Effect of acute nutritional deprivation on diaphragm structure and function in adolescent rats.

The influence of 90 h of acute nutritional deprivation (ND; water ad libitum only) on in vitro contractile and fatigue properties, muscle fiber type proportions, and cross-sectional areas (CSA) of the adolescent rat diaphragm was determined. Diaphragm muscle properties in the ND rats were compared with those in control rats (CTL; food and water ad libitum). Acute ND resulted in a 32% reduction in body mass, whereas the body mass of CTL rats increased by 29%. Acute ND resulted in a significant reduction in the mass of the diaphragm (costal, 36%; crural, 43%), soleus (36%), and medial gastrocnemius (45%) muscles. Isometric twitch characteristics of the diaphragm muscle (contraction and half-relaxation times) were prolonged in the ND animals. Peak twitch and maximum tetanic forces were unaffected by ND. Fatigue resistance of the diaphragm muscle was improved in ND animals. Diaphragm muscle fiber type proportions were similar in ND and CTL groups. The CSA of type I and II diaphragm muscle fibers were reduced by 22 and 40%, respectively, in ND animals compared with CTL. We conclude that, whereas an identical protocol of acute ND had no significant effects on diaphragm muscle structure and function in adult rats, adolescent animals exhibit significantly less nutritional reserve. These differences may be due to curtailment of the rapid anabolic rate in growing animals.     

Neutralization of tumor necrosis factor-alpha reverses insulin resistance in skeletal muscle but not adipose tissue

We examined the possible role of tumor necrosis factor-alpha (TNF-alpha) as a mediator of insulin resistance in maturing male Sprague-Dawley rats. Rats were treated either with goat anti-murine TNF-alpha IgG (anti-TNF-alpha) or goat nonimmune IgG (NI) for 7 days. Vascular catheters were implanted, and rats were fasted overnight before hyperinsulinemic euglycemic clamp (HUC) studies were performed. TNF-alpha neutralization increased the rate of glucose infusion required to maintain euglycemia by 68%. Insulin-stimulated glucose transport into individual tissues was measured after bolus administration of 2-deoxy-[(14)C]glucose during HUC. Anti-TNF-alpha administration increased glucose transport in muscles composed predominantly of fast-twitch fibers: white gastrocnemius muscle (68% increase) and tibialis anterior muscle (64% increase). There were nonsignificant trends for increased glucose transport in the slow-twitch soleus muscle and in the mixed-fiber red gastrocnemius muscle. Glucose transport was unchanged in visceral and subcutaneous fat. Anti-TNF treatment did not alter body weight, muscle mass, or fat mass. Anti-TNF-alpha did not alter the distribution of the 17-kDa and 26-kDa forms of TNF-alpha in either muscle or fat. However, anti-TNF-alpha treatment caused an approximately 50% reduction in the secretion of TNF-alpha bioactivity in vitro by explants of visceral and subcutaneous fat. We conclude that TNF-alpha neutralization reversed insulin resistance substantially in fast-twitch muscle and may have done so in other muscles, while having little effect in fat. TNF-alpha neutralization was accompanied by reduced TNF-alpha bioactivity without tissue depletion of TNF-alpha protein.     

Influence of varying degrees of malnutrition on IGF-I expression in the rat diaphragm.

