[Traumatic brain injury]. / Schädel-Hirn-Trauma.

Traumatic brain injury (TBI) describes parenchymal brain damage caused by external forces to the head. It has a massive personal and socioeconomic impact, as it is a disease with high morbidity and mortality. Both young and old people are affected, as a result of traffic or sports accidents as well as due to falls at home. The term TBI encompasses various clinical pictures, differing considerably in cause, prognosis and therapy. What they all have in common is the pathophysiological cascade that develops immediately after the initial trauma and which can persist for several days and weeks. In this phase, medical treatment, whether surgical or pharmacological, attempts to reduce the consequences of the primary damage. The aim is to maintain adequate cerebral perfusion pressure and to reduce intracranial pressure.

Obesity in regional anesthesia--a risk factor for peripheral catheter-related infections.

BACKGROUND: Obesity is believed to increase the risk of surgical site infections and possibly increase the risk of catheter-related infections in regional anesthesia. We, therefore, analyzed the influence of obesity on catheter-related infections defined within a national registry for regional anesthesia. METHODS: The German Network for Regional Anesthesia database with 25 participating clinical centers was analyzed between 2007 and 2012. Exactly, 28,249 cases (13,239 peripheral nerve and 15,010 neuraxial blocks) of patients ≥ 14 years were grouped in I: underweight (BMI 13.2-18.49 kg/m(2) , n = 597), II: normal weight (BMI 18.5-24.9 kg/m(2) , n = 9272), III: overweight (BMI 25.0-29.9 kg/m(2) , n = 10,632), and IV: obese (BMI 30.0-70.3 kg/m(2) , n = 7,744). The analysis focused on peripheral and neuraxial catheter-related infections. Differences between the groups were tested with non-parametric ANOVA and chi-square (P < 0.05). Binary logistic regression was used to compare obese, overweight, or underweight patients with normal weight patients. Odds ratios (OR and 95% confidence interval) were calculated and adjusted for potential confounders. RESULTS: Confounders with significant influence on the risk for catheter-related infections were gender, age, ASA score, diabetes, preoperative infection, multiple skin puncture, and prolonged catheter use. The incidence (normal weight: 2.1%, obese: 3.6%; P < 0.001) and the risk of peripheral catheter-related infection was increased in obese compared to normal weight patients [adjusted OR: 1.69 (1.25-2.28); P < 0.001]. In neuraxial sites, the incidence of catheter-related infections differed significantly between normal weight and obese patients (normal weight: 3.2%, obese: 2.3%; P = 0.01), whereas the risk was comparable [adjusted OR: 0.95 (0.71-1.28); P = 0.92]. CONCLUSION: This retrospective cohort study suggests that obesity is an independent risk factor for peripheral, but not neuraxial, catheter-related infections.

Transport of energy in muscle: the phosphorylcreatine shuttle.

In order to explain the insulin-like effect of exercise, it was proposed in 1951 that contracting muscle fibers liberate creatine, which acts to produce an acceptor effect--later called respiratory control--on the muscle mitochondria. The development of this notion paralleled the controversy between biochemists and physiologists over the delivery of energy for muscle contraction. With the demonstration of functional compartmentation of creatine kinase on the mitochondrion, it became clear that the actual form of energy transport in the muscle fiber is phosphorylcreatine. The finding of an isoenzyme of creatine phosphokinase attached to the M-line region of the myofibril revealed the peripheral receptor for the mitochondrially generated phosphorylcreatine. This established a molecular basis for a phosphorylcreatine-creatine shuttle for energy transport in heart and skeletal muscle and provided an explanation for the inability to demonstrate experimentally a direct relation between muscle activity and the concentrations of adenosine triphosphate and adenosine diphosphate.

Isozymes of phenylalanine hydroxylase.

Three isozymes of phenylalanine hydroxylase exist in adult rat liver. They are chromatographically unique. Partial chracterization suggests that they are similar in chemical properties and differ only in charge. Estimation of the Stokes radii indicates that the isozymes have similar molecular weights of about 200,000. Two isozymes exist in human fetal liver. Alterations of the relative amounts of these isozymes may control the phenotype of the disease phenylketonuria.

