|Home | About | Journals | Submit | Contact Us | Français|
To review current knowledge about the impact of prolonged mechanical ventilation on diaphragmatic function and biology.
Systematic literature review.
Prolonged mechanical ventilation can promote diaphragmatic atrophy and contractile dysfunction. As few as 18 hrs of mechanical ventilation results in diaphragmatic atrophy in both laboratory animals and humans. Prolonged mechanical ventilation is also associated with diaphragmatic contractile dysfunction. Studies using animal models revealed that mechanical ventilation-induced diaphragmatic atrophy is due to increased diaphragmatic protein breakdown and decreased protein synthesis. Recent investigations have identified calpain, caspase-3, and the ubiquitin-proteasome system as key proteases that contribute to mechanical ventilation-induced diaphragmatic proteolysis. The scientific challenge for the future is to delineate the mechanical ventilation-induced signaling pathways that activate these proteases and depress protein synthesis in the diaphragm. Future investigations that define the signaling mechanisms responsible for mechanical ventilation-induced diaphragmatic weakness will provide the knowledge required for the development of new medicines that can maintain diaphragmatic mass and function during prolonged mechanical ventilation.
Mechanical ventilation (MV) is used clinically to sustain pulmonary gas exchange in patients who are incapable of maintaining sufficient alveolar ventilation. Common indications for MV include respiratory failure due to chronic obstructive pulmonary disease, status asthmaticus, and/or heart failure. MV is also required for many patients with neuromuscular diseases, drug overdoses, and during postsurgical recovery.
Although MV is a lifesaving measure, prolonged MV is often associated with complications, such as tracheal injuries, infection, cardiovascular failure, and lung injury. Abundant evidence also indicates that prolonged MV promotes diaphragmatic weakness due to both atrophy and contractile dysfunction. The detrimental impact of prolonged MV on the diaphragm has been termed ventilator-induced diaphragmatic dysfunction (VIDD) and this phenomenon has received increased research attention in recent years. This review will summarize the current knowledge about the impact of prolonged MV on diaphragmatic function and biology. Our approach will be to provide a synopsis of major concepts and highlight the findings of selected individual studies. Furthermore, in hopes of stimulating additional research, we will highlight specific areas where additional work is warranted. We begin with an overview of the clinical significance of VIDD followed with a discussion of the impact of prolonged MV on diaphragm structure, function, and gene expression.
Although many patients requiring MV can be extubated in <3 days, ~30% of MV patients will experience weaning problems and will require additional time on the ventilator (1). In patients experiencing weaning problems, weaning procedures account for 40% to 60% of the total time on the ventilator (2). Unfortunately, 1% to 5% of all MV patients repeatedly fail attempts to be weaned from MV and therefore may become ventilator dependent (3). Chronic ventilator dependence is a major medical problem and presents significant psychological and financial problems for both patients and families (3).
Difficult weaning has received considerable clinical attention during the past 20 yrs as numerous studies have attempted to determine clinical predictors of weaning problems and to develop strategies to improve the weaning success rate. The pathophysiological problems that contribute to difficult weaning are complex and numerous (4). To further complicate this issue, the factors that contribute to weaning difficulties vary among patients depending on age, existing comorbidities, and nutritional status. Nonetheless, several key factors have been identified as potential contributors to difficult weaning. These include inadequate ventilatory drive, increased work of breathing, cardiac failure, and inspiratory muscle weakness and/or fatigue (4). Of these factors, growing evidence suggests that inspiratory muscle weakness and fatigue are likely significant contributors to weaning problems (5–9). Therefore, although MV is a lifesaving intervention for countless patients, it seems likely that VIDD is an important contributor to weaning problems in many patients. It follows that developing methods to protect patients against weaning problems based on an understanding of the mechanisms responsible for MV-induced diaphragmatic weakness is an important topic for future research.