This study evaluated the impact of varying degrees of prolonged malnutrition on the local insulin-like growth factor-I (IGF-I) system in the costal diaphragm muscle. Adult rats were provided with either 60 or 40% of usual food intake over 3 wk. Nutritionally deprived (ND) animals (i.e., ND60 and ND40) were compared with control (Ctl) rats fed ad libitum. Costal diaphragm fiber types and cross-sectional areas were determined histochemically. Costal diaphragm muscle IGF-I mRNA levels were determined by RT-PCR. Serum and muscle IGF-I peptide levels were determined by using a rat-specific radioimmunoassay. The body weights of Ctl rats increased by 5%, whereas those of ND60 and ND40 animals decreased by 16 and 26%, respectively. Diaphragm weights were reduced by 17 and 27% in ND60 and ND40 animals, respectively, compared with Ctl. Diaphragm fiber proportions were unaffected by either ND regimen. Significant atrophy of both type IIa and IIx fibers was noted in the ND60 group, whereas atrophy of all three fiber types was observed in the diaphragm of ND40 rats. Serum IGF-I levels were reduced by 62 and 79% in ND60 and ND40 rats, respectively, compared with Ctl. Diaphragm muscle IGF-I mRNA levels in both ND groups were similar to those noted in Ctl. In contrast, IGF-I concentrations were reduced by 36 and 42% in the diaphragm muscle of ND60 and ND40 groups, respectively, compared with Ctl. We conclude that the local (autocrine/paracrine) muscle IGF-I system is affected in our models of prolonged ND. We propose that this contributes to disordered muscle protein turnover and muscle cachexia with atrophy of muscle fibers. This is particularly so in view of recent data demonstrating the importance of the autocrine/paracrine system in muscle growth and maintenance of fiber size.     

Effect of nutritional deprivation on diaphragm contractility and muscle fiber size

The influence of nutritional deprivation on the contractile and fatigue properties of the diaphragm was studied in adult rats. Food access was restricted to one-third of normal daily intake until the body weight of nutritionally deprived (ND) animals was approximately 50% of controls (CTL). Isometric contractile properties were studied in an in vitro nerve muscle strip preparation. Both twitch (Pt) and tetanic (Po) tensions of diaphragms from the ND animals were markedly reduced compared with CTL; however, Pt/Po was higher for the ND group. The shape of the force-frequency curve (normalized to Po) was generally similar between the two groups, except at 5 and 10 pulses/s stimulation, where greater relative tensions were produced in diaphragms from the ND animals. Diaphragm fatigue was induced by repetitive stimulation at either 20 or 100 pulses/s. Endurance time (defined as the time required for tension to fall to 50% of initial) of diaphragms from ND animals was prolonged at both 20 and 100 pulses/s. Immediately after induction of fatigue, force-frequency curves for both ND and CTL diaphragms were shifted to the right. However, this rightward shift was attenuated in the ND group compared with CTL. Nutritional deprivation had no effect on the proportions of different fiber types within the diaphragm but did result in a significant decrease in the cross-sectional area of both fast-and slow-twitch fibers. This decrease in cross-sectional area was significantly greater for fast-twitch fibers. We conclude that these changes in diaphragm contractile and fatigue properties occur as a result of the influence of malnutrition on muscle fiber cross-sectional area.     

Effect of corticosteroids on diaphragm fatigue, SDH activity, and muscle fiber size.

The influence of dexamethasone on diaphragm (DIA) fatigue, oxidative capacity, and fiber cross-sectional areas (CSA) was determined in growing hamsters. One group received dexamethasone by daily subcutaneous injection for 21 days (D animals), while pair-weight (P) and free-eating controls (CTL) received saline subcutaneously. Isometric contractile properties of the DIA were determined in vitro by supramaximal direct muscle stimulation in the presence of curare. DIA fatigue resistance was determined through repetitive stimulation at 40 pulses/s for 2 min. A computer-based image-processing system was used to histochemically determine muscle fiber-type proportions, CSA, and succinate dehydrogenase activities. The medial gastrocnemius muscle (MG) was used as a limb muscle control, with histochemical studies being performed on both the superficial (s) and deep/red (r) portions. Dexamethasone markedly attenuated the normal increment in body weight over the 3-wk period. DIA fatigue resistance was significantly reduced in the D compared with CTL and P animals. Dexamethasone had no effect on fiber-type proportions of the DIA or MGr (MGs contained only type II fibers). In the DIA, the CSA of type II fibers was reduced 33% in D and 18.5% in P animals compared with CTL. Although no significant atrophy was noted in the type I DIA fibers of either D or P animals, a trend toward significance was noted in D animals compared with CTL. In the MGs, the CSA of type II fibers was reduced 33% in D and 16.5% in P animals compared with CTL. Significant atrophy of type I and II fibers of the MGr was noted in D animals compared with CTL (33.8 and 35% reductions, respectively).(ABSTRACT TRUNCATED AT 250 WORDS)     

