Effect of hypoxia exposure on the recovery of skeletal muscle phenotype during regeneration.

Hypoxia impairs the muscle fibre-type shift from fast-to-slow during post-natal development; however, this adaptation could be a consequence of the reduced voluntary physical activity associated with hypoxia exposure rather than the result of hypoxia per se. Moreover, muscle oxidative capacity could be reduced in hypoxia, particularly when hypoxia is combined with additional stress. Here, we used a model of muscle regeneration to mimic the fast-to-slow fibre-type conversion observed during post-natal development. We hypothesised that hypoxia would impair the recovery of the myosin heavy chain (MHC) profile and oxidative capacity during muscle regeneration. To test this hypothesis, the soleus muscle of female rats was injured by notexin and allowed to recover for 3, 7, 14 and 28 days under normoxia or hypobaric hypoxia (5,500 m altitude) conditions. Ambient hypoxia did not impair the recovery of the slow MHC profile during muscle regeneration. However, hypoxia moderately decreased the oxidative capacity (assessed from the activity of citrate synthase) of intact muscle and delayed its recovery in regenerated muscle. Hypoxia transiently increased in both regenerated and intact muscles the content of phosphorylated AMPK and Pgc-1α mRNA, two regulators involved in mitochondrial biogenesis, while it transiently increased in intact muscle the mRNA level of the mitophagic factor BNIP3. In conclusion, hypoxia does not act to impair the fast-to-slow MHC isoform transition during regeneration. Hypoxia alters the oxidative capacity of intact muscle and delays its recovery in regenerated muscle; however, this adaptation to hypoxia was independent of the studied regulators of mitochondrial turn-over.

Effect of hypoxia exposure on the phenotypic adaptation in remodelling skeletal muscle submitted to functional overload.

AIM: To determine whether hypoxia influences the phenotypic adaptation of skeletal muscle induced by mechanical overload. METHODS: Plantaris muscles of female rats were submitted to mechanical overload following synergist ablation. After 3 days of overload, rats were exposed to either hypobaric hypoxia (equivalent to 5500 m) or normoxia. Muscles were collected after 5, 12 and 56 days of overload (i.e. after 3, 9 and 53 days of hypoxia). We determined the myosin heavy chain (MHC) distribution, mRNA levels of myocyte-enriched calcineurin-integrating protein 1 (MCIP1) to indirectly assess calcineurin activity, the changes in oxidative capacity from the activities of citrate synthase (CS) and cytochrome c oxidase (COX), and the expression of regulators involved in mitochondrial biogenesis (Pgc-1α, NRF1 and Tfam) and degradation (BNIP-3). RESULTS: Hypoxia did not alter the fast-to-slow MHC shift and the increase in calcineurin activity induced by overload; it only transiently slowed down the overload-induced transition in MHC isoforms. Hypoxia similarly decreased CS and COX activities in overloaded and control muscles. Nuclear respiratory factor 1 (NRF1) and transcription factor A (Tfam) mRNA and BNIP-3 protein were not influenced by hypoxia in overloaded muscles, whereas Pgc-1α mRNA and protein contents did not correlate with changes in oxidative capacity. CONCLUSION: Hypoxia is not a critical stimulus to modulate the fast-to-slow MHC transition associated with overload. Thus, the impairment of the fast-to-slow fibre shift often observed during post-natal development in hypoxia could be explained by the lower voluntary locomotor activity associated with hypoxia. Hypoxia alters mitochondrial oxidative capacity, but this adaptive response is similar in overloaded and control muscles.

Atypical Toxoplasma gondii strain from a free-living jaguar (Panthera onca) in French Guiana.

Like domestic cats, wild felids are involved in the complete infective cycle of Toxoplasma gondii because they can host in their gastrointestinal tract sexually mature parasites and shed infective oocysts in their feces. We report, to our knowledge, the first isolation and molecular characterization of a T. gondii strain from the heart tissue of a free-living jaguar (Panthera onca) in French Guiana. Sequencing at six polymorphic markers indicated that the jaguar isolate had an atypical genotype, including an allele at TgM-A previously found only in isolates from South America, and an allele at GRA6, which was previously reported only in Californian sea otter isolates. These findings are consistent with the recent description of atypical T. gondii strains involved in severe toxoplasmoses in immunocompetent patients in French Guiana that seemed to be linked to a neotropical forest-based cycle involving wild cats and their prey.

Musclin gene expression is strongly related to fast-glycolytic phenotype.

