Biomedical subjects

P O Hasselgren

Publications and source records attributed to P O Hasselgren.

At least 19 recordsLinked to original sources

PubMed1998

Sepsis: stimulation of energy-dependent protein breakdown resulting in protein loss in skeletal muscle.

Muscle catabolism is a characteristic metabolic response to sepsis, severe infection, and injury. In patients with severe and protracted sepsis, the catabolic response results in muscle wasting and fatigue, which may adversely affect the outcome in these patients. An understanding of the regulation of muscle protein breakdown during sepsis and the mechanisms involved is important from a clinical standpoint and is essential for the development of new therapeutic modalities to prevent protein loss from muscle tissue. Studies in septic patients and experimental animals have provided evidence that the myofibrillar proteins actin and myosin are particularly sensitive to the effects of sepsis. Among the factors that regulate muscle protein breakdown during sepsis, the proinflammatory cytokines tumor necrosis factor and interleukin-1, together with glucocorticoids, are the principal mediators. Intracellular protein breakdown is regulated by multiple proteolytic pathways. Among these, the energy-ubiquitin-dependent pathway accounts for a major portion of muscle protein breakdown during sepsis. The development of specific proteasome inhibitors may make it possible in the future to target the molecular mechanisms of sepsis-induced increase in muscle proteolysis. Such treatment may prove an important avenue to reduce the metabolic cost in patients with severe infection or sepsis.

Energy Metabolism↗

PubMed1998

Sepsis-induced increase in muscle proteolysis is blocked by specific proteasome inhibitors.

Recent studies suggest that sepsis stimulates ubiquitin-dependent protein breakdown in skeletal muscle. The 20S proteasome is the catalytic core of the ubiquitin-dependent proteolytic pathway. We tested the effects in vitro of the proteasome inhibitors N-acetyl-L-leucinyl-L-leucinal-L-norleucinal (LLnL) and lactacystin on protein breakdown in incubated muscles from septic rats. LLnL resulted in a dose- and time-dependent inhibition of protein breakdown in muscles from septic rats. Lactacystin blocked both total and myofibrillar muscle protein breakdown. In addition to inhibiting protein breakdown, LLnL reduced muscle protein synthesis and increased ubiquitin mRNA levels, probably reflecting inhibited proteasome-associated ribonuclease activity. Inhibited muscle protein breakdown caused by LLnL or lactacystin supports the concept that the ubiquitin-proteasome pathway plays a central role in sepsis-induced muscle proteolysis. The results suggest that muscle catabolism during sepsis may be inhibited by targeting specific molecular mechanisms of muscle proteolysis.

Acetylcysteine↗

PubMed1998

IGF-I stimulates protein synthesis but does not inhibit protein breakdown in muscle from septic rats.

Sepsis is associated with reduced protein synthesis and increased protein degradation in skeletal muscle. We examined the effects of insulin-like growth factor I (IGF-I) on protein synthesis and breakdown in muscles from nonseptic and septic rats. Sepsis was induced by cecal ligation and puncture; control rats were sham operated. Extensor digitorum longus muscles were incubated in the absence or presence of IGF-I at concentrations ranging from 100 ng/ml to 10 micrograms/ml. Total and myofibrillar protein breakdown rates were measured as net release of tyrosine and 3-methylhistidine, respectively. Protein synthesis was determined by measuring incorporation of [U-14C]phenylalanine into protein. IGF-I stimulated protein synthesis in a dose-dependent fashion in muscles from both sham-operated and septic rats, with a maximal effect seen at a hormone concentration between 500 and 1,000 ng/ml. IGF-I inhibited total and myofibrillar protein breakdown in muscles from sham-operated rats, whereas in muscles from septic rats, IGF-I had no effect on protein breakdown, even at high concentrations. The results suggest that protein breakdown in skeletal muscle becomes resistant to IGF-I during sepsis and that this resistance reflects a postreceptor defect.

