Dexamethasone stimulates store-operated calcium entry and protein degradation in cultured L6 myotubes through a phospholipase A₂-dependent mechanism

Abstract

Hasselgren PO. Dexamethasone stimulates store-operated calcium entry and protein degradation in cultured L6 myotubes through a phospholipase A 2-dependent mechanism. Am J Physiol Cell Physiol 298: C1127-C1139, 2010. First published January 27, 2010; doi:10.1152/ajpcell.00309.2009.-Muscle wasting in various catabolic conditions is at least in part regulated by glucocorticoids. Increased calcium levels have been reported in atrophying muscle. Mechanisms regulating calcium homeostasis in muscle wasting, in particular the role of glucocorticoids, are poorly understood. Here we tested the hypothesis that glucocorticoids increase intracellular calcium concentrations in skeletal muscle and stimulate storeoperated calcium entry (SOCE) and that these effects of glucocorticoids may at least in part be responsible for glucocorticoid-induced protein degradation. Treatment of cultured myotubes with dexamethasone, a frequently used in vitro model of muscle wasting, resulted in increased intracellular calcium concentrations determined by fura-2 AM fluorescence measurements. When SOCE was measured by using calcium "add-back" to muscle cells after depletion of intracellular calcium stores, results showed that SOCE was increased 15-25% by dexamethasone and that this response to dexamethasone was inhibited by the store-operated calcium channel blocker BTP2. Dexamethasone treatment stimulated the activity of calcium-independent phospholipase A 2 (iPLA2), and dexamethasone-induced increase in SOCE was reduced by the iPLA 2 inhibitor bromoenol lactone (BEL). In additional experiments, treatment of myotubes with the store-operated calcium channel inhibitor gadolinium ion or BEL reduced dexamethasone-induced increase in protein degradation. Taken together, the results suggest that glucocorticoids increase calcium concentrations in myocytes and stimulate iPLA 2-dependent SOCE and that glucocorticoid-induced muscle protein degradation may at least in part be regulated by increased iPLA 2 activity, SOCE, and cellular calcium levels. glucocorticoids; muscle; wasting MUSCLE WASTING, mainly reflecting stimulated degradation of myofibrillar proteins, is commonly seen in patients with sepsis, severe injury, and cancer (24, 52). Loss of muscle mass also occurs in patients with muscular dystrophy (30, 31). In previous studies, we found evidence (4, 15, 79) that sepsis-induced muscle proteolysis was associated with increased uptake and tissue levels of calcium in skeletal muscle. Additional reports suggest that muscle atrophy caused by other catabolic conditions, including denervation, burn injury, and muscular dystrophy, is also associated with increased tissue concentrations of calcium (2,3,10,17,64,73). These observations are significant because calcium is an important regulator of muscle protein balance and may link systemic catabolic responses to stimulated protein breakdown in skeletal muscle (16,34,50). The mechanisms regulating calcium entry and cellular levels of calcium in muscle-wasting conditions are not known.Glucocorticoids are prom...

Proinflammatory treatment of astrocytes with lipopolysaccharide results in augmented Ca2+ signaling through increased expression of VIA phospholipase A2 (iPLA2)

“…The critical role of VIA iPLA 2 for CCE was extensively investigated before (9,21,37). We have shown that the mechanism that underlies the increased load of the Ca 2ϩ stores and duration of the responses to ATP in LPStreated astrocytes is based on the increased CCE.…”

Corticosteroids and muscle wasting: role of transcription factors, nuclear cofactors, and hyperacetylation

“…These observations are significant because calcium is an important regulator of muscle protein breakdown [40]. In a recent study, we found evidence that glucocorticoids may increase muscle calcium levels secondary to stimulated store-operated calcium entry (SOCE) and that this effect of glucocorticoids was regulated by increased phospholipase A2 activity [41]. Additional experiments in the same study suggested that dexamethasone-induced protein degradation was calcium- and SOCE-dependent, suggesting that inhibition of SOCE may be a therapeutic tool to prevent glucocorticoid-induced muscle wasting.…”

Loss of muscle strength during sepsis is in part regulated by glucocorticoids and is associated with reduced muscle fiber stiffness

“…In previous studies, the increase in muscle proteolysis and changes in the expression of muscle wasting-associated transcription factors and nuclear cofactors during sepsis were more extensively prevented by RU38486 (1,36,45,53,57), supporting the concept that reduced muscle strength during early sepsis does not primarily reflect loss of muscle mass. Of note, we recently found that muscle calcium homeostasis, in particular store-operated calcium entry, is regulated by glucocorticoids (21), and it is possible that the effect of RU38486 observed in the present study reflected improved calcium homeostasis, although additional experiments are needed to test that hypothesis.…”