Muscle sarcomere lesions and thrombosis after spaceflight and suspension unloading

Abstract

Spaceflight (flight) and tail suspension-hindlimb unloading (unloaded) produced significant decreases in fiber cross-sectional areas of the adductor longus (AL), a slow-twitch antigravity muscle. However, the mean wet weight of the flight AL muscles was near normal, whereas that of the suspension unloaded AL muscles was significantly reduced. Interstitial edema within the flight AL, but not in the unloaded AL, appeared to account for this apparent disagreement. In both experimental conditions, the slow-twitch oxidative fibers atrophied more than the fast-twitch oxidative-glycolytic and fast-twitch glycolytic fibers. Immunostaining showed that slow-twitch oxidative fibers expressed fast myosin, producing hybrid fibers containing slow and fast myosin isoforms. Two-dimensional gel electrophoresis of flight AL muscles revealed increased content of fast myosin light chains and decreased amounts of slow myosin light chains and fatty acid-binding protein. In the flight AL, absolute mitochondrial content decreased, but the relatively greater breakdown of myofibrillar proteins maintained mitochondrial concentration near normal in the central intermyofibrillar regions of fibers. Subsarcolemmal mitochondria were preferentially lost and reduced below normal concentration. Elevated fiber immunostaining for ubiquitin conjugates was suggestive of ubiquitin-mediated breakdown of myofibrillar proteins. On return to weight bearing for 8-11 h, the weakened atrophic muscles exhibited eccentric contraction-like lesions (hyperextension of sarcomeres with A-band filaments pulled apart and fragmented), tearing of the supporting connective tissue, and thrombosis of the microcirculation. Segmental necrosis of muscle fibers, denervation of neuromuscular junctions, and extravasation of red blood cells were minimal. Lymphocyte antibody markers did not indicate a significant immune reaction. The flight AL exhibited threefold more eccentric-like lesions than the unloaded AL; the high reentry G forces experienced by the flight animals, but not the unloaded group, possibly accounted for this difference. Muscle atrophy appears to increase the susceptibility to form eccentric contraction-like lesions after reloading; this may reflect weakening of the myofibrils and extracellular matrix. Microcirculation was also compromised by spaceflight, such that there was increased formation of thrombi in the post-capillary venules and capillaries. This blockage led to edema by 8-11 h after resumption of weight bearing by the COSMOS 2044 rats. The present findings indicate that defective microcirculation most likely accounted for the extensive tissue necrosis and microhemorrhages observed for COSMOS 1887 rats killed 2 days after landing.

Macrophage invasion does not contribute to muscle membrane injury during inflammation

“…Frozen sections were fixed in acetone, quenched in 0.3% H 2 O 2 in PBS, and then immersed in PBS containing 3% bovine serum albumin, 0.5% Tween-20, and 0.2% gelatin before immunolabeling with one of the following antibodies: (1) anti-ED1 ϩ (diluted 1:50; Bioproducts for Science, Indianapolis, IN), which binds an antigen specific for subpopulations of monocytes and macrophages [23], (2) anti-ED2 ϩ (diluted 1:100; Bioproducts for Science), which binds an antigen specific for a subpopulation of tissue macrophages [23], and (3) anti-W3/13 (diluted 1:100; Bioproducts for Science), which binds an antigen present on neutrophils and T cells; in these preparations T cells are scarce so virtually all of the cells recognized by anti-W3/13 in reloaded muscle are neutrophils [5]. Although anti-ED1 binds an antigen present in both monocytes and a subclass of tissue macrophages [23], in this investigation it is used as a macrophage marker because monocytes would remain in the vasculature and cells located within blood vessels were not included in the data obtained here.…”

“…Morphological observations support this general sequence of events in response to many different types of injuries, including crush injuries, strain injuries, freezing, exposure to toxins, and modified muscle use [3][4][5][6][7]. In each of these instances, the initial defect is thought to be damage to the muscle cell membrane, which allows unregulated influx of calcium into the damaged cell, thereby activating calpains that are capable of proteolyzing a wide variety of structural and regulatory proteins in muscle [8].…”

Nitric oxide synthase inhibition reduces muscle inflammation and necrosis in modified muscle use

“…These disruptions to the normal structure and function of muscle are apparent within minutes after injury and may become more extensive during the following hours to days [2,[4][5][6]. Muscle injury also typically causes edema and the extravasation of neutrophils followed by ED1 ϩ macrophages and later the proliferation of resident ED2 ϩ macrophages [2,5,6].…”

“…In this study, we tested whether modifying NO production in vivo affects muscle inflammation and necrosis or influences the occurrence of inflammatory cell apoptosis during modified muscle use. Rats were subjected to periods of muscle unloading followed by reloading, which has been shown previously to cause muscle inflammation and necrosis [2,5,6] and inflammatory cell apoptosis [29]. Experimental manipulations were performed on animals to which NOS inhibitors were administered, and the concentrations of neutrophils, macrophages, necrotic fibers, and apoptotic inflammatory cells were compared to control animals not receiving NOS inhibitors.…”

Potential exercise countermeasures to attenuate skeletal muscle deterioration in space

“…Important data from Riley et al 11 in the 8-11 hours following space flight revealed substantial eccentric contraction-like damage of muscle fibers in antigravity muscles which included hyperextension of sarcomeres with Aband filaments pulled apart and fragmented. These results have been replicated in a number of microgravity and simulated microgravity conditions (for a review see Riley et Table 1.…”