Sterile soil from Antarctica: organic analysis.

Soils from the dry-valley region of Antarctica can be sterile by the usual microbiological criteria and yet contain significant amounts of organic carbon. Examination of one such soil shows that the organic material is finely divided anthracite coal. These findings have significant implications for the biological exploration of Mars.

[Incidence of infection from catheter procedures for regional anesthesia: first results from the network of DGAI and BDA]. / Infektionsinzidenz von Katheterverfahren zur Regionalanästhesie : Erste Ergebnisse aus dem Netzwerk von DGAI und BDA.

BACKGROUND: To analyze safety issues of regional anaesthesia and analgesia in Germany only a few single center studies are available. Therefore, the German Society for Anaesthesiology and Intensive Care Medicine (Deutschen Gesellschaft für Anästhesiologie und Intensivmedizin, DGAI) and the Professional Association of German Anaesthetists (Berufsverband Deutscher Anästhesisten, BDA) initiated a network for safety in regional anaesthesia. From this the first results on infectious complications will be reported. MATERIALS AND METHODS: In a Delphi process the documentation of the setup and maintenance of regional anaesthesia and analgesia was agreed with the participants in a working group from the DGAI. After approval by the officially authorized representative for patient data privacy protection a registry was programmed to collect anonymous data. Up to October 2008 data from 6 centers could be analyzed. RESULTS: After testing for plausibility 8,781 regional anaesthesia procedures (22,112 catheter days) could be analyzed. The 5,057 neuraxial and 3,724 peripheral catheter-based procedures were in place for a median of 2.48 days (range 1.0-3.0 days) and 4 severe, 15 moderate and 128 mild infections were recorded. Diabetics were not found to show a statistically significant increase in risk (2.6% compared to 1.9% for non-diabetics: n.s.). Neuraxial procedures seem to have a higher rate of infections than peripheral procedures (2.7% vs. 1.3%, p<0.0001). Multiple punctures of the skin also seem to be associated with a higher infection rate than single skin punctures (4.1% vs. 1.6%, p<0.0001). CONCLUSIONS: Infectious complications of catheter-based regional anaesthesia are common. Strict hygienic standards must therefore be complied with. More data are necessary to calculate risk factors. The registry provided can also be used as a benchmark to reduce these rates further.

Selenoperoxidase-mediated cytoprotection against merocyanine 540-sensitized photoperoxidation and photokilling of leukemia cells.

Photodynamic therapy with the lipophilic sensitizing dye merocyanine 540 (MC540) is a promising new approach for extracorporeal purging of neoplastic cells from autologous remission bone marrow grafts. Resistance-conferring cellular defenses against the cytotoxic effects of MC540/photodynamic therapy have not been well characterized. This study focuses on the cytoprotective effects of the glutathione-dependent selenoperoxidases GPX and PHGPX, which can detoxify a wide variety of hydroperoxides, including lipid-derived species (LOOHs). Murine leukemia L1210 cells were grown in 1% serum media without [L.Se(-)] and with [L.Se(+)] selenium supplementation. L.Se(-) cells expressed 10- to 20-fold lower GPX and PHGPX activities than L.Se(+) controls and were markedly more sensitive to MC540-mediated photoperoxidation (LOOH formation) and clonally assessed photokilling. Susceptibility of L.Se(-) cells to photoperoxidation and photokilling could be fully reversed to L.Se(+) levels by replenishing Se, and partially reversed by treating with Ebselen, a selenoperoxidase mimetic. Altered lipid composition, greater uptake of MC540, and defective catabolism of H2O2 were all ruled out as possible factors in the elevated photosensitivity of L.Se(-) cells. Human leukemia K562 cells (capable of expressing PHGPX but not GPX) exhibited 5- to 10-fold lower PHGPX activity under Se-deficient relative to Se-sufficient conditions. Although MC540 uptake (nmol/mg lipid) by K562 and L1210 cells was essentially the same, the former were more resistant to photoinactivation. However, like murine counterparts, Se-deficient cells were more susceptible to photoperoxidation and photokilling than Se-sufficient controls. These results clearly demonstrate that GPX and/or PHGPX in L1210 cells and PHGPX in K562 cells play an important cytoprotective role during photooxidative stress. Whether membrane damage due to lipid photoperoxidation is causally related to cell death is not certain; however, the parallel effects of Se deficiency on LOOH formation and cell killing are at least consistent with this possibility.

