Effects of vigorous exercise training and beta-agonist administration on bone response to hindlimb suspension.

The effectiveness of dobutamine (Dob) in preventing bone loss during 14 days of hindlimb suspension (Sus) was tested in exercise-trained (Ex; n = 25) and sedentary (Sed; n = 22) rats (age 155 days). One-half of each group was given Dob (2 mg . kg-1 . day-1) or saline (Sal). Histomorphometric measurements at midfemur revealed a 17% smaller cortical bone area (CBA) and a 32% lower periosteal mineral apposition rate (MAR) in suspended vs. nonsuspended Sed/Sal rats. Dob abolished this decline in CBA in Sed/Sus rats, probably via an attenuation of the decrease in periosteal MAR; similar but nonsignificant effects on cross-sectional moment of inertia were observed. Nonsuspended Ex rats had no change in bone CBA when CBA is indexed to body weight. Sus appeared to uncouple the relationship between soleus weight and CBA. Dob attenuated the 43% decline in soleus weight after Sus in Ex but not in Sed rats. In summary, vigorous Ex before Sus does not affect loss of bone mass due to unloading; Dob effectively maintains CBA in Sed rats subjected to suspension.

Preventing annoyance from odors in spaceflight: a method for evaluating the sensory impact of rodent housing.

For the scientific community, the ability to fly mice under weightless conditions in space offers several advantages over the use of rats. These advantages include the option of testing a range of transgenic animals, the ability to increase the number of animals that can be flown, and reduced demands on shuttle resources (food, water, animal mass) and crew time (for water refill). Mice have been flown in animal enclosure module (AEM) hardware only once [Space Shuttle Transport System (STS)-90] and were dissected early in the mission, whereas rats have been flown in the AEM on >20 missions. This has been due, in part, to concerns that strong and annoying odors from mouse urine (vs. rat urine) will interfere with crew performance in the shuttle middeck. To screen and approve mice for flight, a method was developed to evaluate the odor containment performance of AEMs housing female C57BL/6J mice compared with AEMs housing Sprague-Dawley rats across a 21-day test period. Based on the results of this test, consensus was reached that mice could fly in the AEM hardware for up to 17 days (including prelaunch and contingency) and that the AEM hardware would likely contain odors beyond this duration. Human sensory and electronic nose analysis of the AEMs postflight demonstrated their success in containing odors from mice for the mission duration of STS-108 (13 days). Although this paper focuses specifically on odor evaluations for the space shuttle, the concern is applicable to any confined, closed-system environment for human habitation.

Skeletal muscle adaptations to microgravity exposure in the mouse.

To investigate the effects of microgravity on murine skeletal muscle fiber size, muscle contractile protein, and enzymatic activity, female C57BL/6J mice, aged 64 days, were divided into animal enclosure module (AEM) ground control and spaceflight (SF) treatment groups. SF animals were flown on the space shuttle Endeavour (STS-108/UF-1) and subjected to approximately 11 days and 19 h of microgravity. Immunohistochemical analysis of muscle fiber cross-sectional area revealed that, in each of the muscles analyzed, mean muscle fiber cross-sectional area was significantly reduced (P < 0.0001) for all fiber types for SF vs. AEM control. In the soleus, immunohistochemical analysis of myosin heavy chain (MHC) isoform expression revealed a significant increase in the percentage of muscle fibers expressing MHC IIx and MHC IIb (P < 0.05). For the gastrocnemius and plantaris, no significant changes in MHC isoform expression were observed. For the muscles analyzed, no alterations in MHC I or MHC IIa protein expression were observed. Enzymatic analysis of the gastrocnemius revealed a significant decrease in citrate synthase activity in SF vs. AEM control.

The respiratory effects of the cytokine regulating agent HP 228 alone and in combination with morphine in human volunteers.