Animals↗

PubMed1998

Proteasome blockers inhibit protein breakdown in skeletal muscle after burn injury in rats.

1. Burn injury stimulates ubiquitin-dependent protein breakdown in skeletal muscle. The 20S proteasome is the proteolytic core of the 26S proteasome that degrades ubiquitin conjugates. We examined the effects of the proteasome inhibitors N-acetyl-L-leucinyl-L-leucinal-L-norleucinal (LLnL), lactacystin and beta-lactone on protein breakdown in muscles from burned rats. 2. A full-thickness burn of 30% total body surface area was inflicted on the back of rats. Control rats underwent a sham procedure. After 24 h, extensor digitorum longus muscles were incubated in the absence or presence of 20S proteasome blocker and protein turnover rates and ubiquitin mRNA levels were determined. 3. LLnL resulted in a dose- and time-dependent inhibition of total protein breakdown in incubated muscles from burned rats. Lactacystin and beta-lactone blocked both total and myofibrillar muscle protein breakdown. In addition to inhibiting protein breakdown, LLnL increased ubiquitin mRNA levels, possibly reflecting inhibited proteasome-associated RNase activity. 4. Inhibited muscle protein breakdown caused by LLnL, lactacystin and beta-lactone supports the concept that the ubiquitin-proteasome pathway plays a central role in burn-induced muscle proteolysis. Because the proteasome has multiple important functions in the cell, in addition to regulating general protein breakdown, further studies are needed to test the role of proteasome blockers in the treatment or prevention of muscle catabolism.

Acetylcysteine↗

PubMed1997

Sepsis is associated with increased mRNAs of the ubiquitin-proteasome proteolytic pathway in human skeletal muscle.

Previous studies provided evidence that sepsis-induced muscle proteolysis in experimental animals is caused by increased ubiquitin-proteasome-dependent protein breakdown. It is not known if a similar mechanism accounts for muscle proteolysis in patients with sepsis. We determined mRNA levels for ubiquitin and the 20 S proteasome subunit HC3 by Northern blot analysis in muscle tissue from septic (n = 7) and non-septic (n = 11) patients. Plasma and muscle amino acid concentrations and concentrations in urine of 3-methylhistidine (3-MH), creatinine, and cortisol were measured at the time of surgery to assess the catabolic state of the patients. A three- to fourfold increase in mRNA levels for ubiquitin and HC3 was noted in muscle tissue from the septic patients concomitant with increased muscle levels of phenylalanine and 3-MH and reduced levels of glutamine. Total plasma amino acids were decreased by approximately 30% in the septic patients. The 3-MH/creatinine ratio in urine was almost doubled in septic patients. The cortisol levels in urine were higher in septic than in control patients but this difference did not reach statistical significance. The results suggest that sepsis is associated with increased mRNAs of the ubiquitin-proteasome pathway in human skeletal muscle.

Aged↗

PubMed1997

Treatment of endotoxemic mice with anti-interleukin-6 antibody paradoxically increases interleukin-6 levels and stimulates mucosal protein synthesis.

OBJECTIVE: To test the role of interleukin-6 (IL-6) in the regulation of mucosal protein synthesis during endotoxemia. EXPERIMENTAL DESIGN: Three series of experiments were performed: (1) mice were injected with different doses of lipopolysaccharide (LPS) and protein synthesis was measured in jejunal mucosa at various time points up to 24 hours after injection of LPS; (2) mice were pretreated with anti-IL-6 antibody before injection of LPS; and (3) mice were injected with recombinant IL-6 and mucosal protein synthesis was measured 24 hours later. RESULTS: Treatment of mice with LPS resulted in a dose- and time-dependent increase in mucosal protein synthesis. Pretreatment of endotoxemic mice with anti-IL-6 antibody resulted in a paradoxical increase in plasma and tissue levels of IL-6 and a further increase in mucosal protein synthesis. Administration of IL-6 to normal mice resulted in increased mucosal protein synthesis. CONCLUSION: The increase in mucosal protein synthesis seen during endotoxemia may be regulated at least in part by IL-6.