Enzymatic reduction of phospholipid and cholesterol hydroperoxides in artificial bilayers and lipoproteins.

Lipid hydroperoxides (LOOHs) in various lipid assemblies are shown to be efficiently reduced and deactivated by phospholipid hydroperoxide glutathione peroxidase (PHGPX), the second selenoperoxidase to be identified and characterized. Coupled spectrophotometric analyses in the presence of NADPH, glutathione (GSH), glutathione reductase and Triton X-100 indicated that photochemically generated LOOHs in small unilamellar liposomes are substrates for PHGPX, but not for the classical glutathione peroxidase (GPX). PHGPX was found to be reactive with cholesterol hydroperoxides as well as phospholipid hydroperoxides. Kinetic iodometric analyses during GSH/PHGPX treatment of photoperoxidized liposomes indicated a rapid decay of total LOOH to a residual level of 35-40%; addition of Triton X-100 allowed the reaction to go to completion. The non-reactive LOOHs in intact liposomes were shown to be inaccessible groups on the inner membrane face. In the presence of iron and ascorbate, photoperoxidized liposomes underwent a burst of thiobarbituric acid-detectable lipid peroxidation which could be inhibited by prior GSH/PHGPX treatment, but not by GSH/GPX treatment. Additional experiments indicated that hydroperoxides of phosphatidylcholine, cholesterol and cholesteryl esters in low-density lipoprotein are also good substrates for PHGPX. An important role of PHGPX in cellular detoxification of a wide variety of LOOHs in membranes and internalized lipoproteins is suggested from these findings.

Selenoperoxidase-dependent glutathione cycle activity in peroxide-challenged leukemia cells.

Murine leukemia L1210 cells rendered deficient in glutathione peroxidase (GPX) and phospholipid hydroperoxide glutathione peroxidase (PHGPX) by Se deprivation (L.Se(-) cells) were found to be more sensitive to tert-butyl hydroperoxide (t-BuOOH) cytotoxicity than Se-replete controls (L.Se(+) cells). Human K562 cells, which express PHGPX, but not GPX, were also more sensitive to t-BuOOH in the Se-deficient (K.Se(-)) than Se-satisfied (K.Se(+)) condition. In examining the metabolic basis for selenoperoxidase-dependent resistance, we found that glucose-replete Se(-) cells reduce t-BuOOH to t-butanol far more slowly than Se(+) cells, the ratio of the first-order rate constants approximating that of the GPX activities (L1210 cells) or PHGPX activities (K562 cells). Monitoring peroxide-induced changes in GSH and GSSG gave consistent results; e.g., glucose-depleted L.Se(+) cells exhibited a first order loss of GSH that was substantially faster than that of glucose-depleted L.Se(-) cells. Under the conditions used, peroxide-induced conversion of GSH to GSSG could be stoichiometrically reversed by resupplying D-glucose, indicating that no significant lysis or GSSG efflux and/or interchange had taken place. The apparent first-order rate constant for GSH decay increased progressively for L1210 cells expressing a range of GPX activities from approximately 5% to 100%, demonstrating that peroxide detoxification is strictly dependent on enzyme content. The initial rate of 14CO2 release from D-[1-14C]glucose supplied in the medium was much greater for L.Se(+) or K.Se(+) cells than for their respective Se(-) counterparts, consistent with greater hexose monophosphate shunt activity in the former. These results highlight the importance of selenoperoxidase action in the glutathione cycle as a means by which tumor cells cope with hydroperoxide stress.

