Transcription Factors in Muscle Atrophy Caused by Blocked Neuromuscular Transmission and Muscle Unloading In Rats

Abstract

The muscle wasting associated with long-term intensive care unit (ICU) treatment has a negative effect on muscle function resulting in prolonged periods of rehabilitation and a decreased quality of life. To identify mechanisms behind this form of muscle wasting, we have used a rat model designed to mimic the conditions in an ICU. Rats were pharmacologically paralyzed with a postsynaptic blocker of neuromuscular transmission, and mechanically ventilated for one to two weeks, thereby unloading the limb muscles. Transcription factors were analyzed for cellular localization and nuclear concentration in the fast-twitch muscle extensor digitorum longus (EDL) and in the slow-twitch soleus. Significant muscle wasting and upregulation of mRNA for the ubiquitin ligases MAFbx and MuRF1 followed the treatment. The IκB family-member Bcl-3 displayed a concomitant decrease in concentration, suggesting altered κB controlled gene expression, although NFκB p65 was not significantly affected. The nuclear levels of the glucocorticoid receptor (GR) and the thyroid receptor α1 (TRα1) were altered and also suggested as potential mediators of the MAFbx-and MuRF1-induction in the absence of induced Foxo1. We believe that this model, and the strategy of quantifying nuclear proteins, will provide a valuable tool for further, more detailed, analyses of the muscle wasting occurring in patients kept on a mechanical ventilator. variations in the response. For example, cachexia associated with disease states such as cancer and sepsis involves an increase in inflammatory cytokine production, which activates transcription factors that are, at least partly, distinct from those activated by disuse (12). How, and if, these differences affect the net result of the atrophic process is still unclear.Muscle wasting and impaired muscle function impose a risk to critically ill ICU patients during treatment. Specifically, neuromuscular abnormalities have been reported as the dominant cause for the reduced quality of life in critically ill ICU survivors, and are remaining even up to two years after hospital discharge (13,14). Therefore, our research is focused on the molecular events mediating the muscle atrophy in ICU patients, because successful treatment of this side effect would be highly beneficial for the survival, recovery, and quality of life in these patients. To address this issue, we have used an experimental rat model mimicking the conditions for many ICU patients, such as mechanical ventilation for long durations (weeks), postsynaptic block of neuromuscular transmission to facilitate ventilation, and unloading of muscles. In this study, the levels of a number of transcription factors in nuclear extracts from muscle tissue, and their spatial organization in the muscle fiber, have been investigated. The transcription factors we selected for investigation had been implicated in the expression of MAFbx and MuRF1 or as downstream effectors of signal transduction pathways previously linked to muscle atrophy or in the regulation of differentiation an...

Mechanisms underlying ICU muscle wasting and effects of passive mechanical loading

“…In this study, all mechanically ventilated, sedated and immobilized ICU patients had lower than normal myosin:actin ratios. This is consistent with our previous studies using a rat experimental ICU model; that is, a preferential myosin loss in both fast-twitch or slow-twitch muscles and fiber types was observed in response to mechanical ventilation, sedation and immobilization at durations longer than 5 days [8,35,36]. …”

“…However, the myosin loss is not only caused by enhanced degradation but also by decreased synthesis, as indicated by a significant downregulation of contractile proteins at the transcriptional level. This is consistent with previous observations in patients with CIM and in experimental ICU models [5,6,8,35,36]. The sparing of the thin filament protein actin in spite of a similar downregulation at the transcriptional level has been suggested to be secondary to differences in protein turnover rate or the upregulation of the small αB-crystalline chaperone protecting actin from degradation [8].…”

Myofibrillar protein and gene expression in acute quadriplegic myopathy

“…In AQM, muscles are not depolarized due to the post-synaptic blockade of neuromuscular transmission, an in-excitable muscle membrane, or sedation or a combination of these factors. Further, the lack of both depolarization and weight bearing result in unloading of cytoskeletal and contractile proteins, which results in the activation of specific intracellular signaling pathways, severe muscle wasting and a preferential loss of myosin and myosin-associated proteins in patients with AQM, as well as in an experimental rodent AQM model, separating this muscle atrophy from other muscle wasting conditions [19,42,69,70]. Thus, a difference in the sensitivity to contractile and mechanical loading is forwarded as one possible mechanism underlying the relative sparing of intrafusal muscle fibers and craniofacial muscles.…”

The role and regulation of MAFbx/atrogin-1 and MuRF1 in skeletal muscle atrophy

“…MAFbx and/ or MuRF1 mRNA are up-regulated following immobilisation in humans [11,37,46,95,96], hibernating squirrels [15], mice [9] and rats [4,38,47,48,78,100,112]. The increases in MAFbx and MuRF1 are associated with immobilisation-induced increases in proteosomedependent proteolysis but not inhibition of protein synthesis [48].…”