The role of mechanical strain in maintaining bone balance was investigated by altering the strain history. Mechanical strain was increased and decreased in long bones of ovariectomized rats by treadmill exercise and functional unloading, respectively. Functional unloading was achieved during orbital spaceflight and following unilateral sciatic neurotomy. Increasing mechanical loading reduced bone loss in the metaphysis. In contrast, decreasing loading accentuated bone loss in the metaphysis and resulted in bone loss in the epiphysis. Finally, administration of estrogen to ovariectomized rats reduced bone loss in the unloaded and prevented loss in the loaded limb following unilateral sciatic neurotomy in part by reducing indices of bone turnover. These results suggest that estrogen regulates the rate of bone turnover, but the overall balance between bone formation and bone resorption is influenced by prevailing levels of mechanical strain.

STS-62 Shuttle Project↗

Cortical bone growth and maturational changes in dwarf rats induced by recombinant human growth hormone

The growth hormone (GH)-deficient dwarf rat was used to investigate recombinant human (rh) GH-induced bone formation and to determine whether rhGH facilitates simultaneous increases in bone formation and bone maturation during rapid growth. Twenty dwarf rats, 37 days of age, were randomly assigned to dwarf plus rhGH (GH; n = 10) and dwarf plus vehicle (n = 10) groups. The GH group received 1.25 mg rhGH/kg body wt two times daily for 14 days. Biochemical, morphological, and X-ray diffraction measurements were performed on the femur middiaphysis. rhGH stimulated new bone growth in the GH group, as demonstrated by significant increases (P < 0.05) in longitudinal bone length (6%), middiaphyseal cross-sectional area (20%), and the amount of newly accreted bone collagen (28%) in the total pool of middiaphyseal bone collagen. Cortical bone density, mean hydroxyapatite crystal size, and the calcium and collagen contents (microgram/mm3) were significantly smaller in the GH group (P < 0.05). Our findings suggest that the processes regulating new collagen accretion, bone collagen maturation, and mean hydroxyapatite crystal size may be independently regulated during rapid growth.

NASA Discipline Musculoskeletal↗

Hindlimb suspension diminishes femoral cross-sectional growth in the rat

Growth, functional adaptation, and torsional strength were examined in the femora of 39-day-old male Sprague-Dawley rats subjected to hindlimb suspension for 0, 1, 2, 3, or 4 weeks and were compared with measurements for age-matched control animals. Our goal was to understand the effect of reduced loading on the normal age-related changes in femoral properties during growth. The control animals exhibited growth-related increases in all geometric and torsional properties of the femur. The mean body mass and femoral length of the hindlimb-suspended rats were similar to those of the controls throughout the experiment. Over 4 weeks, the femoral cross-sectional and torsional measurements from the hindlimb-suspended rats demonstrated increases in comparison with the basal values (+33% cross-sectional area, +64% polar moment of inertia, +67% ultimate torque, and +181% torsional rigidity), but the age-matched controls showed significantly greater growth-related increases (+71% cross-sectional area, +136% polar moment of inertia, +127% ultimate torque, and +367% torsional rigidity). The differences in femoral structural strength between the hindlimb-suspended animals and the age-matched controls were attributable to differences in altered cross-sectional geometry.

NASA Discipline Musculoskeletal↗

Skeletal unloading induces resistance to insulin-like growth factor I

In previous studies with a hindlimb elevation model, we demonstrated that skeletal unloading transiently inhibits bone formation. This effect is limited to the unloaded bones (the normally loaded humerus does not cease growing), suggesting that local factors are of prime importance. IGF-I is one such factor; it is produced in bone and stimulates bone formation. To determine the impact of skeletal unloading on IGF-I production and function, we assessed the mRNA levels of IGF-I and its receptor (IGF-IR) in the proximal tibia and distal femur of growing rats during 2 weeks of hindlimb elevation. The mRNA levels for IGF-I and IGF-IR rose during hindlimb elevation, returning toward control values during recovery. This was accompanied by a 77% increase in IGF-I levels in the bone, peaking at day 10 of unloading. Changes in IGF binding protein levels were not observed. Infusion of IGF-I (200 micrograms/day) during 1 week of hindlimb elevation doubled the increase in bone mass of the control animals but failed to reverse the cessation of bone growth in the hindlimb-elevated animals. We conclude that skeletal unloading induces resistance to IGF-I, which may result secondarily in increased local production of IGF-I.

NASA Center ARC↗