Prevalence of Corynebacteria in diabetic foot infections.

OBJECTIVE: Microbiological flora of diabetic foot infections are usually polymicrobial and frequently include bacteria of the Corynebacterium sp. (diphtheroids). The purpose of this study was to determine the prevalence of these bacteria in both deep and superficial cultures in diabetic patients with foot infections. RESEARCH DESIGN AND METHODS: The charts of 50 patients of successive admissions to the Orthopedic-Diabetes Service at our hospital were reviewed to obtain the following data: age, sex, ethnic origin, method of treatment of diabetes, blood glucose level, prior antibiotics, and reports of cultures taken from bedside and intraoperative sites. Data were analyzed to compare the prevalence of diphtheroids in reliable versus nonreliable cultures and the influence of other parameters on the presence of these organisms. RESULTS: Fourteen of 19 (74%) of the intraoperative specimens grew diphtheroids compared with 25 of 65 (39%) of the bedside cultures, a highly significant difference. In addition, there was a somewhat greater occurrence of diphtheroids in women compared with men. The likelihood that contamination is the cause for the presence of diphtheroids is highly unlikely, because one arm of the study included cultures derived from deep tissue at the time of the surgical procedure (i.e., the intraoperative cultures). Cultures always grew at least one other organism in addition to the diphtheroid. CONCLUSIONS: Corynebacteria, commonly known as diphtheroids, are present as a part of the polymicrobial flora in a large percentage of diabetic patients with foot infections. Because the diphtheroids were identified in culture material taken in the operating room or at the time of incision and drainage in a higher percentage of patients than in specimens from superficial cultures, it is highly unlikely that they are contaminants.

Blue light-induced retinal lesions, intraretinal vascular leakage and edema formation in the all-cone mouse retina.

Little is known about the mechanisms underlying macular degenerations, mainly for the scarcity of adequate experimental models to investigate cone cell death. Recently, we generated R91W;Nrl(-/-) double-mutant mice, which display a well-ordered all-cone retina with normal retinal vasculature and a strong photopic function that generates useful vision. Here we exposed R91W;Nrl(-/-) and wild-type (wt) mice to toxic levels of blue light and analyzed their retinas at different time points post illumination (up to 10 days). While exposure of wt mice resulted in massive pyknosis in a focal region of the outer nuclear layer (ONL), the exposure of R91W;Nrl(-/-) mice led to additional cell death detected within the inner nuclear layer. Microglia/macrophage infiltration at the site of injury was more pronounced in the all-cone retina of R91W;Nrl(-/-) than in wt mice. Similarly, vascular leakage was abundant in the inner and outer retina in R91W;Nrl(-/-) mice, whereas it was mild and restricted to the subretinal space in wt mice. This was accompanied by retinal swelling and the appearance of cystoid spaces in both inner and ONLs of R91W;Nrl(-/-) mice indicating edema in affected areas. In addition, basal expression levels of tight junction protein-1 encoding ZO1 were lower in R91W;Nrl(-/-) than in wt retinas. Collectively, our data suggest that exposure of R91W;Nrl(-/-) mice to blue light not only induces cone cell death but also disrupts the inner blood-retinal barrier. Macular edema in humans is a result of diffuse capillary leakage and microaneurysms in the macular region. Blue light exposure of the R91W;Nrl(-/-) mouse could therefore be used to study molecular events preceding edema formation in a cone-rich environment, and thus potentially help to develop treatment strategies for edema-based complications in macular degenerations.

Selenoperoxidase-mediated cytoprotection against the damaging effects of tert-butyl hydroperoxide on leukemia cells.