Animals↗

PubMed1997

Complement C3 production in human intestinal epithelial cells is regulated by interleukin 1beta and tumor necrosis factor alpha.

BACKGROUND: Sepsis and endotoxemia are associated with increased mucosal production of complement component C3; the enterocyte may be a source of C3 in these conditions. OBJECTIVE: To test the hypothesis that interleukin 1beta (IL-1beta) and tumor necrosis factor alpha (TNF-alpha) regulate the production of C3 in the enterocyte at the transcriptional level and that this regulation is potentiated by interferon gamma (IFN-gamma). METHODS: Cultured Caco-2 cells, a human intestinal epithelial cell line, were treated with various concentrations of human recombinant IL-1beta (0.005-1.25 ng/mL) or TNF-alpha (1-1000 U/mL) with or without the addition of IFN-gamma (250 U/mL). C3 levels in the culture medium were measured by enzyme-linked immunosorbent assay and cellular messenger RNA levels by Northern blot analysis. RESULTS: Treatment of the Caco-2 cells with IL-1beta or TNF-alpha resulted in a time- and dose-dependent increase in C3 production. The use of IFN-gamma alone did not affect C3 production but potentiated the effect of IL-1beta and TNF-alpha in a synergistic manner. C3 messenger RNA levels were increased following stimulation with either cytokine. CONCLUSIONS: C3 production in the enterocyte is regulated by IL-1beta and TNF-alpha at the transcriptional level, and this response is potentiated by IFN-gamma. The results suggest that C3 production in the intestinal mucosa may be regulated locally by cytokines in a paracrine or autocrine manner.

Caco-2 Cells↗

PubMed1997

IL-6 production in human intestinal epithelial cells following stimulation with IL-1 beta is associated with activation of the transcription factor NF-kappa B.

Recent studies suggest that interleukin-1 beta (IL-1 beta) stimulates interleukin-6 (IL-6) production in human intestinal epithelial cells, but the intracellular mechanisms of this response are not known. In other reports, the nuclear factor-kappa B (NF-kappa B) regulated IL-6 production in certain cell types. We tested the hypothesis that IL-6 production in the enterocyte is associated with activation of NF-kappa B. Caco-2 cells, a human intestinal epithelial cell line, were grown in tissue culture whereafter they were treated with IL-1 beta (0.5 ng/ml). Cells were preincubated with pyrrolidine dithiocarbamate (PDTC; 10-500 microM), tosyl-lys-chloromethylketone (TLCK; 10-500 microM), or genistein (25-75 microM), all of which are known inhibitors of NF-kappa B. IL-6 levels in the culture media were measured after 24 hr by enzyme-linked immunosorbent assay (ELISA) and IL-6 messenger RNA (mRNA) levels were determined after 4 hr by competitive reverse-transcriptase polymerase chain reaction (RT-PCR). NF-kappa B activity was determined by electrophoretic gel mobility shift assay (EMSA). PDTC, TLCK, and genistein each inhibited IL-1 beta-induced IL-6 production by the Caco-2 cells in a dose-dependent fashion. These responses were also associated with a decrease in IL-6 mRNA levels. There was no NF-kappa B activity in untreated cells, but the addition of IL-1 beta resulted in the activation of NF-kappa B as determined by EMSA. The results suggest that IL-1 beta-induced IL-6 production in the enterocyte is associated with activation of NF-kappa B. The inhibition of IL-6 production by the NF-kappa B inhibitors indicates that the IL-6 production is regulated by NF-kappa B, although further experiments are needed to test that hypothesis.

Caco-2 Cells↗

PubMed1997

Increased glutamine consumption in small intestine epithelial cells during sepsis in rats.