HP 228 is a synthetic heptapeptide analog of alpha-MSH that attenuates the production and release of inflammatory cytokines. The purpose of this study was to define HP 228's effects, alone and in combination with morphine, on resting ventilation and the ventilatory response to hypoxia and hypercarbia. Six healthy nonsmoking young adult males completed the four-session experiment. Subjects first underwent an initial training session. During subsequent sessions, each subject was tested for the respiratory effects of intravenous HP 228 (30 microg/kg), morphine (0.15 mg/kg), or HP 228 (30 microg/kg) plus morphine (0.15 mg/kg) in a double-blind placebo-controlled randomized balanced within-subjects experimental design. Sessions began with baseline measurement of resting ventilation, oxygen consumption, the isocapnic hypoxic ventilatory response (HVR), and normoxic hypercapnic ventilatory response (HCVR). A second set of respiratory measurements were obtained 10 min after completion of HP 228 or placebo infusion. Morphine or placebo was then administered and ventilatory responses were determined 15 and 40 min postinfusion. HP 228 produced cutaneous flushing, but had no significant effect on respiration or hemodynamics. Morphine significantly decreased metabolism, resting ventilation, and hypoxic and hypercarbic ventilatory responsiveness, independent of prior HP 228 administration. A seventh subject experienced a significant cardiac arrhythmia upon exposure to hypoxia after receiving both HP 228 and morphine and was withdrawn from further study. In conclusion, in this early Phase I clinical trial, HP 228 was found to neither depress ventilation nor augment morphine-induced respiratory depression in healthy young males.

Life sciences flight hardware development for the International Space Station.

During the construction phase of the International Space Station (ISS), early flight opportunities have been identified (including designated Utilization Flights, UF) on which early science experiments may be performed. The focus of NASA's and other agencies' biological studies on the early flight opportunities is cell and molecular biology; with UF-1 scheduled to fly in fall 2001, followed by flights 8A and UF-3. Specific hardware is being developed to verify design concepts, e.g., the Avian Development Facility for incubation of small eggs and the Biomass Production System for plant cultivation. Other hardware concepts will utilize those early research opportunities onboard the ISS, e.g., an Incubator for sample cultivation, the European Modular Cultivation System for research with small plant systems, an Insect Habitat for support of insect species. Following the first Utilization Flights, additional equipment will be transported to the ISS to expand research opportunities and capabilities, e.g., a Cell Culture Unit, the Advanced Animal Habitat for rodents, an Aquatic Facility to support small fish and aquatic specimens, a Plant Research Unit for plant cultivation, and a specialized Egg Incubator for developmental biology studies. Host systems (Figure 1A, B: see text), e.g., a 2.5 m Centrifuge Rotor (g-levels from 0.01-g to 2-g) for direct comparisons between g and selectable g levels, the Life Sciences Glovebox for contained manipulations, and Habitat Holding Racks (Figure 1B: see text) will provide electrical power, communication links, and cooling to the habitats. Habitats will provide food, water, light, air and waste management as well as humidity and temperature control for a variety of research organisms. Operators on Earth and the crew on the ISS will be able to send commands to the laboratory equipment to monitor and control the environmental and experimental parameters inside specific habitats. Common laboratory equipment such as microscopes, cryo freezers, radiation dosimeters, and mass measurement devices are also currently in design stages by NASA and the ISS international partners.

Lack of effect of gallium nitrate on bone density in a rat model of simulated microgravity.

The effect of gallium (Ga) nitrate upon bone density was studied in 24 male Sprague-Dawley rats (275-335 g) in an established model of simulated microgravity. Rats, tail-suspended for 14 d from a system of double pulleys, were allowed free mobility with their hind limbs unloaded. Animals were randomized into four groups of six: 1) unsuspended, saline; 2) suspended, saline; 3) unsuspended, Ga; and 4) suspended, Ga. On day 0, 1 d prior to suspension, rats received equal volumes of a single subcutaneous injection of either saline or 30 mg/kg of elemental Ga in the form of Ga nitrate. On day 15, all animals were euthanized under anesthesia, and their hind limbs analyzed in vitro using single-photon absorptiometry. Although we previously demonstrated that Ga inhibits bone resorption in patients with Paget's disease of bone, and although Ga exhibits many apparent similarities in efficacy to diphosphonates, which prevent bone loss in ovariectomized, paralyzed, and single-limb immobilized rats, Ga failed to prevent bone loss in the tail-suspended rats.

