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Microgravity Stress: Bone and Connective Tissue

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ABSTRACT

The major alterations in bone and the dense connective tissues in humans and animals exposed to microgravity illustrate the dependency of these tissues' function on normal gravitational loading. Whether these alterations depend solely on the reduced mechanical loading of zero g or are compounded by fluid shifts, altered tissue blood flow, radiation exposure, and altered nutritional status is not yet well defined. Changes in the dense connective tissues and intervertebral disks are generally smaller in magnitude but occur more rapidly than those in mineralized bone with transitions to 0 g and during recovery once back to the loading provided by 1 g conditions. However, joint injuries are projected to occur much more often than the more catastrophic bone fracture during exploration class missions, so protecting the integrity of both tissues is important. This review focuses on the research performed over the last 20 years in humans and animals exposed to actual spaceflight, as well as on knowledge gained from pertinent ground‐based models such as bed rest in humans and hindlimb unloading in rodents. Significant progress has been made in our understanding of the mechanisms for alterations in bone and connective tissues with exposure to microgravity, but intriguing questions remain to be solved, particularly with reference to biomedical risks associated with prolonged exploration missions. © 2016 American Physiological Society. Compr Physiol 6:**‐**, 2016.

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Figure 1. Figure 1. A typical long bone: anatomical regions, envelopes, compartments and bone cells. Epiphyses are distal and proximal to growth plates while they are still open, but fuse with metaphyseal compartments once the growth plates close. The major bone “envelopes” or surfaces are the periosteum, endosteum (any bone surface bordering bone marrow in the medullary canal; also called “endocortical” if referring to inner border of cortical shaft), and intracortical Haversian canals, within which are housed blood vessels and nerves. The insert at top right is an enlargement of cancellous bone just below the growth plate. In the primary spongiosa adjacent to the growth plate, cartilage spicules are gradually mineralized (black); secondary spongiosa refers to the lattice‐work of cancellous rods and plates that are fully mineralized. The cross‐section of the cortical mid‐diaphysis illustrates multiple Haversian systems (osteons) and multiple osteocytes in lacunae, connected by canaliculi. Bone cells illustrated at lower right are (from left) flat, inactive lining cells, active osteoblasts (bone‐forming cells), osteocytes embedded in the mineralized matrix, and an active osteoclast, residing in the Howship's lacuna it has created by its resorptive action. Figure from Morey‐Holton et al., 1996 (), used with permission.
Figure 2. Figure 2. BFR, as measured by fluorochrome label administration, decreases dramatically in rodents subjected to hindlimb unloading, simulating the disuse of microgravity. Calcein, injected 2 and 9 days before the end of the experiment, binds to circulating calcium and effectively labels those bone surfaces that are actively mineralizing in the 36 to 48 h postinjection. Measuring the average distance between labels yields mineral apposition rate (MAR); quantitating surfaces with single and double labels yields % mineralizing surface (%MS/BS). The product of these two measures is BFR. Skeletally mature rats exhibit 80% to 90% declines in BFR within 2 weeks of hindlimb unloading.
Figure 3. Figure 3. Fluid shear stress on osteocytes and/or their dendritic processes is the key mechanical signal that is transduced to a biochemical response, resulting in the stimulation of osteoblast activity and new bone formation. Only slight bone deformation is required to generate areas of tension and compression, which creates movement of interstitial fluid within the millions of canalicula connecting lacunae housing osteocytes, thought to be the key mechanosensors inside bone. Figure from Duncan & Turner, 1995 (), used with permission.
Figure 4. Figure 4. Proximal femur strength deteriorates over 4.5 to 6 months in low earth orbit. Panel A illustrates sample finite element model (FEM) of one crew member's proximal femur derived from QCT data illustrating loading in (a) single‐leg standing and (b) a fall to the side impacting the greater trochanter. In panel B, individual data for 13 ISS crew members (40‐55 years, 12 males, 1 female) are detailed for pre‐ and postflight FEM‐computed proximal femur strength normalized to body weight (FFE/BW) for both stance (a) and fall (b) loading. The dotted lines represent median values for femoral neck strength assessed by the same patient‐specific FEM procedure in populations of healthy premenopausal (36‐45 years; top line) and elderly (62‐81 years; bottom line) Caucasian women. Figure from Keyak et al., 2009 (), used with permission.
Figure 5. Figure 5. Foot forces aboard ISS are much reduced compared to 1 g. In‐shoe peak force magnitudes recorded during activities of daily living on Earth and aboard ISS. Both magnitude and frequency of the GRFs observed in 1 g are reduced onboard ISS. No GRF's > 2 × BW were recorded in microgravity (as seen in 1 g), with the vast majority of peak forces during running in 0 g at 1.6 × BW or less. Figure from Cavanagh et al., 2010 (), used with permission.
Figure 6. Figure 6. Recovery of proximal femur (trochanter) BMD in Shuttle crew members takes many‐fold longer than short duration of mission (<23 days). Dotted lines represent 95% confidence limits for the BMD data. Immediate postflight BMD deficits were ∼8% of preflight levels; 50% recovery time (dashed vertical line) was estimated to be ∼8.5 months. Repeat fliers' data are denoted by “+” signs. Figure from Sibonga et al., 2007 (), used with permission.
Figure 7. Figure 7. Partial weight‐bearing (A) and hindlimb suspension (B) rodent models. Partial weight‐bearing ranging from 16% to 70% of total body weight can be achieved by daily adjustment of tension on the spring above the triangular harness support. In the traditional hindlimb (tail) suspension model, hindlimbs are elevated to disallow weight bearing without immobilization; the head‐down posture also induces a cephalic fluid shift. Figure from Ellman et al., 2013 (), used with permission.
Figure 8. Figure 8. Graded reductions in weight bearing produce graded reductions in bone volume. Micro computed tomography results obtained at (A) distal femur metaphysis and (B) femur mid‐diaphysis. HLU = hindlimb unloading; PWB = partial weight bearing at indicated %BW; CON = control; * = different from CON (P < 0.05); # = HLU different from PWB100 (P < 0.05); brackets = pairwise differences between PWB groups. Figure adapted from Ellman et al., 2013 (), used with permission.
Figure 9. Figure 9. Radiation produces dramatic reductions in cancellous bone volume and altered microarchitecture. Micro‐CT images of the proximal tibia in young mice that were (A) SHAM exposed, (B) exposed to 2 Gy gamma radiation, or (C) exposed to 2 Gy proton radiation. Both gamma and proton exposure led to decreased trabecular volume fraction (−29 and −35%, respectively) and increased trabecular spacing (+19% and +21%, respectively). Figure adapted from Hamilton et al., 2006 (), used with permission.
Figure 10. Figure 10. Proposed radiation‐induced inflammatory cascade leading to increased fracture risk following crew member exposure to radiation. Rapidly dividing marrow cells in exposed bone are damaged and die, in part from damage from ROS (OH, O2 ); phagocytes are then recruited to remove dead cells. These phagocytes also release proinflammatory cytokines like TNF‐α and IL‐1, which in turn stimulate osteoclasts directly or indirectly by stimulating receptor activator of NF‐κβ ligand (RANKL) release from nearby osteoblasts. Accelerated loss of bone by osteoclastic resorption contributes to impaired bone structural integrity, elevating fracture risk for the affected individual. Figure from Willey et al., 2011 (), used with permission.
Figure 11. Figure 11. Sclerostin regulation of canonical Wnt signaling. Binding of Wnt to the frizzled receptor complexed with low‐density lipoprotein 5 or 6 (Lrp5/Lrp6) receptor complex initiates a signaling cascade resulting in B‐catenin translocating to the nucleus, where it activates transcription factors regulating Wnt target genes. Wnt pathway activation promotes osteoblast proliferation, maturation and differentiated activity and, via mature osteoblast production of osteoprotogerin, decreasing osteoclast differentiation and activation (). With reduced mechanical loading, osteocyte production of sclerostin increases, blocking Wnt binding and resulting ultimately in reduced osteoblast and increased osteoclast activity.