Effect of acute nutritional deprivation on diaphragm structure and function

The influence of 90 h of acute nutritional deprivation (ND) on the cross-sectional areas of muscle fibers and the contractile and fatigue properties of the adult rat diaphragm were determined. Isometric contractile properties and fatigue resistance of the diaphragm were measured by means of an in vitro nerve-muscle strip preparation. Contractions were evoked by using phrenic nerve stimulation (left hemidiaphragm) or direct muscle stimulation (right hemidiaphragm) in the presence of curare. Acute ND resulted in a 20% reduction in body weight. No significant decrements in diaphragm or soleus weights were noted in the ND animals compared with controls (CTL), whereas the weight of the medial gastrocnemius was reduced by 20% in the ND animals. Peak twitch and tetanic tensions (normalized for the weight of the diaphragm strip) were not reduced in ND compared with CTL animals after either nerve or muscle stimulation. The fatigue index of the diaphragm was significantly reduced in ND animals only after nerve stimulation. After the fatigue test, there was rapid recovery of the additional fatigue noted with nerve stimulation. The proportions of type I and II muscle fibers of the diaphragm were similar in the CTL and ND animals. No differences in diaphragm cross-sectional areas were noted for either type I or II muscle fibers in the CTL and ND animals. It is concluded that acute ND has no effect on diaphragm contractility or morphometry and only an inconsequential influence on diaphragm fatigue.     

Insulin-like growth factor I prevents corticosteroid-induced diaphragm muscle atrophy in emphysematous hamsters

The aim of this study was to evaluate whether recombinant human insulin-like growth factor I (rhIGF-I) could attenuate or prevent diaphragm (DIA) fiber atrophy with corticosteroid (CS) administration to emphysematous (EMP) hamsters. DIA muscle IGF-I responses to CS administration with and without exogenous rhIGF-I administration were evaluated. Three groups were studied: 1) EMP; 2) EMP + triamcinolone (T; 0.4 mg.kg-1.day-1 im); and 3) EMP + T + IGF-I (600 microg/day by constant infusion). After 4 wk, the DIA was analyzed histochemically and biochemically (IGF-I mRNA levels by RT-PCR and endogenous and exogenous IGF-I peptide levels immunochemically). Body weights of EMP-T progressively decreased, while those of EMP and EMP-T-IGF-I remained stable despite similarly reduced food intake in both T groups. DIA weight was reduced with T but preserved with rhIGF-I infusion. DIA fiber proportions were similar among the groups. The cross-sectional areas of types I, IIa, and IIx fibers were reduced (17 to 31%) with T administration but unchanged with rhIGF-I infusion. DIA IGF-I mRNA levels were similar across all groups. By contrast, the endogenous DIA IGF-I levels were reduced (41%) in the EMP-T-IGF-I animals. Total DIA IGF-I levels (endogenous + exogenous) were still significantly reduced. IGF-I immunoreactivity confirmed this reduction in all DIA fibers. We conclude that DIA fiber atrophy with T was completely prevented by exogenous rhIGF-I administration. This effect was likely mediated by the pharmacological influences of exogenously administered rhIGF-I. We speculate that this results from increased bioavailability of free IGF-I to react with muscle receptors. Reduced endogenous IGF-I levels in the DIA likely reflect a negative-feedback influence. These results may have important clinical implications for treatment options to offset the adverse effects of CS on the respiratory muscles in patients with chronic lung disorders.     