Controlled mechanical ventilation (CMV) is a mode of MV whereby the ventilator provides all of the work of breathing and patient triggering of the ventilator is not possible. Clinical indications for CMV include general anesthesia, drug overdoses, coma, and spinal cord injuries (10). Numerous studies have demonstrated that prolonged CMV results in a rapid-onset of diaphragmatic atrophy in several species (e.g., rats, rabbits, pigs) (11–16). For example, as few as 12 hrs to 18 hrs of CMV results in significant fiber atrophy in both slow and fast muscle fibers of the rat diaphragm (Fig. 1) (13, 16, 17). In contrast, the limb muscles of mechanically ventilated animals show no signs of atrophy after 12 hrs to 18 hrs of inactivity (13, 16). The rapid rate of MV-induced diaphragmatic atrophy greatly exceeds the time course of atrophy observed in locomotor skeletal muscles during periods of disuse (18). Specifically, it would take approximately 96 hrs to achieve the same level of atrophy in unloaded locomotor skeletal muscles as observed in the diaphragm after 12 hrs of CMV (18). Furthermore, the rate of CMV-induced atrophy exceeds that reported for the diaphragm after denervation (19). Hence, CMV-induced diaphragmatic atrophy is an extremely rapid and unique type of skeletal muscle wasting.
Similar to the findings in animal studies, Levine et al confirmed that 18 hrs to 69 hrs of CMV also results in atrophy of the human diaphragm (20). This landmark study demonstrated that, compared with diaphragm biopsy samples from control subjects, diaphragm fibers from mechanically ventilated subjects showed a large decrease in fiber cross-sectional area of both type I and type II fibers. Specifically, 18 hrs to 69 hrs of CMV resulted in a decreased diaphragm fiber cross-sectional area of 57% and 53%, respectively, in type I and type II fibers (Figs. 2 and and3).3). This magnitude of CMV-induced human diaphragmatic atrophy is similar to that reported for the rat diaphragm after 48 hrs of CMV (21).
Although it is clear that prolonged CMV promotes a rapid onset of diaphragmatic atrophy, it is unknown whether pressure-support MV can retard or prevent ventilator-induced diaphragmatic atrophy. However, two recent reports suggested that MV using a pressure-support mode can limit ventilator-induced protein catabolism (22, 23). Nonetheless, neither of these studies performed measurements of diaphragmatic atrophy.
Animal studies revealed that CMV results in time-dependent alterations in the ultrastructure of diaphragm muscle fibers (24–27). In contrast, inactivity does not promote ultrastructure damage in locomotor muscles of mechanically ventilated rats (24). In reference to MV-induced changes in diaphragmatic structure, CMV results in diverse areas of abnormal diaphragmatic myofibrils as indicated by myofibrillar disarray and alterations in Z-line structure (24). Furthermore, prolonged CMV also promotes focal areas of diaphragm fiber regeneration without signs of inflammation (27, 28). Finally, prolonged CMV (i.e., 3 days) also results in an increase in cytoplasmic lipid vacuoles (26, 27). However, it is unclear whether these vacuoles represent a pathologic or a physiologic adaptive process. In several forms of myopathy, cytoplasmic vacuoles represent secondary lysosomes involved in the autophagic process (29, 30). Currently, the role of autophagy in CMV-induced changes in diaphragmatic ultrastructure remains unknown.
To date, limited information exists regarding the impact of prolonged CMV on respiratory muscles other than the diaphragm. However, recent evidence indicated that prolonged CMV promotes damage to the external intercostal muscles (26). In this regard, the pattern of CMV-induced ultrastructual damage to the external intercostal muscle fibers seems similar to MV-induced diaphragmatic damage as demonstrated by focal areas of fragmented myofibrils along with increases in lipid vacuoles (26).