BACKGROUND: Previous reports have emphasized increased intestinal glutamine consumption during sepsis. This may be of clinical significance as glutamine is a (and perhaps the) physiologic fuel for the intestine. A problem remains, however, as glutaminase, the major enzyme for the degradation of glutamine, is decreased rather than increased in sepsis. This implies an alteration in the metabolic fate of glutamine in sepsis, which may be of clinical significance. METHODS: We determined the effect of sepsis on glutamine metabolism in mucosa of small intestine. Sepsis was induced in rats by cecal ligation and puncture. Control rats were sham-operated. After 16 hours, glutamine consumption was measured in isolated enterocytes, and glutaminase, glutamine synthetase, and glutamine transaminase activities and DNA synthesis were determined in mucosa. RESULTS: Glutamine consumption was increased during sepsis in enterocytes from the tips of the villi and was unchanged in enterocytes from the midportions of the villi and the crypts. As previously shown, mucosal glutaminase activity was reduced. However, glutamine synthetase and glutamine transaminase activities were stimulated in septic rats, suggesting an increase in metabolism of glutamine via alternate pathways. DNA synthesis was increased during sepsis, especially in crypt cells. CONCLUSIONS: Increased or unchanged glutamine consumption in enterocytes from septic rats, despite reduced glutaminase activity, appears to reflect increased activity of other enzyme systems and/or increased utilization of the amino acid for DNA and protein synthesis. Since other aspects of glutamine metabolism may also be deranged in sepsis, a continual enteral supply of glutamine to the intestinal lumen during sepsis may be clinically useful.

Animals↗

PubMed1997

Sepsis and endotoxaemia in mice stimulate the expression of interleukin-I and interleukin-6 in the central nervous system.

1. In previous studies, experimental endotoxaemia was found to stimulate cytokine production in the central nervous system. The effect of sepsis on brain cytokines is not fully known. We compared the effect of endotoxaemia and sepsis on brain interleukin-1 and interleukin-6 expression. 2. Male A/J mice were injected subcutaneously with lipopolysaccharide (10 mg/kg) or an equal volume of saline as control. Sepsis was induced by caecal ligation and puncture (CLP); control mice underwent sham-operation. Brain tissue was assayed for interleukin-1 and interleukin-6 by ELISA. Northern blotting or the polymerase chain reaction was used to determine cytokine mRNA levels. 3. Administration of endotoxin induced a greater than fourfold increase in brain interleukin-1, a greater than threefold increase in interleukin-6 and an increase in mRNA for both cytokines. Caecal ligation and puncture resulted in increased brain interleukin-1 and interleukin-6 levels, but the changes were less pronounced and occurred later than after injection of endotoxin. There was no detectable difference in brain interleukin-1 mRNA between septic and sham-operated mice, whereas interleukin-6 mRNA was increased in brains of septic animals. 4. Sepsis and endotoxaemia resulted in similar, although not identical, changes in brain interleukin-1 and interleukin-6 concentrations and mRNA levels, suggesting that increased cytokine production in the central nervous system is part of the systemic response to sepsis and may be mediated by endotoxin.

Animals↗

PubMed1997

The ubiquitin-proteasome pathway: review of a novel intracellular mechanism of muscle protein breakdown during sepsis and other catabolic conditions.

SUMMARY BACKGROUND DATA: Patients with sepsis and other catabolic conditions, such as severe trauma, cancer, and fasting, suffer significant loss of body protein, the majority of which originates from skeletal muscle. Recent evidence suggests that muscle protein breakdown during sepsis is caused by upregulated activity in the ubiquitin-proteasome pathway and is associated with increased expression of the ubiquitin gene. PURPOSE: The purpose of the study was to review the role of the ubiquitin-proteasome pathway in the regulation of muscle proteolysis during sepsis and other catabolic conditions. REVIEW: Proteins that are degraded by the ubiquitin-proteasome mechanism are first conjugated to ubiquitin, a 76-amino-acid, highly conserved residue. Ubiquitinated proteins are recognized by the 26S proteasome, which is a large proteolytic complex consisting of the 19S cap complex and the 20S proteasome. The 20S proteasome is a cylindrical particle composed of four stacked rings, making it look like a barrel. The rings form a "tunnel" in which the target proteins are hydrolyzed, after which ubiquitin is released to be reused in the proteolytic pathway. A unique feature of the ubiquitin-proteasome proteolytic pathway is its energy dependency. CONCLUSIONS: An understanding of the molecular regulation of protein metabolism in patients with sepsis and other catabolic conditions is important because it may form the basis for improved treatment in the future.