TNF-alpha is crucial for the development of mast cell-dependent colitis in mice

Inflammatory bowel disease (IBD) describes chronic inflammatory conditions of the gastrointestinal tract, and TNF-alpha plays a pivotal role in mediating the response. The proinflammatory cytokine TNF-alpha is rapidly released by mast cells after degranulation. In the present study, we hypothesized TNF-alpha to be an important player in our recently described mast cell-dependent murine model for IBD. The effect of neutralizing anti-TNF-alpha MAb was studied on colonic hypersensitivity in mice induced by a skin application of dinitrofluorobenzene (DNFB) followed by an intrarectal challenge with dinitrobenzene sulfonic acid. Features of the colonic hypersensitivity response included diarrhea, mast cell infiltration and activation, infiltration of inflammatory cells in the colon, colonic patch hypertrophy, and increased mast cell-derived TNF-alpha levels in the colon. Anti-TNF-alpha MAb could effectively abrogate diarrhea in DNFB-sensitized mice 72 h after the challenge. The numbers of colonic patches and total tissue damage scores were reduced by anti-TNF-alpha MAb treatment in DNFB-sensitized mice 72 h after the challenge. Mast cell infiltration and activation remained unaffected by neutralizing anti-TNF-alpha MAb. Treatment with the corticosteroid dexamethasone, a frequently used therapeutic treatment in IBD, resulted in a reduction of diarrhea, cellular infiltration, and total tissue damage scores to the same extent as anti-TNF-alpha MAb. Additionally, dexamethasone treatment could also reduce total TNF-alpha levels in the colon, mast cell numbers, and mast cell activation in both vehicle- and DNFB-sensitized mice 72 h after the challenge. These findings suggest that TNF-alpha can play an instrumental role in causing inflammatory responses in the present murine model for IBD downstream from mast cell activation.     

Effects of anti-tumor necrosis factor-alpha and anti-intercellular adhesion molecule-1 antibodies on ischemia/reperfusion lung injury

Inhibition of neutrophil activation and adherence to endothelium by antibodies to tumor necrosis factor-alpha (TNF-alpha) and intercellular adhesion molecules (ICAM-1), respectively, might attenuate ischemia-reperfusion injury (I/R). I/R was conducted in an isolated rat lung model. Anti-TNF-alpha antibody and/or anti-ICAM-1 antibody were added before ischemia or after reperfusion. Hemodynamic changes, lung weight gain (LWG), capillary filtration coefficients (Kfc), and pathologic changes were assessed to evaluate the severity of I/R. The LWG, Kfc, pathological changes and lung injury score of treatment groups with anti-TNF-alpha antibody treatment, either pre-ischemia or during reperfusion, were less than those observed in control groups. Similar findings were found in group treated with anti-ICAM-1 antibody or combination therapy during reperfusion. In contrast, pre-I/R treatment with anti-ICAM-1 antibody induced severe lung edema and failure to complete the experimental procedure. No additional therapeutic effect was found in combination therapy. We conclude that TNF-alpha and ICAM-1 play important roles in I/R. Anti-TNF-alpha antibody has therapeutic and preventive effects on I/R. However, combined therapy with anti-TNF-alpha antibody and anti-ICAM-1 antibody may have no additive effect and need further investigation.     

Role of endogenous tumor necrosis factor alpha and gamma interferon in resistance to Corynebacterium pseudotuberculosis infection in mice

The production and roles of endogenous tumor necrosis factor alpha (TNF-alpha) and gamma interferon (IFN-gamma) in the infection of Corynebacterium (C.) pseudotuberculosis were investigated in mice. The maximum levels of TNF-alpha and IFN-gamma were detected on day 4 after infection. The administration of anti-TNF-alpha monoclonal antibody (mAb) as well as anti-IFN-gamma mAb increased bacterial proliferation in the organs, leading to the death of infected mice, but anti-IFN-gamma mAb showed a less marked effect than anti-TNF-alpha mAb. The suppressive effect of anti-TNF-alpha and anti-IFN-gamma mAbs on anticorynebacterial resistance was augmented by the simultaneous administration of these antibodies. Anti-TNF-alpha mAb was found to be highly effective when administered on day 0 and day 4, suggesting that TNF-alpha produced during the early stage of infection is critical for the generation of resistance. Histologically, many microabscesses, severe follicular swelling and lymphocyte destruction were observed in mice treated with anti-TNF-alpha or anti-IFN-gamma mAb. Injection of anti-CD4 or anti-CD8 mAb also resulted in significantly increased mortality and a marked suppression of IFN-gamma production, but had no effect on TNF-alpha production. Carrageenan also showed a marked effect on the exacerbation of infection. Taken together, these results suggest that endogenously produced TNF-alpha and IFN-gamma are both essential to the host defense against C. pseudotuberculosis infection and that these cytokines may have an additive effect.     