The first report that CMV promotes diaphragmatic contractile dysfunction appeared in 1994 (21). Le Bourdelles et al, using a rat model of mechanical ventilation, demonstrated that 48 hrs of CMV results in a large (i.e., ~40%) reduction in diaphragmatic maximal tetanic specific force production (i.e., force per cross-sectional area) (21). Since this initial observation, numerous studies using several animal species (i.e., rats, rabbits, pigs, baboons) have confirmed that prolonged CMV results in diaphragmatic contractile dysfunction (12, 24, 25, 28, 31–38). Prolonged CMV promotes a time-dependent and progressive decrease in diaphragmatic specific force production at both submaximal and maximal stimulation frequencies (35) (Fig. 4). Identical to the time course of CMV-induced atrophy, only 12 hrs of CMV is required to promote significant reductions in diaphragmatic-specific force production (35, 37, 38).
As aging impairs diaphragmatic contractile function (39), it is feasible that the senescent diaphragm may be more sensitive to CMV-induced diaphragmatic dysfunction. To address this issue, Criswell et al investigated the cumulative effects of aging and CMV on diaphragmatic function in young adult and senescent rats. Their findings indicated that aging does not exacerbate the relative CMV-induced impairment in diaphragmatic-specific force production (37). However, compared with nonventilated young adult animals, maximal diaphragmatic-specific force production was 13% lower in senescent rats. Therefore, despite the similar relative responses of young and old diaphragms to CMV, the negative effects of MV are additive to the age-related deficit in diaphragmatic contractile performance (Fig. 5). This additive effect of aging and CMV-induced diaphragmatic contractile dysfunction may explain why patient age is an independent predictor of difficulties in patient weaning (40).
The aforementioned animal models of MV have consistently shown that CMV leads to depressed diaphragmatic force production. However, many MV patients are also treated with drugs that can exacerbate the impact of CMV on diaphragmatic function (e.g., neuromuscular blockers and/or glucocorticoids). For example, prolonged administration of non-depolarizing neuromuscular blocking agents are used in ~13% of MV patients (41). These agents are typically used to facilitate MV, assist in the treatment of intracranial pressure, and reduce oxygen consumption (42). A possible complication of using neuromuscular blocking agents in the intensive care unit is skeletal muscle myopathy. In this regard, two recent studies confirmed that use of the neuromuscular blocking agent rocuronium exacerbates CMV-induced contractile dysfunction in rats (31, 43).
Also, many patients suffering from acute respiratory failure are treated with corticoidsteroids for the underlying lung disease or other diseases (44). This is significant because glucocorticoid treatment is associated with steroid-induced myopathy of both locomotor and respiratory muscles (45). To investigate the extent to which corticosteroids contribute to diaphragm weakness in mechanically ventilated rats, Maes et al studied the combined effects of corticosteroid (methylprednisolone) treatment on diaphragm function in rats after 24 hrs of CMV (44). Surprisingly, their results revealed that corticosteroid treatment provided partial protection against CMV-induced diaphragmatic atrophy and contractile dysfunction. This protective action of glucocorticoids seems to be associated with the inhibition of the protease calpain (44).
Although CMV remains a common mode of MV for use with specific patient populations (e.g., drug overdose, surgery), other MV modes are often used in the adult intensive care unit. Specifically, modes of MV that provide partial ventilatory support are commonly used with adult patients inflicted with respiratory failure (10). The key question becomes: Do ventilatory modes that provide partial support for the work of breathing result in the same magnitude of diaphragmatic contractile dysfunction as that observed with CMV? Sassoon et al addressed this issue and reported that, compared with CMV, assist-control MV partially attenuates the diaphragmatic contractile dysfunction associated with CMV (22). Specifically, after 3 days of MV in rabbits, peak diaphragmatic power output decreased 41% with CMV but declined only by 20% with assist-control MV. Hence, compared with CMV, modes of MV that provide partial ventilatory support seem to reduce the magnitude of VIDD.
Further evidence that diaphragmatic inactivity is a key factor in promoting CMV-induced VIDD has been provided by Gayan-Ramirez et al (14). This group demonstrated that short (i.e., 5-min) periods of intermittent spontaneous breathing during CMV can retard the damaging effects of CMV on diaphragmatic contractile dysfunction (14). These findings reinforce the concept that diaphragmatic inactivity is an important factor in promoting VIDD.