Murine leukemia L1210 cells grown for 5-7 d in the presence of 1% serum without added selenium [Se(-) cells] expressed < 5% of the glutathione peroxidase (GPX) activity of selenium-supplemented controls [Se(+) cells]. Clonogenic survival assays indicated that t-butyl hydroperoxide (t-BuOOH) is much more toxic to Se(-) cells (LC50 approximately 10 microM) than to Se(+) or selenium-repleted [Se(-/+)] cells (LC50 approximately 250 microM). Hypersensitivity of Se(-) cells to t-BuOOH was partially reversed by treating them with Ebselen, a selenoperoxidase mimetic; thus, selenoperoxidase insufficiency was probably the most serious defect of Se deprivation. Cytotoxicity of t-BuOOH was inhibited by desferrioxamine and by alpha-tocopherol, indicating that redox iron and free radical intermediates are involved. Elevated sensitivity of Se(-) cells to t-BuOOH was accompanied by an increased susceptibility to free radical lipid peroxidation, which became even more pronounced in cells that had been grown in arachidonate (20:4, n-6) supplemented media. That glutathione (GSH) is required for cytoprotection was established by showing that Se(+) cells are less resistant to t-BuOOH after exposure to buthionine sulfoximine (BSO), an inhibitor of GSH synthesis, or 1,3-bis(2-chloroethyl)-1-nitrosourea (BCNU), an inhibitor of glutathione reductase. Coupled enzymatic assays indicated that Se(+) or Se(-/+) cells metabolize t-BuOOH 20-25 times more rapidly than Se(-), consistent with the measured difference in GPX activities of these cells. Correspondingly, when challenged with t-BuOOH, Se(+) cells showed an initial loss of GSH and elevation of GSSG that exceeded that of Se(-) cells. It was further shown that like Se(-) cells, BSO- or BCNU-treated Se(+) cells metabolize t-BuOOH more slowly than nontreated controls. These results clearly indicate that selenoperoxidase action in the glutathione cycle is a vital element in cellular defense against toxic hydroperoxides.

Lipid peroxidation in photodynamically stressed mammalian cells: use of cholesterol hydroperoxides as mechanistic reporters.

Photodynamic action of merocyanine 540, an antileukemic sensitizing dye, on murine L1210 cells results in the formation of lipid hydroperoxides and loss of cell viability. High-performance liquid chromatography with mercury cathode electrochemical detection was used for determining lipid oxidation products, including the following cholesterol-derived hydroperoxides: 5 alpha-OOH, 6 alpha-OOH, 6 beta-OOH, and unresolved 7 alpha, 7 beta-OOH. Among these species, 5 alpha-, 6 alpha-, and 6 beta-OOH (singlet oxygen adducts) were predominant in the early stages of photooxidation, whereas 7 alpha- and 7 beta-OOH (products of free radical reactions) became so after prolonged irradiation or during dark incubation after exposure to a light dose. These mechanistic changes were studied in a unique way by monitoring shifts in the peroxide ratio, i.e., 7-OOH/5 alpha-OOH, or 7-OOH/6-OOH. When cells (10(7)/ml) were exposed to a visible light fluence of 0.6 J/cm2 in the presence of 10 microM merocyanine 540, 7-OOH/5 alpha-OOH increased by approximately 100% after 2 h of dark incubation at 37 degrees C. The increase was much larger (approximately 250%) when cells were photooxidized after treatment with 1 microM ferric-8-hydroxyquinoline, a lipophilic iron donor, whereas no increase was observed when cells were pretreated with 100 microM desferrioxamine, an avid iron chelator/redox inhibitor. Correspondingly, postirradiation formation of thiobarbituric acid-reactive material was markedly enhanced by ferric-8-hydroxyquinoline and suppressed by desferrioxamine, as was the extent of cell killing. When added to cells after a light dose, chain-breaking antioxidants such as butylated hydroxytoluene and alpha-tocopherol strongly protected against cell killing and slowed the increase in 7-OOH/5 alpha-OOH ratio. It is apparent from these results that (1) the 7-OOH/5 alpha-OOH or 7-OOH/6-OOH ratio can be used as a highly sensitive index of singlet oxygen vs. free radical dominance in photodynamically stressed cells; and (2) that postirradiation chain peroxidation plays an important role in photodynamically initiated cell killing.