Microtubule disassembly induces cytoskeletal remodeling and lung vascular barrier dysfunction: role of Rho-dependent mechanisms

Barrier dysfunction of pulmonary endothelial monolayer is associated with dramatic cytoskeletal reorganization, activation of actomyosin contractility, and gap formation. The linkage between the microtubule (MT) network and the contractile cytoskeleton has not been fully explored, however, clinical observations suggest that intravenous administration of anti-cancer drugs and MT inhibitors (such as the vinca alkaloids) can lead to the sudden development of pulmonary edema in breast cancer patients. In this study, we investigated the crosstalk between MT and actomyosin cytoskeleton and characterized specific molecular mechanisms of endothelial cells (EC) barrier dysfunction induced by MT inhibitor nocodazole (ND). Our results demonstrate that MT disassembly by ND induced rapid decreases in transendothelial electrical resistance (TER) and actin cytoskeletal remodeling, indicating EC barrier dysfunction. These effects involved ND-induced activation of Rho GTPase. Rho-mediated activation of its downstream target, Rho-kinase, induced phosphorylation of Rho-kinase effector EC MLC phosphatase (MYPT1) at Thr(696) and Thr(850) resulting in MYPT1 inactivation. Phosphatase inhibition leaded to accumulation of diphospho-MLC, which induced acto-myosin polymerization, stress fiber formation and gap formation. Inhibition of Rho-kinase by Y27632 abolished ND-induced MYPT1 phosphorylation, MLC phosphorylation, and stress fiber formation. In addition, MT preservation via the MT stabilizer paclitaxel, Rho inhibition (via C3 exotoxin, or dominant negative (DN)-Rho, or DN-Rho-kinase) attenuated ND-induced TER decreases, stress fiber formation and MLC phosphorylation. Collectively, our results demonstrate a leading role for Rho-dependent mechanisms in crosstalk between the MT and actomyosin cytoskeleton, and suggest Rho-kinase and MYPT1 as major Rho effectors mediating pulmonary EC barrier disruption in response to ND-induced MT disassembly.     

Influences of IGF-I gene disruption on the cellular profile of the diaphragm

The impact of a targeted disruption of the Igf1 gene, encoding the insulin-like growth factor I (IGF-I), on diaphragm (DIA) cellularity was studied in 2-mo-old homozygous mutant [IGF-I(-/-)] mice and their wild-type [WT; i.e., IGF-I(+/+)] littermates. DIA fiber types were classified histochemically. DIA fiber cross-sectional areas (CSA) were determined from digitized muscle sections, and fiber succinate dehydrogenase (SDH) activity was determined histochemically using a microdensitometric procedure. An acidic ATPase reaction was used to visualize capillaries. Myosin heavy chain (MyHC) isoforms were identified by SDS-PAGE, and their proportions were determined by scanning densitometry. The body weight of IGF-I(-/-) animals was 32% that of WT littermates. DIA fiber type proportions were unchanged between the groups. The CSAs of types I, IIa, and IIx DIA fibers of IGF-I(-/-) mutants were 63, 68, and 65%, respectively, those of WT animals (P < 0.001). The DIA thickness and the number of fibers spanning its entire thickness were reduced by 36 and 25%, respectively, in IGF-I(-/-) mice (P < 0. 001). SDH activity was significantly increased in all three types of DIA fibers of IGF-I(-/-) mutants (P < 0.05). The number of capillaries per fiber was reduced approximately 30% in IGF-I(-/-) animals, whereas the capillary density was preserved. The proportions of MyHC isoforms were similar between the groups. Muscle hypoplasia likely reflects the importance of IGF-I on cell proliferation, differentiation, and apoptosis (alone or in combination) during development, although reduced cell size highlights the importance of IGF-I on rate and/or maintenance of DIA fiber growth in the postnatal state. Reduced capillarity may result from both direct and indirect influences on angiogenesis. Improved oxidative capacity likely reflects DIA compensatory mechanisms in IGF-I(-/-) mutants.     