Figure 1. A typical long bone: anatomical regions, envelopes, compartments and bone cells. Epiphyses are distal and proximal to growth plates while they are still open, but fuse with metaphyseal compartments once the growth plates close. The major bone “envelopes” or surfaces are the periosteum, endosteum (any bone surface bordering bone marrow in the medullary canal; also called “endocortical” if referring to inner border of cortical shaft), and intracortical Haversian canals, within which are housed blood vessels and nerves. The insert at top right is an enlargement of cancellous bone just below the growth plate. In the primary spongiosa adjacent to the growth plate, cartilage spicules are gradually mineralized (black); secondary spongiosa refers to the lattice‐work of cancellous rods and plates that are fully mineralized. The cross‐section of the cortical mid‐diaphysis illustrates multiple Haversian systems (osteons) and multiple osteocytes in lacunae, connected by canaliculi. Bone cells illustrated at lower right are (from left) flat, inactive lining cells, active osteoblasts (bone‐forming cells), osteocytes embedded in the mineralized matrix, and an active osteoclast, residing in the Howship's lacuna it has created by its resorptive action. Figure from Morey‐Holton et al., 1996 (), used with permission.


Figure 2. BFR, as measured by fluorochrome label administration, decreases dramatically in rodents subjected to hindlimb unloading, simulating the disuse of microgravity. Calcein, injected 2 and 9 days before the end of the experiment, binds to circulating calcium and effectively labels those bone surfaces that are actively mineralizing in the 36 to 48 h postinjection. Measuring the average distance between labels yields mineral apposition rate (MAR); quantitating surfaces with single and double labels yields % mineralizing surface (%MS/BS). The product of these two measures is BFR. Skeletally mature rats exhibit 80% to 90% declines in BFR within 2 weeks of hindlimb unloading.