Lipid hydroperoxide analysis by high-performance liquid chromatography with mercury cathode electrochemical detection.

In addition to the applications described, HPLC-EC(Hg) can be used for determining LOOHs in lipoproteins and for monitoring LOOH detoxification in cells. As it continues to be developed and refined, this approach should prove to be valuable not only for ultrasensitive determination of lipid-derived peroxides, but protein- and nucleic acid-derived peroxided as well.

Force-calcium relationship depends on myosin heavy chain and troponin isoforms in rat diaphragm muscle fibers.

The present study examined Ca(2+) sensitivity of diaphragm muscle (Dia(m)) fibers expressing different myosin heavy chain (MHC) isoforms. We hypothesized that Dia(m) fibers expressing the MHC(slow) isoform have greater Ca(2+) sensitivity than fibers expressing fast MHC isoforms and that this fiber-type difference in Ca(2+) sensitivity reflects the isoform composition of the troponin (Tn) complex (TnC, TnT, and TnI). Studies were performed in single Triton-X-permeabilized Dia(m) fibers. The Ca(2+) concentration at which 50% maximal force was generated (pCa(50)) was determined for each fiber. SDS-PAGE and Western analyses were used to determine the MHC and Tn isoform composition of single fibers. The pCa(50) for Dia(m) fibers expressing MHC(slow) was significantly greater than that of fibers expressing fast MHC isoforms, and this greater Ca(2+) sensitivity was associated with expression of slow isoforms of the Tn complex. However, some Dia(m) fibers expressing MHC(slow) contained the fast TnC isoform. These results suggest that the combination of TnT, TnI, and TnC isoforms may determine Ca(2+) sensitivity in Dia(m) fibers.

Effect of unilateral denervation on maximum specific force in rat diaphragm muscle fibers.

We hypothesize that 1) the effect of denervation (DNV) is more pronounced in fibers expressing fast myosin heavy chain (MHC) isoforms and 2) the effect of DNV on maximum specific force reflects a reduction in MHC content per half sarcomere or the number of cross bridges in parallel. Studies were performed on single Triton X-100-permeabilized fibers activated at a pCa (-log Ca2+ concentration) of 4.0. MHC content per half sarcomere was determined by densitometric analysis of SDS-PAGE gels and comparison to a standard curve of known MHC concentrations. After 2 of wk DNV, the maximum specific force of fibers expressing MHC2X was reduced by approximately 40% (MHC(2B) expression was absent), whereas the maximum specific force of fibers expressing MHC2A and MHC(slow) decreased by only approximately 20%. DNV also reduced the MHC content in fibers expressing MHC2X, with no effect on fibers expressing MHC2A and MHC(slow). When normalized for MHC content per half sarcomere, force generated by DNV fibers expressing MHC2X and MHC2A was decreased compared with control fibers. These results suggest the force per cross bridge is also affected by DNV.

Nitric oxide impairs Ca2+ activation and slows cross-bridge cycling kinetics in skeletal muscle.

The effects of the nitric oxide (NO) donor spermine NONOate (Sp-NO, 1.0 mM) on cross-bridge recruitment and cross-bridge cycling kinetics were studied in permeabilized rabbit psoas muscle fibers. Fibers were activated at various Ca2+ concentrations (pCa, negative logarithm of Ca2+ concentration), and the pCa at which force was maximal (pCa 4.0) and approximately 50% of maximal (pCa50 5.6) were determined. Fiber stiffness was determined using 1-kHz sinusoidal length perturbations, and the fraction of cross bridges in the force-generating state was estimated by the ratio of stiffness during maximal (pCa 4.0) and submaximal (pCa 5.6) Ca2+ activation to stiffness during rigor (at pCa 4.0). Cross-bridge cycling kinetics were evaluated by measuring the rate constant for force redevelopment after quick release (by 15% of optimal fiber length, L(o)) and restretch of the fiber to L(o). Exposing fibers to Sp-NO for 10 min reduced force and the fraction of cross bridges in the force-generating state at maximal and submaximal (pCa50) Ca2+ activation. However, the effects of Sp-NO were more pronounced during submaximal Ca2+ activation. Sp-NO also reduced the rate constant for force redevelopment but only during submaximal Ca2+ activation. We conclude that Sp-NO reduces Ca2+ sensitivity by decreasing the number of cross bridges in the strongly bound state and also impairs cross-bridge cycling kinetics during submaximal activation.