Musclin has been described as a muscle-derived secretory peptide, responsive to insulin in vivo, and inducing insulin resistance in vitro. Because muscle fibers display very different metabolic properties and insulin sensitivity, we tested the hypothesis that musclin expression could depend on myofiber type. Musclin mRNA was detected at high level in fast gastrocnemius and plantaris muscles, but only as traces in soleus, a slow-twitch muscle. A single fiber analysis showed that musclin was produced by muscle fibers themselves, almost exclusively type IIb fibers. Slow to fast transition of soleus phenotype after hindlimb suspension increased musclin mRNA levels, whereas fast to slow transition of plantaris phenotype after functional overload decreased musclin mRNA levels. This clearly suggests that musclin transcription is strongly related to fast-glycolytic phenotype. We conclude that musclin is produced by myocytes in a highly fiber-type specific manner and that physiological changes in type IIb MHC lead to coordinated musclin expression.

Human umbilical vein endothelial cells increase ex vivo expansion of human CD34(+) PBPC through IL-6 secretion.

BACKGROUND: Ex vivo expansion of hematopoietic stem cells (HSC) can help reduce cytopenia following transplantation, especially in NHL patients whose BM is deficient because of extensive chemotherapy. We have previously reported that human umbilical vein endothelial cells (HUVEC) can contribute to improved PBPC expansion when used in co-culture with CD34(+) cells. METHODS: We evaluated the roles of direct HUVEC CD34(+) contact and HUVEC-produced soluble factors. We cultured CD34(+) PBPC harvested from NHL patients in four different conditions: (1) liquid culture without HUVEC; (2) co-culture in contact with HUVEC; (3) co-culture with HUVEC but without direct contact; (4) liquid culture with HUVEC-conditioned medium (CM). Thrombopoietin (Tpo), Flk2Flt3 ligand (FL) and c-kit ligand (KL) with or without rhIL-6 were added to these four culture conditions. RESULTS AND DISCUSSION: Our results showed that HUVEC co-culture or addition of HUVEC-CM to Tpo, FL and KL (TFK) improved CD34(+) PBPC expansion compared with liquid culture, as determined by total viable nucleated cells (TNC), colony-forming cell assay (CFC) and week-6 cobblestone area-forming cells (Wk-6 CAFC) expansions. Non-contact culture led to similar PBPC expansion as contact co-culture; moreover, HUVEC-CM improved PBPC expansion. However, when rhIL-6 was added to HUVEC-CM with TFK, no significant difference was observed. Finally, high quantities of IL-6 were detected in HUVEC-CM and addition of anti-IL-6 Ab inhibited the positive effect of HUVEC on PBPC expansion. Our results thus suggest that HUVEC may improve PBPC expansion, at least through IL-6 secretion.

Adaptive changes in cardiac myosin heavy chain and creatine kinase isozymic profiles in rats native of altitude.

AIM: The developmental changes in the myosin heavy chain (MHC) profile, creatine kinase (CK) and lactate dehydrogenase (LDH) activities and isozyme expression occurring in heart were examined in rats born and living at altitude (La Paz, Bolivia, 3700 m, H(LP)) for 16 generations. We hypothesized that H(LP) rats respond differently to hypoxia than rats born and living at sea level, and secondarily exposed to altitude during 3 weeks (H(3W)). METHODS: The cardiac expression of MHC, CK and LDH was studied in left (LV) and right ventricle (RV) of H(LP) animals 1, 2, 3, 4 and 18 weeks after birth, and compared with control normoxic (C groups) and H(3W) animals. RESULTS: Rats secondarily exposed to hypoxia showed a lower alpha-MHC content than C or H(LP) rats in both LV and RV, 3 weeks after birth (P < 0.05), consistent with a delay in the maturation of the heart contractile phenotype. A global increase in the total CK activity was observed in the LV of H(3W) animals in comparison with C rats (P < 0.05), while no change was reported in H(LP) animals. In both ventricles, M-LDH activity was higher in H(3W) than in H(LP) and C rats (P < 0.05). The relative amount of alpha-MHC decreased by 20% in RV of 18-week-old H(LP) and H(3W) rats in comparison with C animals, consistent with the hypoxia-induced ventricular enlargement (P < 0.01). An increased activity of the foetal B-CK subunit was observed in both LV and RV of H(3W) rats in comparison with H(LP) and C animals (P < 0.05). CONCLUSION: This study demonstrates that rats native and living at altitude for several generations present some features relevant to genetic selection to altitude.

Differential effects of thyroid hormones on energy metabolism of rat slow- and fast-twitch muscles.