Effect of spaceflight on oxidative and antioxidant enzyme activity in rat diaphragm and intercostal muscles.

There are limited data regarding changes in oxidative and antioxidant enzymes induced by simulated or actual weightlessness, and any additional information would provide insight into potential mechanisms involving other changes observed in muscles from animals previously flown in space. Thus, the NASA Biospecimen Sharing Program was an opportunity to collect valuable information. Oxidative and antioxidant enzyme levels, as well as lipid perioxidation, were measured in respiratory muscles from rats flown on board Space Shuttle mission STS-54. The results indicated that there was an increasing trend in citrate synthase activity in the flight diaphragm when compared to ground based controls, and there were no significant changes observed in the intercostal muscles for any of the parameters. However, lipid peroxidation was significantly (p<0.05) decreased in the flight diaphragm. These results indicate that 6 day exposure to microgravity may have a different effect on oxidative and antioxidant activity in rat respiratory muscles when compared to data from previous 14 day hindlimb suspension studies.

Effects of aminohydroxybutane bisphosphonate on bone growth when administered after hind-limb bone loss in tail-suspended rats.

The effects of aminohydroxybutane bisphosphonate (AHBuBP) on bone after disuse osteopenia were studied in tail-suspended rats. Male Sprague-Dawley rats (weight range, 313-352 g) randomized into four groups of eight animals received 2 ml kg-1 day-1 of either AHBuBP (0.3 mg kg-1 day-1) or normal saline (vehicle) subcutaneously on days 14 and 15 of a 28-day experiment. The groups were 1) nonsuspended, saline; 2) suspended on days 14 to 28, saline; 3) suspended on days 0 to 28, AHBuBP; and 4) suspended on days 0 to 28, saline. On days 19 and 26, all rats received 15 mg/kg (1 ml/kg) of calcein. On day 28, they were sacrificed and their tibias and femurs were analyzed in vitro for bone density, strength and stiffness. The tibias were also analyzed histomorphometrically. The tibias and femurs from AHBuBP-treated rats were as dense as those in the nonsuspended group, whereas tail suspension in the untreated rats for 14 and 28 days caused a significant decrease in bone density. However, in measurements of bone strength and stiffness, the samples from the rats that received AHBuBP were similar to those of untreated rats suspended for 14 days, suggesting the newly formed bone was weaker. In the AHBuBP group, compared with all others, static histologic measurements of the proximal tibial metaphyses showed an increased bone area and perimeter and a decreased percentage of osteoid perimeter without a difference in the percentage of eroded perimeter. Dynamic histologic studies showed a decreased bone formation rate and decreased longitudinal growth rate.(ABSTRACT TRUNCATED AT 250 WORDS)

Effectiveness of a cytokine restraining agent (CRA (TM)) in attenuating disuse deconditioning induced by hindlimb unloading in rats.

Spaceflight alters many immune responses and among the regulatory components of an organisms response system that have been to be affected by spaceflight is the cytokine network. Spaceflight, as well as ground-based model systems of spaceflight, have been shown to affect the production and activation of various cytokines including interleukins (IL) and tumor necrosis factor (TNF). Levels of urinary IL-2 are elevated on the first day of spaceflight and again after returning from space. Most results from ground-based studies in rodents indicate either no alterations in cytokines or decreased levels. Results from this experiment indicate that HP 228, a potent cytokine restraining agent (CRA (TM)) was effective in attentuating many of the disuse deconditioning changes induced by the ground-based hindlimb suspension model that simulates weightlessness in rats. HP 228 is a novel heptapeptide with unnatural amino acids and can effectively restrain lipopolysaccharide (LPS)-induced increased levels of several key cytokines, including plasma TNF alpha, IL-1 beta and IL-6. HP 228 has also been shown to be effective in several rodent models of pain, inflammation and LPS-induced lethality, as well as in reducing inducible nitric oxide synthase.