The mechanism of superantigen-mediated toxic shock: not a simple Th1 cytokine storm

The profound clinical consequences of Gram-positive toxic shock are hypothesized to stem from excessive Th1 responses to superantigens. We used a new superantigen-sensitive transgenic model to explore the role of TCRalphabeta T cells in responses to staphylococcal enterotoxin B (SEB) in vitro and in two different in vivo models. The proliferative and cytokine responses of HLA-DR1 spleen cells were 100-fold more sensitive than controls and were entirely dependent on TCRalphabeta T cells. HLA-DR1 mice showed greater sensitivity in vivo to two doses of SEB with higher mortality and serum cytokines than controls. When d-galactosamine was used as a sensitizing agent with a single dose of SEB, HLA-DR1 mice died of toxic shock whereas controls did not. In this sensitized model of toxic shock there was a biphasic release of cytokines, including TNF-alpha, at 2 h and before death at 7 h. In both models, mortality and cytokine release at both time points were dependent on TCRalphabeta T cells. Anti-TNF-alpha pretreatment was protective against shock whereas anti-IFN gamma pretreatment and delayed anti-TNF-alpha treatment were not. Importantly, anti-TNF-alpha pretreatment inhibited the early TNF-alpha response but did not inhibit the later TNF-alpha burst, to which mortality has previously been attributed. Splenic T cells were shown definitively to be the major source of TNF-alpha during the acute cytokine response. Our results demonstrate unequivocally that TCRalphabeta T cells are critical for lethality in toxic shock but it is the early TNF-alpha response and not the later cytokine surge that mediates lethal shock.     

In vivo neutralization of TNF-alpha promotes humoral autoimmunity by preventing the induction of CTL.

Neutralization of TNF-alpha in humans with rheumatoid arthritis or Crohn's disease has been associated with the development of humoral autoimmunity. To determine the effect of TNF-alpha neutralization on cell-mediated and humoral-mediated responses, we administered anti-TNF-alpha mAb to mice undergoing acute graft-vs-host disease (GVHD) using the parent-into-F(1) model. In vivo neutralization of TNF-alpha blocked the lymphocytopenic features characteristic of acute GVHD and induced a lupus-like chronic GVHD phenotype (lymphoproliferation and autoantibody production). These effects resulted from complete inhibition of detectable antihost CTL activity and required the presence of anti-TNF-alpha mAb for the first 4 days after parental cell transfer, indicating that TNF-alpha plays a critical role in the induction of CTL. Moreover, an in vivo blockade of TNF-alpha preferentially inhibited the production of IFN-gamma and blocked IFN-gamma-dependent up-regulation of Fas; however, cytokines such as IL-10, IL-6, or IL-4 were not inhibited. These results suggest that a therapeutic TNF-alpha blockade may promote humoral autoimmunity by selectively inhibiting the induction of a CTL response that would normally suppress autoreactive B cells.     

Evaluation of TNF-alpha and IL-1 blockade in collagen-induced arthritis and comparison with combined anti-TNF-alpha/anti-CD4 therapy

We have evaluated the effects of anti-TNF-alpha, anti-IL-1, and combined anti-TNF-alpha/anti-CD4 therapy in collagen-induced arthritis. Blockade of TNF-alpha or IL-1 before disease onset delayed, but did not prevent, the induction of arthritis. When treatment was initiated after onset of arthritis, anti-TNF-alpha, anti-IL-1beta, and anti-IL-1R (which blocks IL-1alpha and IL-1beta) were all found to be effective in reducing the severity of arthritis, with anti-IL-1R and anti-IL-1beta showing greater efficacy than anti-TNF-alpha. Anti-IL-1beta was equally as effective as anti-IL-1R, indicating that IL-1beta plays a more prominent role than IL-1alpha in collagen-induced arthritis. An additive effect was observed between anti-TNF-alpha and anti-IL-1R in the prevention of joint erosion and in normalization of the levels of serum amyloid P. Combined anti-TNF-alpha/anti-CD4 therapy also caused normalization of serum amyloid P levels. The therapeutic effect of anti-TNF-alpha plus anti-CD4 was comparable to that of anti-TNF-alpha plus anti-IL-1R, suggesting that combined anti-TNF-alpha/anti-CD4 therapy prevents both TNF-alpha- and IL-1-mediated pathology. Anti-TNF-alpha treatment reduced IL-1beta expression in the joint and, conversely, anti-IL-1beta treatment reduced TNF-alpha expression. Combined anti-TNF-alpha/anti-CD4 treatment almost completely blocked the expression of IL-1beta, thereby confirming the ability of this form of combination therapy to prevent IL-1ss-mediated pathology.     

