Ionizing radiation can cause substantial tissue degeneration, which may threaten the long-term health of astronauts and radiotherapy patients. To determine whether a single dose of radiation acutely compromises structural integrity in the postpubertal skeleton, 18-week-old male mice were exposed to (137)Cs gamma radiation (1 or 2 Gy). The structure of high-turnover, cancellous bone was analyzed by microcomputed tomography (microCT) 3 or 10 days after irradiation and in basal controls (tissues harvested at the time of irradiation) and age-matched controls. Irradiation (2 Gy) caused a 20% decline in tibial cancellous bone volume fraction (BV/TV) within 3 days and a 43% decline within 10 days, while 1 Gy caused a 28% reduction 10 days later. The BV/TV decrement was due to increased spacing and decreased thickness of trabeculae. Radiation also increased ( approximately 150%) cancellous surfaces lined with tartrate-resistant, acid phosphatase-positive osteoclasts, an index of increased bone resorption. Radiation decreased lumbar vertebral BV/TV 1 month after irradiation, showing the persistence of cancellous bone loss, although mechanical properties in compression were unaffected. In sum, a single dose of gamma radiation rapidly increased osteoclast surface in cancellous tissue and compromised cancellous microarchitecture in the remodeling appendicular and axial skeleton of postpubertal mice.

Exposure of astronauts in space to radiation during weightlessness may contribute to subsequent bone loss. Gamma irradiation of postpubertal mice rapidly increases the number of bone-resorbing osteoclasts and causes bone loss in cancellous tissue; similar changes occur in skeletal diseases associated with oxidative stress. Therefore, we hypothesized that increased oxidative stress mediates radiation-induced bone loss and that musculoskeletal disuse changes the sensitivity of cancellous tissue to radiation exposure. Musculoskeletal disuse by hindlimb unloading (1 or 2 wk) or total body gamma irradiation (1 or 2 Gy of (137)Cs) of 4-mo-old, male C57BL/6 mice each decreased cancellous bone volume fraction in the proximal tibiae and lumbar vertebrae. The extent of radiation-induced acute cancellous bone loss in tibiae and lumbar vertebrae was similar in normally loaded and hindlimb-unloaded mice. Similarly, osteoclast surface in the tibiae increased 46% as a result of irradiation, 47% as a result of hindlimb unloading, and 64% as a result of irradiation + hindlimb unloading compared with normally loaded mice. Irradiation, but not hindlimb unloading, reduced viability and increased apoptosis of marrow cells and caused oxidative damage to lipids within mineralized tissue. Irradiation also stimulated generation of reactive oxygen species in marrow cells. Furthermore, injection of alpha-lipoic acid, an antioxidant, mitigated the acute bone loss caused by irradiation. Together, these results showed that disuse and gamma irradiation, alone or in combination, caused a similar degree of acute cancellous bone loss and shared a common cellular mechanism of increased bone resorption. Furthermore, irradiation, but not disuse, may increase the number of osteoclasts and the extent of acute bone loss via increased reactive oxygen species production and ensuing oxidative damage, implying different molecular mechanisms. The finding that alpha-lipoic acid protected cancellous tissue from the detrimental effects of irradiation has potential relevance to astronauts and radiotherapy patients.