Synthetic hexa- and heptapeptides that inhibit IL-8 from binding to and activating human blood neutrophils.

IL-8 is a member of the chemokine alpha subfamily that activates and is chemotactic for neutrophils. In these studies, we have synthesized and characterized a hexapeptide inhibitor of IL-8. This peptide, with an acetylated amino terminus and an amidated carboxyl terminus (Ac-RRWWCR-NH2), inhibited the specific binding of 125I-IL-8 to neutrophils. The inhibition was biphasic and apparent Ki was estimated to be approximately 2.7 microM and 13 microM for two different IL-8 binding sites. The peptide inhibited neutrophil chemotaxis, beta-glucuronidase release from neutrophils, and rabbit skin edema induced by IL-8 with an EC50 of 90 microM, 0.8 microM, respectively. Ac-RRWWCR-NH2 also suppressed the binding of macrophage inflammatory protein (MIP) 2 beta to neutrophils. However, it did not inhibit the binding of MIP-1 alpha, C5a, or leukotriene B4 to neutrophils, chemotaxis induced by FMLP, or beta-glucuronidase release induced by FMLP, C5a, or leukotriene B4. Additional peptides were analyzed to identify a better inhibitor. Inhibition of binding by Ac-rrwwcrc-NH2 synthesized with all D-amino acids was almost four times more potent than Ac-RRWWCR-NH2. Small peptide homologues of the amino-terminal end of IL-8 failed to inhibit IL-8 binding to neutrophils. These studies have identified several peptides that significantly inhibit IL-8 function. Because IL-8 seems to be an important inflammatory mediator of several human illnesses, these peptides may have pharmacologic potential.

Aminohydroxybutane bisphosphonate and clenbuterol prevent bone changes and retard muscle atrophy respectively in tail-suspended rats.

Hind-limb unloading by tail suspension of rats, an established model of simulated microgravity, was used to examine the efficacy of aminohydroxybutane bisphosphonate (AHBuBP) and clenbuterol in preventing bone loss and muscle atrophy, respectively. Male Sprague-Dawley rats (299-372 g) were randomized into six groups of six: 1) unsuspended, saline, 2) unsuspended, saline, pair fed with group 3, 3) suspended, saline, 4) suspended, 0.03 mg/kg/day x 2 of AHBuBP, 5) suspended, 0.3 mg/kg/day x 2 of AHBuBP and 6) suspended, 0.3 mg/kg/day x 2 of AHBuBP + clenbuterol (0.5 mg/kg/day i.p. x 6, then 1 mg/kg/day i.p. x 6). Animals in groups 3 to 6 were tail suspended for 14 days from a system of double pulleys and allowed free mobility with their hind limbs unloaded. On days -2 and -1, before suspension on day 0, all rats received a single s.c. injection of either 2 ml/kg of normal saline (vehicle) or AHBuBP. All rats were tested for exercise tolerance before day -2 and on day 10, and grip strength before day -2 and on day 13. On day 14, the rats were euthanized and their humeri, tibias and femurs analyzed in vitro for bone density (by single-photon absorptiometry), strength and stiffness (by 3-point bending). Muscles were analyzed for weight, protein concentration and enzyme activity. Pair feeding had no effect other than on food consumption and body weight. AHBuBP caused a dose-dependent increase in bone density in humeri, tibias and femurs, even in tail-suspended rats, relative to control unsuspended animals, with no significant difference in bone strength or stiffness between AHBuBP groups and unsuspended animals.(ABSTRACT TRUNCATED AT 250 WORDS)