Thyroid hormone (TH) is an important regulator of mitochondrial content and activity. As mitochondrial content and properties differ depending on muscle-type, we compared mitochondrial regulation and biogenesis by T3 in slow-twitch oxidative (soleus) and fast-twitch mixed muscle (plantaris). Male Wistar rats were treated for 21 to 27 days with T3 (200 microg/kg/day). Oxidative capacity, regulation of mitochondrial respiration by substrates and phosphate acceptors, and transcription factors were studied. In soleus, T3 treatment increased maximal oxygen consumption (Vmax) and the activities of citrate synthase (CS) and cytochrome oxidase (COX) by 100%, 45%, and 71%, respectively (P < 0.001), whereas in plantaris only Vmax increased, by 39% (P < 0.01). ADP-independent respiration rate was increased in soleus muscle by 216% suggesting mitochondrial uncoupling. Mitochondrial substrate utilization in soleus was also influenced by T3, as were mitochondrial enzymes. Lactate dehydrogenase (LDH) activity was elevated in soleus and plantaris by 63% and 11%, respectively (P < 0.01), and soleus creatine kinase was increased by 48% (P < 0.001). T3 increased the mRNA content of the transcriptional co-activator of mitochondrial genes, PGC-1alpha, and the I and IV COX subunits in soleus. The muscle specific response to thyroid hormones could be explained by a lower content of TH receptors in plantaris than soleus. Moreover, TRalpha mRNA level decreased further after T3 treatment. These results demonstrate that TH has a major effect on mitochondrial content, regulation and coupling in slow oxidative muscle, but to a lesser extent in fast muscle, due to the high expression of TH receptors and PGC-1alpha transcription factor.

Clenbuterol treatment affects myosin heavy chain isoforms and MyoD content similarly in intact and regenerated soleus muscles.

AIMS: Pharmacological treatment with the beta2-adrenoceptor agonist clenbuterol is known to induce a slow-to-fast fibre type and myosin heavy chain (MHC) isoform transition in intact muscle. This study examined the sensitivity of regenerated soleus muscle to 4 weeks of clenbuterol treatment (2 mg kg-1 day-1). METHODS: Female Wistar rats were divided into two groups: vehicle treated (n = 8) and clenbuterol treated (n = 8). The clenbuterol effects on MHC and MyoD expression were examined in soleus muscles either intact, or previously degenerated by venom of the Notechis scutatus scutatus snake. RESULTS: Post-treatment body weights and skeletal muscle weights were not affected by clenbuterol treatment. Muscle protein concentration was higher, and body fat lower in clenbuterol-treated rats than in vehicle-treated animals (P < 0.05). Polyacrylamide gel electrophoresis of soleus myofibrillar protein indicated a clenbuterol-induced decrease in the relative percentage of type I MHC with a concomitant increase in type IIa MHC (31%, P < 0.001). No degeneration effect was observed after 28 days of recovery on the MHC isoform content, and regenerated soleus muscles exhibited the same phenotypical profile as intact soleus muscles, whether or not they were treated with clenbuterol. In intact and in regenerated soleus muscles, MyoD protein levels were significantly increased by clenbuterol treatment (90 and 77%, respectively, P < 0.001). CONCLUSION: These results show that regenerated soleus muscles, comprising a homogeneous population of fibres deriving from satellite cells, have a similar response to clenbuterol as intact muscle arising from at least two discrete populations of myotubes; it is suggested that the activity of signalling pathways involved in the effects of clenbuterol on MHC transitions is not related to the developmental history of myofibres.

Recovery of contractile and metabolic phenotypes in regenerating slow muscle after notexin-induced or crush injury.

The recovery of metabolic pathways after muscle damage has been poorly studied. We investigated the myosin heavy chain (MHC) isoform transitions and the recovery of citrate synthase (CS) activity, isoform distribution of lactate dehydrogenase (LDH) and creatine kinase (CK) in slow muscles after two types of injury. Muscle degeneration was induced in left soleus muscles of male Wistar rats by either notexin injection or crushing and the regenerative process was examined from 2 to 56 days after injury. Myosin transition occurred earlier after notexin than after crush injury. Fast-type IIx and more particularly type IIa MHC isoform disappeared by day 28 after notexin inoculation, while they were still detected long after in crushed muscles. A full recovery of both the CS activity and the specific activity of the H-LDH subunit was observed from day 42 in notexin-treated muscles, while values measured in crushed muscles remained significantly lower than in non-injured muscles (P < 0.05). The activity of the mitochondrial isoform of CK (mi-CK) was markedly affected by the type of injury (P < 0.001), and failed to reach normal levels after crush injury (P < 0.05). The results of this study show that the relatively rapid MHC transitions during regeneration contrasts with the slow recovery in the oxidative capacity. The recovery of the oxidative capacity remained incomplete after crush injury, a model of injury known to lead to disruption of the basal lamina and severe interruption of the vascular and nerve supply.

Physical activity changes the regulation of mitochondrial respiration in human skeletal muscle.