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Space travel and prolonged bed rest cause bone loss due to musculoskeletal disuse. In space, radiation fields may also have detrimental consequences because charged particles traversing the tissues of the body can elicit a wide range of cytotoxic and genotoxic lesions. The effects of heavy-ion radiation exposure in combination with musculoskeletal disuse on bone cells and tissue are not known. To explore this, normally loaded 16-week-old male C57BL/6 mice were exposed to (56)Fe ions (1 GeV/nucleon) at doses of 0 cGy (sham), 10 cGy, 50 cGy or 2 Gy 3 days before tissue harvest. Additional mice were hindlimb unloaded by tail traction continuously for 1 week to simulate weightlessness and exposed to (56)Fe-ion radiation (0 cGy, 50 cGy, 2 Gy) 3 days before tissue harvest. Despite the short duration of this study, low-dose (10, 50 cGy) irradiation of normally loaded mice reduced trabecular volume fraction (BV/TV) in the proximal tibiae by 18% relative to sham-irradiated controls. Hindlimb unloading together with 50 cGy radiation caused a 126% increase in the number of TRAP(+) osteoclasts on cancellous bone surfaces relative to normally loaded, sham-irradiated controls. Together, radiation and hindlimb unloading had a greater effect on suppressing osteoblastogenesis ex vivo than either treatment alone. In sum, low-dose exposure to heavy ions (50 cGy) caused rapid cancellous bone loss in normally loaded mice and increased osteoclast numbers in hindlimb unloaded mice. In vitro irradiation also was more detrimental to osteoblastogenesis in bone marrow cells that were recovered from hindlimb unloaded mice compared to cells from normally loaded mice. Furthermore, irradiation in vitro stimulated osteoclast formation in a macrophage cell line (RAW264.7) in the presence of RANKL (25 ng/ml), showing that heavy-ion radiation can stimulate osteoclast differentiation even in the absence of osteoblasts. Thus heavy-ion radiation can acutely increase osteoclast numbers in cancellous tissue and, under conditions of musculoskeletal disuse, can enhance the sensitivity of bone cells, in particular osteoprogenitors, to the effects of radiation.

Influence of increased mechanical loading by hypergravity on the microtubule cytoskeleton and prostaglandin E 2 release in primary osteoblasts. Am J Physiol Cell Physiol 289: C148 -C158, 2005. First published February 23, 2005 doi:10.1152/ajpcell.00524.2003.-Cells respond to a wide range of mechanical stimuli such as fluid shear and strain, although the contribution of gravity to cell structure and function is not understood. We hypothesized that bone-forming osteoblasts are sensitive to increased mechanical loading by hypergravity. A centrifuge suitable for cell culture was developed and validated, and then primary cultures of fetal rat calvarial osteoblasts at various stages of differentiation were mechanically loaded using hypergravity. We measured microtubule network morphology as well as release of the paracrine factor prostaglandin E 2 (PGE2). In immature osteoblasts, a stimulus of 10ϫ gravity (10 g) for 3 h increased PGE 2 2.5-fold and decreased microtubule network height 1.12-fold without affecting cell viability. Hypergravity (3 h) caused dose-dependent (5-50 g) increases in PGE 2 (5.3-fold at 50 g) and decreases (1.26-fold at 50 g) in microtubule network height. PGE 2 release depended on duration but not orientation of the hypergravity load. As osteoblasts differentiated, sensitivity to hypergravity declined. We conclude that primary osteoblasts demonstrate dose-and duration-dependent sensitivity to gravitational loading, which appears to be blunted in mature osteoblasts. mechanotransduction; differentiation; bone LIFE AS WE KNOW IT HAS EVOLVED under the influence of Earth's gravity. Changes in other physical conditions in the environment (e.g., temperature) have profound effects on cells, and gravity may act at a fundamental level as well. Gravity is ubiquitous and influences the mechanical environment within tissues by affecting cell weight, extracellular hydrostatic pressure, and fluid convection. Despite the central importance of gravity, little is known about how it influences cellular physiology.Because the weight of a cell depends on the amount of gravity acting on its mass, changes in gravity may serve as mechanical stimuli to adherent cells. By applying increased gravity to cells (hypergravity), the direction and magnitude of gravity can be varied to provide insight into its effects on cellular physiology. Coordinated shape and cytoskeletal changes may result from shifts in the position of organelles of different densities within the cell. In a gravity field, the denser nucleus (26,48,49) may shift downward, pulling the interconnected cytoskeleton and plasma membrane with it, resulting in a reduced cell height that would be expected to continue to decrease as the gravity field increased. Microtubules are thought to resist compressive loads (21, 22) and may themselves be compressed as a result of the hydrostatic pressure generated by extracellular fluid. Studies of osteoblasts and endothelial cells have shown that reorganization of the cytoskeleton correlates with increased release of prostagl...