To date, there are no published reports that have directly evaluated the impact of prolonged MV on diaphragmatic contractile function in humans. Nonetheless, respiratory muscle weakness is a well-known problem in the intensive care unit (4). Many studies indicated that, compared with controls, maximal inspiratory pressure is markedly lower in patients after prolonged MV (5). Furthermore, patients who fail to wean from the ventilator have significantly weaker inspiratory muscles compared with patients who can be weaned from MV (9). Nonetheless, in patient populations, it is often difficult to determine the direct effects of MV on respiratory muscle function vs. the impact of the patient’s illness (e.g., infection).
Several studies using laboratory animals have demonstrated that prolonged CMV depresses diaphragmatic protein synthesis and accelerates protein breakdown. For example, as few as 6 hrs of CMV is associated with a 30% decrease in mixed protein synthesis and a 65% decline in the rate of myosin heavy-chain protein synthesis (46). These depressed levels of protein synthesis remained consistent throughout 18 hrs of CMV.
Furthermore, animal studies have consistently reported that CMV results in increased diaphragmatic proteolysis. Specifically, in vitro measurements of diaphragmatic protein breakdown reveal that 12 hrs to 18 hrs of CMV results in a large increase (i.e., >46%) in diaphragmatic proteolysis (13, 38). Furthermore, numerous studies have demonstrated that prolonged CMV activates several proteases in the diaphragm including calpain, caspase-3, and the ubiquitin proteasome system of proteolysis (13, 16, 22, 33, 38, 47).
At present, no studies have performed direct measurements of CMV-induced diaphragmatic proteolysis in humans. However, Levine et al reported that 18 hrs to 69 hrs of CMV increases caspase-3 activity in the diaphragm along with increased messenger RNA (mRNA) for two key components of the ubiquitin-proteasome system of proteolysis (20). These findings in humans mimic the numerous animal studies and are consistent with the notion that prolonged CMV results in a significant increase in proteolysis in the human diaphragm.
CMV lasting ≥6 hrs results in diaphragmatic redox disturbances as demonstrated by increases in biomarkers of oxidative injury (i.e., protein carbonyls and lipid peroxidation) (13, 38, 48, 49). For example, key contractile proteins, such as actin and myosin, are oxidized in the diaphragm during prolonged CMV (49). This redox disturbance occurs because prolonged CMV results in increased reactive oxygen species (ROS) production and a diminished antioxidant capacity in the diaphragm (48). The pathways responsible for CMV-induced ROS production in the diaphragm remain unclear but nitric oxide synthase and nicotinamide adenine dinucleotide phosphate oxidase are not strong candidates (50, 51). Although xanthine oxidase is a source of ROS production in the diaphragm during CMV, it is clear that additional unknown sources also exist (52). Therefore, by elimination, it is feasible that mitochondria are an important source of diaphragmatic ROS production during prolonged MV.
In addition to increased ROS production, prolonged CMV also results in a diminished total antioxidant capacity in the diaphragm as evidenced by decreases in glutathione levels and diminished glutathione peroxidase and CuZn superoxide dismutase levels (48). Interestingly, the changes in protein abundance of these antioxidant enzymes are not associated with alterations in the mRNA level for either enzyme. Therefore, factors other than the amount of mRNA expression dictate the abundance of these antioxidant proteins in the diaphragm during prolonged CMV. It follows that decreased rates of mRNA translation and/or enhanced degradation of the antioxidant protein could be responsible for the dissociation between mRNA and diaphragmatic protein levels (48).