Figure 3. Fluid shear stress on osteocytes and/or their dendritic processes is the key mechanical signal that is transduced to a biochemical response, resulting in the stimulation of osteoblast activity and new bone formation. Only slight bone deformation is required to generate areas of tension and compression, which creates movement of interstitial fluid within the millions of canalicula connecting lacunae housing osteocytes, thought to be the key mechanosensors inside bone. Figure from Duncan & Turner, 1995 (), used with permission.


Figure 4. Proximal femur strength deteriorates over 4.5 to 6 months in low earth orbit. Panel A illustrates sample finite element model (FEM) of one crew member's proximal femur derived from QCT data illustrating loading in (a) single‐leg standing and (b) a fall to the side impacting the greater trochanter. In panel B, individual data for 13 ISS crew members (40‐55 years, 12 males, 1 female) are detailed for pre‐ and postflight FEM‐computed proximal femur strength normalized to body weight (FFE/BW) for both stance (a) and fall (b) loading. The dotted lines represent median values for femoral neck strength assessed by the same patient‐specific FEM procedure in populations of healthy premenopausal (36‐45 years; top line) and elderly (62‐81 years; bottom line) Caucasian women. Figure from Keyak et al., 2009 (), used with permission.


Figure 5. Foot forces aboard ISS are much reduced compared to 1 g. In‐shoe peak force magnitudes recorded during activities of daily living on Earth and aboard ISS. Both magnitude and frequency of the GRFs observed in 1 g are reduced onboard ISS. No GRF's > 2 × BW were recorded in microgravity (as seen in 1 g), with the vast majority of peak forces during running in 0 g at 1.6 × BW or less. Figure from Cavanagh et al., 2010 (), used with permission.


Figure 6. Recovery of proximal femur (trochanter) BMD in Shuttle crew members takes many‐fold longer than short duration of mission (<23 days). Dotted lines represent 95% confidence limits for the BMD data. Immediate postflight BMD deficits were ∼8% of preflight levels; 50% recovery time (dashed vertical line) was estimated to be ∼8.5 months. Repeat fliers' data are denoted by “+” signs. Figure from Sibonga et al., 2007 (), used with permission.


Figure 7. Partial weight‐bearing (A) and hindlimb suspension (B) rodent models. Partial weight‐bearing ranging from 16% to 70% of total body weight can be achieved by daily adjustment of tension on the spring above the triangular harness support. In the traditional hindlimb (tail) suspension model, hindlimbs are elevated to disallow weight bearing without immobilization; the head‐down posture also induces a cephalic fluid shift. Figure from Ellman et al., 2013 (), used with permission.


Figure 8. Graded reductions in weight bearing produce graded reductions in bone volume. Micro computed tomography results obtained at (A) distal femur metaphysis and (B) femur mid‐diaphysis. HLU = hindlimb unloading; PWB = partial weight bearing at indicated %BW; CON = control; * = different from CON (P < 0.05); # = HLU different from PWB100 (P < 0.05); brackets = pairwise differences between PWB groups. Figure adapted from Ellman et al., 2013 (), used with permission.


Figure 9. Radiation produces dramatic reductions in cancellous bone volume and altered microarchitecture. Micro‐CT images of the proximal tibia in young mice that were (A) SHAM exposed, (B) exposed to 2 Gy gamma radiation, or (C) exposed to 2 Gy proton radiation. Both gamma and proton exposure led to decreased trabecular volume fraction (−29 and −35%, respectively) and increased trabecular spacing (+19% and +21%, respectively). Figure adapted from Hamilton et al., 2006 (), used with permission.


Figure 10. Proposed radiation‐induced inflammatory cascade leading to increased fracture risk following crew member exposure to radiation. Rapidly dividing marrow cells in exposed bone are damaged and die, in part from damage from ROS (OH, O2 ); phagocytes are then recruited to remove dead cells. These phagocytes also release proinflammatory cytokines like TNF‐α and IL‐1, which in turn stimulate osteoclasts directly or indirectly by stimulating receptor activator of NF‐κβ ligand (RANKL) release from nearby osteoblasts. Accelerated loss of bone by osteoclastic resorption contributes to impaired bone structural integrity, elevating fracture risk for the affected individual. Figure from Willey et al., 2011 (), used with permission.


Figure 11. Sclerostin regulation of canonical Wnt signaling. Binding of Wnt to the frizzled receptor complexed with low‐density lipoprotein 5 or 6 (Lrp5/Lrp6) receptor complex initiates a signaling cascade resulting in B‐catenin translocating to the nucleus, where it activates transcription factors regulating Wnt target genes. Wnt pathway activation promotes osteoblast proliferation, maturation and differentiated activity and, via mature osteoblast production of osteoprotogerin, decreasing osteoclast differentiation and activation (). With reduced mechanical loading, osteocyte production of sclerostin increases, blocking Wnt binding and resulting ultimately in reduced osteoblast and increased osteoclast activity.
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Susan A. Bloomfield, Daniel A. Martinez, Ramon D. Boudreaux, Anita V. Mantri. Microgravity Stress: Bone and Connective Tissue. Compr Physiol 2016, 6: 645-686. doi: 10.1002/cphy.c130027