Sepsis stimulates calpain activity in skeletal muscle by decreasing calpastatin activity but does not activate caspase-3

Abstract

, and Per-Olof Hasselgren. Sepsis stimulates calpain activity in skeletal muscle by decreasing calpastatin activity but does not activate caspase-3. Am J Physiol Regul Integr Comp Physiol 288: R580 -R590, 2005. First published November 24, 2004 doi:10.1152/ajpregu.00341.2004.-We examined the influence of sepsis on the expression and activity of the calpain and caspase systems in skeletal muscle. Sepsis was induced in rats by cecal ligation and puncture (CLP). Control rats were sham operated. Calpain activity was determined by measuring the calcium-dependent hydrolysis of casein and by casein zymography. The activity of the endogenous calpain inhibitor calpastatin was measured by determining the inhibitory effect on calpain activity in muscle extracts. Protein levels of -and m-calpain and calpastatin were determined by Western blotting, and calpastatin mRNA was measured by real-time PCR. Caspase-3 activity was determined by measuring the hydrolysis of the fluorogenic caspase-3 substrate Ac-DEVD-AMC and by determining protein and mRNA expression for caspase-3 by Western blotting and real-time PCR, respectively. In addition, the role of calpains and caspase-3 in sepsis-induced muscle protein breakdown was determined by measuring protein breakdown rates in the presence of specific inhibitors. Sepsis resulted in increased muscle calpain activity caused by reduced calpastatin activity. In contrast, caspase-3 activity, mRNA levels, and activated caspase-3 29-kDa fragment were not altered in muscle from septic rats. Sepsis-induced muscle proteolysis was blocked by the calpain inhibitor calpeptin but was not influenced by the caspase-3 inhibitor Ac-DEVD-CHO. The results suggest that sepsis-induced muscle wasting is associated with increased calpain activity, secondary to reduced calpastatin activity, and that caspase-3 activity is not involved in the catabolic response to sepsis.

Calpain inhibition attenuates right ventricular contractile dysfunction after acute pressure overload

“…Sorimachi et al and Enns et al both found that ischemia/reperfusion results in a decline in calpastatin inhibitory activity with no change in fluorometrically determined calpain activity [39,40], but that calpain inhibition improves post-ischemic contractile function. Wei et al found altered calpain activity due to a change in calpastatin inhibitory activity in a model of sepsis-mediated muscle proteolysis [41]. Shi et al found down regulation of the 41 kDa isoform of calpastatin in kidney following renal ischemia/reperfusion similar to what we observed in the RV [42].…”

Development of a Diagnostic Method for Detecting Increased Muscle Protein Degradation in Patients with Catabolic Conditions

“…However, measuring caspase-3 activity in muscle samples is not useful as a diagnostic method because we and others have found that caspase-3 activity in muscle samples is low and the "signal to noise" ratio is high even in muscles of rodents with demonstrably high rates of protein degradation. Wei et al (21) reported difficulty in detecting changes in caspase-3 activity in muscle of septic rats. Sepsis activates the UPP in muscle, so their results might be related to degradation of caspase-3 by the ATP-dependent UPP (22).…”

Protein Degradation by the Ubiquitin–Proteasome Pathway in Normal and Disease States

“…Calpains are calcium-dependent cysteine proteases that have been suggested to play roles in several disorders, especially muscular dystrophy. Evidence for calpain's activation has been reported in muscles of animals with sepsis (95). Although inhibition of the calcium-activated proteases in muscles from rodents with uremia or several other types of atrophy has not been found to block the increase in protein degradation or the degradation of myofibrillar proteins or the accumulation of the 14-kD actin fragment in muscle cells (92), the transgenic expression of calpastatin, an endogenous calpain inhibitor, can reduce denervation atrophy and progression of muscular dystrophy in mice (96).…”