Study On The Mechanism Of Skeletal Muscle Hyperproteolysis During Burn Sepsis

Sepsis in severely burned patients is associated with high morbidity and mortality rates. It is well known that hypermetabolism invariably accompanies sepsis, and the process has been labeled as self-cannibalistic. This intensely accelerates the breakdown of body proteins, malnutrition will devalop rapidly, impairing visceral organs. Hyperproteolysis of skeletal muscle holding 50 percent dry weight of body is the main cause for negative nitrogen balance. However, the mechanism of hyperproteolysis in skeletal muscle during burn sepsis is still obscure.OBJECTIVEPrevious studies had found that proteolytic rate of skeletal muscle was positively related to the plasma concentrations of glucocorticoid, interleukin (IL) -6 and tumor necrosis factor (TNF)- a during burn sepsis. In addition, there are many protein degradation pathways in cells, however, which is the key pathway for proteolysis of skeletal muscle during burn sepsis ? Ubiquitin system, a new protein degradation pathway, is regarded as the important one for proteolysis of skeletal muscle. Our study also found that the gene and protein expression of ubiquitin system in skeletal muscle were enhanced during burn sepsis.On this background, the purpose of this study was to make further investigation for the effects of glucocorticoid and inflammatory mediators and ubiquitin system on hyperproteolysis in skeletal muscle during burn sepsis.METHODS:1. Animal Experiments:(1)The normal rats were injected with various dosage of glucocorticoid or TNF- a and the proteolytic rates of extensor digitorium longus (EDL) were detected by the in vitro muscle incubation system with sufficient oxygen supply and high performance liquid chromatography. The gene and protein expressions of ubiquitin systemin rats' EDL muscles were determined by Northern blot analysis and immunopathology.(2)We incubated the rats' EDL with medium containing TNF-a and analyzed the gene and protein expressions of ubiquitin system by same methods.(3) We established the burn sepsis rat model and treated them with glucocorticoid antagonist RU38486, and analyzed the proteolytic rate, the gene and protein expressions of ubiquitin system in EDL. The plasma concentrations of cortisol and inflammatory mediums were detected by radio-immunity.2. Cell Culture Experiments:(1) To establish stable methods for culturing skeletal muscle cells.(2) To label the myotube protein with L-[3,5-3H]-tyrosine.(3) We cultured the myotube with medium containing various concentrations of dexamethasone or TNF- a combined with proteosome inhibitor MG132, and analyzed the proteolytic rate and gene expressions of ubiquitin system in myotube.3. Clinical parts:We collected skeletal muscle (Biceps brachii muscle; Quadriceps femoris muscle), blood and urine samples from nine patients with severe burn injury and nine patients waiting for plastic operation. The plasma concentrations of cortisol and TNF- a and the amounts of 3-methylhistidine in urine were detected. The gene and protein expressions in muscle samples were analyzed by the same methods as mentioned above.RESULTS and CONCLUSIONS:1. The relationship between dexamethasone and proteolysis of skeletal muscle(1) Dexamethasone could enhance the gene and proein expressions of ubiquitin system and the degradation rate of total protein, especially that of the myofibrillar protein, in skeletal muscle of rats with dose-dependent-relationship, and these effects could be inhibited by glucocorticoid receptor antagonist RU38486.(2) Glucocorticoid receptor antagonist RUB 8486 could significantly inhibit the gene and proein expressions of ubiquitin system and proteolytic rate in skeletal muscle of rats with burn sepsis.(3) Dexamethasone could upgrade the gene expressions of ubiquitin system and proteolytic rate in myotubes with dose-dependent-relationship, and the enhanced proteolytic rate could be blocked by proteasome inhibitor MG132.2. The relationship between...