Cysteine Endopeptidases↗

PubMed1997

Interleukin-1 beta and interferon-gamma regulate interleukin-6 production in cultured human intestinal epithelial cells.

Recent studies suggest that interleukin-6 (IL-6) is produced in the intestinal mucosa during sepsis and endotoxemia and that the enterocyte may be a source of IL-6 in these conditions. The regulation of IL-6 production in the enterocyte is not fully understood. We tested the hypothesis that IL-6 production in the enterocyte is regulated by proinflammatory cytokines. This was done by treating cultured Caco-2 cells, a transformed human intestinal epithelial cell line, with different concentrations of tumor necrosis factor-alpha (TNF-alpha), IL-1 beta, IL-6 or interferon-gamma (IFN-gamma). IL-6 production by the Caco-2 cells was determined by ELISA. The expression of IL-6 mRNA was determined by reverse-transcriptase polymerase chain reaction. IL-6 was not produced in unstimulated Caco-2 cells. Treatment of the Caco-2 cells with IL-1 beta resulted in a dose- and time-dependent stimulation of IL-6 production with a maximal effect noted at an IL-1 beta concentration of .5 ng/mL at 24 h. IFN-gamma alone did not stimulate IL-6 production but potentiated the effect of IL-1 beta in a synergistic fashion. Treatment of the Caco-2 cells with IL-1 beta induced expression of IL-6 mRNA with a response noticed after 30 min. TNF-alpha and IL-6 did not influence the production of IL-6 in the Caco-2 cells. The results suggest that enterocyte IL-6 production is stimulated by IL-1 beta and that this effect is potentiated by IFN-gamma. The regulation of IL-6 production in the enterocyte may be specific for IL-1 beta, since neither TNF nor IL-6 stimulated IL-6 production.

Adjuvants, Immunologic↗

PubMed1997

Cytokines block the effects of insulin-like growth factor-I (IGF-I) on glucose uptake and lactate production in skeletal muscle but do not influence IGF-I-induced changes in protein turnover.

There is evidence that proinflammatory cytokines are involved in the regulation of muscle protein breakdown in various catabolic conditions but the mechanisms are not fully understood. Previous studies suggest that cytokines reduce circulating and tissue levels of insulin-like growth factor-I (IGF-I) and may block the anabolic effects of the hormone in certain cell types and tissues. We tested the hypothesis that a mixture of tumor necrosis factor alpha, interleukin-1 alpha, and interferon-gamma block the anabolic effects of IGF-I in skeletal muscle. Muscles from burned or unburned rats were incubated in the absence or presence of 1 microgram/mL of IGF-I with or without the addition of the cytokines. As expected, IGF-I stimulated protein synthesis and inhibited protein breakdown in incubated muscles. The cytokines did not influence protein turnover rates in muscles incubated with or without IGF-I. In additional experiments, the effects of IGF-I on glucose uptake and lactate production were tested. IGF-I increased glucose uptake approximately 2.5-fold and stimulated lactate production approximately 5-fold. These effects of the hormone were significantly inhibited by the cytokine mixture. The results suggest that cytokines do not induce protein catabolism by directly inhibiting the anabolic effects of IGF-I in muscle tissue. The inhibitory effects of the cytokines on IGF-I-stimulated glucose transport and lactate production suggest that the lack of effect of cytokines on protein metabolism was not due to a metabolic unresponsiveness of the incubated muscles to the cytokines.

