Mechanical Ventilation–induced Diaphragmatic Atrophy Is Associated with Oxidative Injury and Increased Proteolytic Activity

Abstract

Prolonged mechanical ventilation (MV) results in reduced diaphrag-matic maximal force production and diaphragmatic atrophy. To investigate the mechanisms responsible for MV-induced diaphrag-matic atrophy, we tested the hypothesis that controlled MV results in oxidation of diaphragmatic proteins and increased diaphrag-matic proteolysis due to elevated protease activity. Further, we postulated that MV would result in atrophy of all diaphragmatic muscle fiber types. Mechanically ventilated animals were anesthe-tized, tracheostomized, and ventilated with 21% O2 for 18 hours. MV resulted in a decrease (p � 0.05) in diaphragmatic myofibrillar protein and the cross-sectional area of all muscle fiber types (i.e., I, IIa, IId/x, and IIb). Further, MV promoted an increase (p � 0.05) in diaphragmatic protein degradation along with elevated (p � 0.05) calpain and 20S proteasome activity. Finally, MV was also associated with a rise (p � 0.05) in both protein oxidation and lipid peroxidation. These data support the hypothesis that MV is associated with atrophy of all diaphragmatic fiber types, increased diaphragmatic protease activity, and augmented diaphragmatic ox-idative stress.

Caspase-3 Regulation of Diaphragm Myonuclear Domain during Mechanical Ventilation–induced Atrophy

“…Heart rate (365 Ϯ 9 beats/min) and mean blood pressure (90 Ϯ 4 mm Hg) homeostasis were maintained during MV across all treatment groups. Blood gas homeostasis and pH (data not shown) were maintained within physiologic levels during the experiment as reported previously (1,4,6,18). Because sepsis is associated with diaphragmatic contractile dysfunction, strict aseptic techniques were followed throughout the experiments (1,4,6,18).…”

“…This notion is supported by the observations that new nuclei are incorporated into locomotor skeletal muscle fibers during growth and that nuclei are lost during atrophy (12,14,16). Although it is known that controlled MV decreases protein synthesis and induces proteolysis and myofiber atrophy across all fiber types (6,17,18), it is unknown whether the myonuclear domain of diaphragm myofibers is altered during the fiber atrophy associated with MV.Regulation of myonuclear loss in skeletal muscle could occur through extrinsic death receptor and intrinsic (sarcoplasmic reticulum and mitochondrial) mediated pathways (19). Caspase (cysteine-dependent, aspartate-directed protease) activation results in protein cleavage and functions in intrinsic and extrinsic pathways of apoptosis.…”

“…Although it is clear that MV-induced diaphragm inactivity results in fiber atrophy (3,5,6), it is unknown if prolonged MV is associated with alterations in diaphragmatic myonuclei via apoptotic mechanisms. Based on reports investigating locomotor skeletal muscle atrophy (12,(14)(15)(16), we hypothesized that diaphragmatic inactivity during prolonged MV would result in myofiber atrophy and myonuclear loss, resulting in the maintenance of a constant myonuclear domain.…”

Corticosteroid effects on ventilator-induced diaphragm dysfunction in anesthetized rats depend on the dose administered

“…Animal studies have clearly demonstrated that CMV impacts the diaphragm by promoting contractile dysfunction, increased proteolysis and atrophy [17,36]. Interestingly, our results reveal that administration of a relatively low dose of methylprednisolone (5 mg/kg, corresponding ~0.8 mg/kg in humans [19]) exacerbates the CMV-induced diaphragm dysfunction, whereas a higher dose (30 mg/kg, corresponding ~5 mg/kg in humans [19]) completely protected the diaphragm against VIDD.…”

“…This study and a previous CMV study, in which we used a calpain inhibitor [9], confirm the important role of the calpain system in the development of VIDD. It is known that three major proteolytic systems are upregulated in the diaphragm during mechanical ventilation: the ubiquitin proteasome system (UPP), the Ca 2+ -dependent calpain system and the lysosomal system [15-17]. Although the UPP is considered a major proteolytic system in skeletal muscle, it cannot degrade intact myofilaments.…”