Growing evidence indicates that prevention of CMV-induced oxidative damage to the diaphragm can retard the deleterious effects of prolonged MV on the diaphragm. Treatment of animals with an antioxidant (i.e., Trolox, water-soluble vitamin E analog) can impede CMV-induced diaphragmatic atrophy and contractile dysfunction (17, 33, 38). Prevention of oxidative stress may protect the diaphragm against atrophy and contractile dysfunction through a variety of signaling mechanisms including the regulation of proteolytic signaling pathways (53–55). Furthermore, because the transcription of many genes is under redox control, CMV-induced redox disturbances could have a profound effect on diaphragmatic gene expression.
Recent studies have explored the impact of prolonged MV on diaphragmatic gene expression. Using a gene chip approach in a rat model of MV, Deruisseau et al examined the effect of both short-(6 hrs) and long-term (18 hrs) CMV on diaphragmatic gene expression (56). Compared with diaphragms from control animals, microarray analysis revealed that MV resulted in 354 differentially expressed gene products after 6 hrs to 18 hrs of CMV (56). In general, both stress response and proteolytic genes were up-regulated in the diaphragm, whereas genes in the structural protein and energy metabolism categories were down-regulated.
Diaphragmatic genes that exhibit a large MV-induced increase in expression include heme oxygenase-1 (+19-fold) and metallothionein-1 and -2 (+26-fold) (56). The physiologic significance of these changes remains speculative but it seems likely that both of these genes are up-regulated in response to CMV-induced oxidative stress in the diaphragm. In reference to heme oxygenase-1, this enzyme catalyzes the rate-limiting step in the degradation of heme, resulting in the generation of carbon monoxide, biliverdin, and free iron (Fe2+) (57). Furthermore, biliverdin is then reduced to bilirubin via biliverdin reductase. Both bilirubin and biliverdin exhibit antioxidant effects and, therefore, it is feasible that HO-1 induction in the diaphragm plays a protective role against MV-induced oxidative stress (57).
Metallothionein gene expression is induced by a variety of stimuli including metal exposure and oxidative stress (58, 59). Metallothionein belongs to a family of cysteine-rich, low-molecular weight proteins. These proteins have several physiologic functions including the binding of several metals (e.g., zinc, copper, cadmium) and the scavenging of ROS (58). Therefore, it seems that the CMV-induced expression of metallothionein in the diaphragm may represent an attempt by diaphragm fibers to preserve redox balance by increasing the level of cellular antioxidants.
It is also clear that CMV promotes large increases in the expression of key proteins involved in the ubiquitin-proteasome proteolytic system. CMV results in increased diaphragmatic levels of both atrogin-1 mRNA (eight-fold increase) and muscle ring finger-1 mRNA (19-fold increase) (56). These two muscle-specific E3 ligases are known to play an important role in disuse muscle atrophy (54).
Many diaphragmatic genes involved in energy metabolism (e.g., fat metabolism and oxidative phosphorylation) are down-regulated during CMV (56). Furthermore, diaphragmatic genes involved in calcium homeostasis are also down-regulated during CMV including calsequestrin 2 and sarco(endo)plasmic reticulum calcium ATPase (SERCA) pumps (60). A decrease in the expression of these proteins would likely contribute to increased cytosolic levels of free calcium and, therefore, promote the calcium-induced activation of the calcium-activated proteolytic enzyme calpain (61).
Also, CMV results in decreased insulin-like growth factor-1 mRNA in the diaphragm (15). This is significant because signaling initiated by insulin-like growth factor-1 exerts an important influence on muscle fiber size (62). Specifically, insulin-like growth factor-1 binding to its receptor results in the phosphorylation of phosphatidylinositol 3 kinase and subsequent activation of protein kinase B (Akt) (63). Activated (i.e., phosphorylated) Akt exerts an important regulatory role in the maintenance of skeletal muscle fiber size, in part, because active Akt promotes protein synthesis and represses protein degradation (63). Numerous studies revealed that decreased Akt activation is a hallmark of many forms of locomotor skeletal muscle atrophy (63). In regard to the impact of prolonged CMV on Akt activation in the diaphragm, McClung et al demonstrated that 18 hrs of CMV results in a down-regulation of insulin-like growth factor-1 to Akt signaling (17).