Maximum specific force depends on myosin heavy chain content in rat diaphragm muscle fibers.

In the present study, myosin heavy chain (MHC) content per half sarcomere, an estimate of the number of cross bridges available for force generation, was determined in rat diaphragm muscle (Dia(m)) fibers expressing different MHC isoforms. We hypothesize that fiber-type differences in maximum specific force [force per cross-sectional area (CSA)] reflect the number of cross bridges present per CSA. Studies were performed on single, Triton X-100-permeabilized rat Dia(m) fibers. Maximum specific force was determined by activation of single Dia(m) fibers in the presence of a high-calcium solution (pCa, -log Ca(2+) concentration of 4.0). SDS-PAGE and Western blot analyses were used to determine MHC isoform composition and MHC content per half sarcomere. Differences in maximum specific force across fast MHC isoforms were eliminated when controlled for half-sarcomere MHC content. However, the force produced by slow fibers remained below that of fast fibers when normalized for the number of cross bridges available. On the basis of these results, the lower force produced by slow fibers may be due to less force per cross bridge compared with fast fibers.

Reserve capacity for ATP consumption during isometric contraction in human skeletal muscle fibers.

Maximum velocity of the actomyosin ATPase reaction (V(max) ATPase) and ATP consumption rate during maximum isometric activation (ATP(iso)) were determined in human vastus lateralis (VL) muscle fibers expressing different myosin heavy chain (MHC) isoforms. We hypothesized that the reserve capacity for ATP consumption [1 -- (ratio of ATP(iso) to V(max) ATPase)] varies across VL muscle fibers expressing different MHC isoforms. Biopsies were obtained from 12 subjects (10 men and 2 women; age 21--66 yr). A quantitative histochemical procedure was used to measure V(max) ATPase. In permeabilized fibers, ATP(iso) was measured using an NADH-linked fluorometric procedure. The reserve capacity for ATP consumption was lower for fibers coexpressing MHC(2X) and MHC(2A) compared with fibers singularly expressing MHC(2A) and MHC(slow) (39 vs. 52 and 56%, respectively). Tension cost (ratio of ATP(iso) to generated force) also varied with fiber type, being highest in fibers coexpressing MHC(2X) and MHC(2A). We conclude that fiber-type differences in the reserve capacity for ATP consumption and tension cost reflect functional differences such as susceptibility to fatigue.

Mechanisms underlying increased force generation by rat diaphragm muscle fibers during development.

It has been found that maximum specific force (F(max); force per cross-sectional area) of rat diaphragm muscle doubles from birth to 84 days (adult). We hypothesize that this developmental change in F(max) reflects an increase in myosin heavy chain (MHC) content per half-sarcomere (an estimate of the number of cross bridges in parallel) and/or a greater force per cross bridge in fibers expressing fast MHC isoforms compared with slow and neonatal MHC isoforms (MHC(slow) and MHC(neo), respectively). Single Triton 100-X-permeabilized fibers were activated at a pCa of 4.0. MHC isoform expression was determined by SDS-PAGE. MHC content per half-sarcomere was determined by densitometric analysis and comparison to a standard curve of known MHC concentrations. MHC content per half-sarcomere progressively increased during early postnatal development. When normalized for MHC content per half-sarcomere, fibers expressing MHC(slow) and coexpressing MHC(neo) produced less force than fibers expressing fast MHC isoforms. We conclude that lower force per cross bridge in fibers expressing MHC(slow) and MHC(neo) contributes to the lower F(max) seen in early postnatal development.