Resistance exercise decreases skeletal muscle tumor necrosis factor alpha in frail elderly humans.

Skeletal muscle protein and function decline with advancing age but the underlying pathophysiology is poorly understood. To test the hypothesis that the catabolic cytokine tumor necrosis factor alpha (TNF-alpha) contributes to this process, we studied the effects of aging and resistance exercise on TNF-alpha expression in human muscle. Using in situ hybridization, TNF-alpha message was localized to myocytes in sections of skeletal muscle from elderly humans. Both TNF-alpha mRNA and protein levels were elevated in skeletal muscle from frail elderly (81+/-1 year) as compared to healthy young (23+/-1 year) men and women. To determine whether resistance exercise affects TNF-alpha expression, frail elderly men and women were randomly assigned to a training group or to a nonexercising control group. Muscle biopsies were performed before and after 3 months. Muscle TNF-alpha mRNA and protein levels decreased in the exercise group but did not change in the control group. Muscle protein synthesis rate in the exercise group was inversely related to levels of TNF-alpha protein. These data suggest that TNF-alpha contributes to age-associated muscle wasting and that resistance exercise may attenuate this process by suppressing skeletal muscle TNF-alpha expression.     

Effects of nandrolone decanoate on the neuromuscular junction of rats submitted to swimming

This study addressed the effects of nandrolone decanoate (ND) on contractile properties and muscle fiber characteristics of rats submitted to swimming. Male Wistar rats were grouped in sedentary (S), swimming (Sw), sedentary+ND (SND), and swimming+ND (SwND), six animals per group. ND (3 mg/kg) was injected (subcutaneously) 5 days/week, for 4 weeks. Swimming consisted of 60-min sessions (load 2%), 5 days/week, for 4 weeks. After this period, the sciatic nerve extensor digitorum longus (EDL) muscle was isolated for myographic recordings. Fatigue resistance was assessed by the percent (%) decline of 180 direct tetanic contractions (30 Hz). Safety margin of synaptic transmission was determined from the resistance to the blockade of indirectly evoked twitches (0.5 Hz) induced by pancuronium (5 to 9x10(-7) M). EDL muscles were also submitted to histological and histochemical analysis (haematoxylin-eosin (HE); nicotinamide adenine dinucleotide-tetrazolium reductase (NADH-TR)). Significant differences were detected by two-way ANOVA (p<0.05). ND did not change body mass, fatigue resistance or kinetic properties of indirect twitches in either sedentary or swimming rats. In contrast, ND reduced the safety margin of synaptic transmission in sedentary animals (SND=53.3+/-4.7% vs. S=75.7+/-2.0%), but did not affect the safety margin in the swimming rats (SwND=75.81+/-3.1% vs. Sw=71.0+/-4.0%). No significant difference in fiber type proportions or diameters was observed in EDL muscle of any experimental group. These results indicate that ND does not act as an ergogenic reinforcement in rats submitted to 4 weeks of swimming. On the other hand, this study revealed an important toxic effect of ND, that it reduces the safety margin of synaptic transmission in sedentary animals. Such an effect is masked when associated with physical exercise.     