Finally, CMV also results in altered diaphragmatic expression of several myogenic regulatory factors including myogenin (mRNA increased), MyoD (mRNA decreased), and myf-5 (mRNA increased) (60). During embryogenesis, these myogenic regulatory factors stimulate myoblast determination and differentiation (64). Unfortunately, the role of these myogenic factors in adult muscle is not fully established and, therefore, the physiologic importance of CMV-induced changes in these myogenic factors remains unclear. Nonetheless, diaphragmatic contractile function is depressed in MyoD knockout mice and, therefore, the diminished MyoD expression may play a role in CMV-induced diaphragmatic contractile dysfunction (65).
Prolonged CMV is associated with numerous important structural and functional changes in the diaphragm (Fig. 6). First, CMV results in a rapid onset of diaphragmatic atrophy in both animals and humans. CMV is also associated with a time-dependent induction of diaphragmatic contractile dysfunction, resulting in depressed diaphragmatic-specific force production at both submaximal and maximal stimulation frequencies. Hence, it is feasible that CMV-induced diaphragmatic weakness may play an important role in weaning difficulties.
CMV-induced diaphragmatic atrophy in animal models of MV occurs due to both a reduction in protein synthesis and an increase in diaphragmatic protein degradation. MV-induced diaphragmatic atrophy is also associated with ultrastructural changes in the diaphragm including diverse areas of abnormal sarcomere structure and irregular Z-line structure.
Furthermore, CMV results in significant changes in diaphragmatic gene expression as >350 gene products are either increased or decreased during 6 hrs to 18 hrs of CMV. In general, stress responsive genes are up-regulated whereas genes in the structural protein and energy metabolism categories are down-regulated. This alteration in diaphragmatic gene expression contributes to the extensive remodeling that occurs in the diaphragm during prolonged CMV.
Importantly, prolonged CMV results in oxidative damage to the diaphragm as demonstrated by increased protein oxidation and lipid peroxidation. This is significant because redox disturbances in skeletal muscle promote contractile dysfunction and the activation of proteolytic systems. Prevention of CMV-induced oxidative damage in the diaphragm has been demonstrated to retard MV-induced diaphragmatic atrophy and contractile dysfunction in animals.
Although much research progress has been made toward describing the impact of prolonged CMV on diaphragmatic structure and function, numerous unanswered questions remain. An important clinical question that remains unresolved is: Do pressure-assist modes of MV protect the diaphragm against the rapid onset of diaphragmatic atrophy and contractile dysfunction that is associated with CMV? A related question is: Can periodic activation of the diaphragm (i.e., electrical stimulation) protect the diaphragm against CMV-induced diaphragmatic wasting?
Other important mechanistic questions remain unanswered. For example, the role that autophagy plays in CMV-induced diaphragmatic atrophy remains unclear. Furthermore, limited information exists about which diaphragmatic proteases are essential and/or rate-limiting in MV-induced proteolysis in the diaphragm. Also, it is unclear if major proteolytic systems are independently regulated or coordinately controlled by a unifying regulatory system.
Finally, the principal ROS producing pathways in the diaphragm that are active during prolonged CMV remain to be identified. This is important because CMV-induced diaphragmatic ROS production is a required upstream signal that promotes diaphragmatic atrophy and contractile dysfunction. Furthermore, the specific signaling pathways that link diaphragmatic ROS production to depressed protein synthesis and/or increased proteolysis in the diaphragm remain unresolved. This is a critical area for future research because understanding the cellular signaling pathways that are responsible for MV-induced diaphragmatic atrophy and contractile dysfunction is a prerequisite to develop methods to protect patients against MV-mediated diaphragmatic weakness.
The research reported in the article was supported, in part, by Grants R01HL062361 and R01HL072789 from the National Institutes of Health (SKP).
The authors have not disclosed any potential conflicts of interest.