Animals↗

PubMed1997

Intracellular regulation of protein degradation during sepsis is different in fast- and slow-twitch muscle.

We tested the hypothesis that the difference in the response to sepsis of protein breakdown between fast- and slow-twitch skeletal muscle reflects differential activation of the energy-ubiquitin-dependent proteolytic pathway. In addition, we defined the time course and the tissue specificity of sepsis-induced changes in the expression of the ubiquitin pathway. Sepsis was induced in rats by cecal ligation and puncture; control rats were sham operated. Energy-dependent protein breakdown was measured in incubated extensor digitorum longus (EDL) and soleus muscles. Ubiquitin mRNA levels were determined by Northern blot analysis. Sepsis resulted in increased energy-dependent protein breakdown and upregulated expression of ubiquitin mRNA in the fast-twitch EDL but not in the slow-twitch soleus muscle. The sepsis-induced increase in ubiquitin mRNA levels in the EDL muscle was noticeable before the increase in energy-dependent protein breakdown. Sepsis increased ubiquitin mRNA levels in the diaphragm (a mixed fiber-type muscle) but not in heart, liver, kidney, or intestine, consistent with a tissue-specific regulation of the ubiquitin system during sepsis. The results suggest that the difference in protein breakdown during sepsis between fast- and slow-twitch muscles reflects differential activation of the energy-ubiquitin-dependent proteolytic pathway. The data also suggest that the expression of the ubiquitin pathway is upregulated in a time-dependent fashion during sepsis and that this response is not a generalized phenomenon but is tissue specific.

Animals↗

PubMed1997

The gut as a source of inflammatory cytokines after stimulation with endotoxin.

OBJECTIVE: To find out if endotoxin (LPS) can mediate the production of inflammatory cytokines by enterocytes. DESIGN: Laboratory experiment. SETTING: Teaching hospital and burns unit, USA. MATERIAL: Caco-2 cells (HTB38, human adenocarcinoma, and colon). MAIN OUTCOME MEASURES: Concentrations of tumour necrosis factor alpha (TNF-alpha), interleukin 6 (IL-6) and prostaglandin E2 (PGE2) in cell culture supernatants. RESULTS: LPS significantly increased the production of TNF from 8.9 to 26.4 units/ml in 24 h and this increase persisted at a lower level for 4 days with an increase from 2.3 to 9 units/ml at a cell concentration of 2 x 10(5) cells/ml. There was no increase in TNF production when the cells were cultured at 5 x 10(5) cells ml with LPS. At a concentration of 2 x 10(5) cells/ml, the cells produced small amounts of IL-6 in 24 h or 4 day cultures with or without LPS. At a concentration of 5 x 10(5) cells/ml, LPS significantly increased IL-6 production in 24 h from 142 to 433 units/ml and from 106 to 250 units/ml in 4 days. The amount of IL-6 produced by LPS-stimulated cells was greater at 1 day than at 4 days. There was no significant difference in PGE2 production by the cells under any of the incubation conditions. CONCLUSION: Enterocytes can produce TNF and IL-6, and endotoxin can increase the production of these cytokines by enterocytes. The gut therefore has the potential to become an important source of inflammatory cytokines.

Caco-2 Cells↗

PubMed1996

Linkage of aerobic glycolysis to sodium-potassium transport in rat skeletal muscle. Implications for increased muscle lactate production in sepsis.