Alveolar macrophage activation after trauma-hemorrhage and sepsis is dependent on NF-kappaB and MAPK/ERK mechanisms

The acute respiratory distress syndrome (ARDS) is a major cause of morbidity after injury. We hypothesized that alveolar macrophage (AMPhi) chemokine and cytokine release after hemorrhage and sepsis is regulated by NF-kappaB and MAPK. Adult male rats underwent soft tissue trauma and hemorrhagic shock (~90 min) followed by crystalloid resuscitation. Sepsis was induced by cecal ligation and puncture (CLP) 20 h after resuscitation. AMPhi were harvested, and TNF-alpha, IL-6, and macrophage inflammatory protein (MIP)-2 release and serum IL-6 and TNF-alpha levels were measured at 5 h after HCLP. Lung tissues were analyzed for activation of NF-kappaB, myeloperoxidase activity, and wet/dry weight ratio. In control animals, AMPhi were stimulated with LPS with or without inhibitors of NF-kappaB and MAPK. Serum TNF-alpha and IL-6 levels and spontaneous AMPhi TNF-alpha and MIP-2 release were elevated (P < 0.05) after HCLP, concomitantly with the development of lung edema and leukocyte activation. Activation of NF-kappaB increased in lungs from the hemorrhage and CLP group compared with shams. Inhibition of NF-kappaB or the upstream MAPK significantly decreased LPS-stimulated AMPhi activation. Because enhanced release of inflammatory mediators by AMPhi may contribute to ARDS after severe trauma, inhibition of intracellular signaling pathways represents a target to attenuate organ injury under those conditions.     

Effects of undernutrition on diaphragm fiber size, SDH activity, and fatigue resistance

The influence of prolonged nutritional deprivation on the succinate dehydrogenase (SDH) activity and cross-sectional areas of individual fibers in the rat diaphragm and deep portion of the medial gastrocnemius (MGr) muscles was determined. Fatigue resistance of the diaphragm was measured by means of an in vitro nerve-muscle strip preparation. Fiber SDH activity and cross-sectional area were quantified by means of an image processing system. Diaphragm fatigue resistance was significantly improved in the nutritionally deprived (ND) group. In both muscles, nutritional deprivation resulted in a significant decrease in fiber cross-sectional area (both type I and II), type II fibers showing greater atrophy. The SDH activities of type I and II fibers in the diaphragm were not affected by nutritional deprivation. This contrasted with a significant decrease in the SDH activity of both type I and II fibers in the MGr of ND animals. An assessment of the interrelationships between fiber atrophy and fiber SDH activity revealed a greater effect of malnutrition on those diaphragm type II fibers that had the lowest relative SDH activities and the largest cross-sectional areas. By comparison, the effect of malnutrition on type I and II fibers in the MGr was nonselective with regard to fiber SDH activity. We conclude that the enhanced diaphragm fatigue resistance in the ND animals does not result from an increase in the oxidative capacity of muscle fibers and is best explained by the pattern of diaphragm muscle fiber atrophy.     

A novel receptor tyrosine kinase, Mer, inhibits TNF-alpha production and lipopolysaccharide-induced endotoxic shock

The regulation of monocyte function and the inhibition of TNF-alpha production during bacterial sepsis are critical in attenuating adverse host responses to endotoxemia. To study the function of a novel receptor tyrosine kinase, mer, that is expressed in monocytes, we generated mice (merkd) that lack the signaling tyrosine kinase domain. Upon LPS challenge, merkd animals died of endotoxic shock (15/17, 88.2%), whereas control wild-type mice survived (1/15, 6.7% died). Susceptible merkd mice exhibited edema, leukocyte infiltration, and signs of endotoxic shock that correlated with higher levels of TNF-alpha found in the serum of merkd mice as compared with wild-type control animals. Death due to LPS-induced endotoxic shock in merkd mice was blocked by administration of anti-TNF-alpha Ab, suggesting that overproduction of this cytokine was principally responsible for the heightened suseptibility. The increase in TNF-alpha production appeared to be the result of a substantial increase in the LPS-dependent activation of NF-kappa B nuclear translocation resulting in greater TNF-alpha production by macrophages from merkd mice. Thus, Mer receptor tyrosine kinase signaling participates in a novel inhibitory pathway in macrophages important for regulating TNF-alpha secretion and attenuating endotoxic shock.