Although a linkage between aerobic glycolysis and sodium-potassium transport has been demonstrated in diaphragm, vascular smooth muscle, and other cells, it is not known whether this linkage occurs in skeletal muscle generally. Metabolism of intact hind-leg muscles from young rats was studied in vitro under aerobic incubation conditions. When sodium influx into rat extensor digitorum longus (EDL) and soleus muscles was facilitated by the sodium ionophore monensin, muscle weight gain and production of lactate and alanine were markedly stimulated in a dose-dependent manner. Although lactate production rose in both muscles, it was more pronounced in EDL than in soleus. Monensin-induced lactate production was inhibited by ouabain or by incubation in sodium-free medium. Preincubation in potassium-free medium followed by potassium re-addition also stimulated ouabain-inhibitable lactate release. Replacement of glucose in the incubation medium with pyruvate abolished monensin-induced lactate production but exacerbated monensin-induced weight gain. Muscles from septic or endotoxin-treated rats exhibited an increased rate of lactate production in vitro that was partially inhibited by ouabain. Increases muscle lactate production in sepsis may reflect linked increases in activity of the Na+, K+-ATPase, consumption of ATP and stimulation of aerobic glycolysis.

Aerobiosis↗

PubMed1996

Identification of altered gene expression in skeletal muscle during sepsis using differential display.

Different aspects of muscle metabolism are altered during sepsis and there is evidence that some of these changes may be regulated at the gene level. Differential display is a recently described technique to identify genes whose expression has changed during a biological process. This technique utilizes reverse transcriptase-polymerase chain reaction (RT-PCR) to compare mRNA signals in tissues during two different conditions. We used differential display to test the hypothesis that gene expression is altered in skeletal muscle during sepsis. Sepsis was induced in rats by cecal ligation and puncture (CLP). Control rats were sham-operated. Sixteen hours after CLP or sham operation, extensor digitorum longus muscles were harvested and RNA was extracted. Following differential display, 30 fragments (F1-F30) were identified that appeared to be uniquely expressed in muscles from sham-operated or septic rats. These fragments were reamplified by PCR and used as probes in Northern blot analysis. Messenger RNA levels corresponding to 2 of the 30 fragments (F5 and F24) were confirmed to be increased by Northern blot analysis in septic muscle. Following cloning and sequencing, F5 was found to display significant homology to the gene sequence of the guanine nucleotide releasing protein MSS4. The sequence of F24 did not match any reported gene sequence and may therefore represent a previously unidentified gene. The results support the hypothesis that gene expression is altered in skeletal muscle during sepsis.

Animals↗

PubMed1996

Energy-ubiquitin-dependent muscle proteolysis during sepsis in rats is regulated by glucocorticoids.

Recent studies suggest that sepsis-induced increase in muscle proteolysis mainly reflects energy-ubiquitin-dependent protein breakdown. We tested the hypothesis that glucocorticoids activate the energy-ubiquitin-dependent proteolytic pathway in skeletal muscle during sepsis. Rats underwent induction of sepsis by cecal ligation and puncture or were sham-operated and muscle protein breakdown rates were measured 16 h later. The glucocorticoid receptor antagonist RU 38486 or vehicle was administered to groups of septic and sham-operated rats. In other experiments, dexamethasone (2.5 or 10 mg/kg) was injected subcutaneously in normal rats. Total and myofibrillar proteolysis was determined in incubated extensor digitorum longus muscles as release of tyrosine and 3-methylhistidine, respectively. Energy-dependent proteolysis was determined in incubated muscles depleted of energy with 2-deoxyglucose and 2,4-dinitrophenol. Levels of muscle ubiquitin mRNA and free and conjugated ubiquitin were determined by Northern and Western blot, respectively. RU 38486 inhibited the sepsis-induced increase in total and myofibrillar energy-dependent protein breakdown rates and blunted the increase in ubiquitin mRNA levels and free ubiquitin. Some, but not all, sepsis-induced changes in ubiquitin protein conjugates were inhibited by RU 38486. Injection of dexamethasone in normal rats increased energy-dependent proteolysis and ubiquitin mRNA levels. The results suggest that glucocorticoids regulate the energy-ubiquitin-dependent proteolytic pathway in skeletal muscle during sepsis.

Animals↗