This study explores the importance of creatine kinase (CK) in the regulation of muscle mitochondrial respiration in human subjects depending on their level of physical activity. Volunteers were classified as sedentary, active or athletic according to the total activity index as determined by the Baecke questionnaire in combination with maximal oxygen uptake values (peak V(O2), expressed in ml min(-1) kg(-1)). All volunteers underwent a cyclo-ergometric incremental exercise test to estimate their peak V(O2) and V(O2) at the ventilatory threshold (VT). Muscle biopsy samples were taken from the vastus lateralis and mitochondrial respiration was evaluated in an oxygraph cell on saponin permeabilised muscle fibres in the absence (V(0)) or in the presence (V(max)) of saturating [ADP]. While V(0) was similar, V(max) differed among groups (sedentary, 3.7 +/- 0.3, active, 5.9 +/- 0.9 and athletic, 7.9 +/- 0.5 micromol O2 min(-1) (g dry weight)(-1)). V(max) was correlated with peak V(O2) (P < 0.01, r = 0.63) and with V(T) (P < 0.01, r = 0.57). There was a significantly greater degree of coupling between oxidation and phosphorylation (V(max)/V(0)) in the athletic individuals. The mitochondrial K(m) for ADP was significantly higher in athletic subjects (P < 0.01). Mitochondrial CK (mi-CK) activation by addition of creatine induced a marked decrease in K(m) in athletic individuals only, indicative of an efficient coupling of mi-CK to ADP rephosphorylation in the athletic subjects only. It is suggested that increasing aerobic performance requires an enhancement of both muscle oxidative capacity and mechanisms of respiratory control, attesting to the importance of temporal co-ordination of energy fluxes by CK for higher efficacy.

Nandrolone decanoate treatment induces changes in contractile responses of rat untrained fast-twitch skeletal muscle.

This investigation was designed to examine whether short-term administration of anabolic-androgenic steroids (AAS) (nandrolone decanoate) could produce changes in contractile responses of untrained rat fast- (edl) and slow- (soleus) twitch skeletal muscle. Twenty male rats were divided into two groups, one group received weekly (for 6 weeks) an intramuscular injection of AAS, nandrolone decanoate (15 mg kg(-1)) and the second group received weekly the similar doses of vehicle (sterile peanut oil). In edl intact isolated small bundles (two to four cells), it was found that nandrolone decanoate treatment increases the K+ contracture tension (146 mM) relative to maximum tension by 56%, whereas no change was observed in the time to peak tension and in the time constant of relaxation. By contrast, in treated soleus muscle, compared with control, no significant modification was found in the K+ contracture characteristics. The change in edl contractile responses was associated with a shift to more negative potential of the voltage-dependence activation and the steady-state inactivation curves which also shifted leftward in treated soleus fibres. Furthermore, in edl skinned Triton X-100 fibres, the Ca2+ sensitivity of contractile proteins (pCa50) was increased, while electrophoresis analysis indicates no significant effect of nandrolone decanoate treatment on myosin heavy chain (MHC) isoforms. The present results show that nandrolone decanoate treatment produces more pronounced changes in untrained fast muscle function rather than soleus by acting at different levels of the excitation-contraction coupling mechanism without changes in the MHC isoforms and that contractile responses became similar to those found in soleus muscle.

Influence of overload on phenotypic remodeling in regenerated skeletal muscle.

We studied the effects of 10 wk of functional overload on the expression of myosin heavy chain (MHC), sarcoplasmic reticulum Ca(2+)-ATPase isoforms (SERCA), and the activity of several metabolic enzymes in sham and regenerated plantaris muscles. Overload was accomplished by bilateral surgical ablation of its synergists 4 wk after right plantaris muscles regenerated after myotoxic infiltration. The overload-induced muscle enlargement was slightly less in regenerated than in sham muscles [28% (P < 0.005) and 43% (P < 0.001), respectively]. Overload led to an increase in type I MHC expression (P < 0.01) to a similar extent in sham and regenerated plantaris, while the expected shift from type IIb to type IIa MHC was less marked in regenerated than in sham plantaris. The overload-induced decrease in the expression of the fast SERCA isoform and in the activity of the M subunit of lactate dehydrogenase occurred to a similar extent in sham and regenerated plantaris [66% (P < 0.01) and 27% (P < 0.005), respectively]. In conclusion, the lesser responses of muscle mass and fast MHC composition of regenerated plantaris to mechanical overload suggest an alteration of the transcriptional, translational, and/or posttranslational control of gene expression in regenerated muscle.

Response of mitochondrial function to hypothyroidism in normal and regenerated rat skeletal muscle.

Although thyroid hormones induce a well known decrease in muscle oxidative capacity, nothing is known concerning their effects on mitochondrial function and regulation in situ. Similarly, the influence of regeneration process is not completely understood. We investigated the effects of hypothyroidism on mitochondrial function in fast gastrocnemius (GS) and slow soleus (SOL) muscles either intact or having undergone a cycle of degeneration/regeneration (Rg SOL) following a local injection of myotoxin. Thyroid hormone deficiency was induced by thyroidectomy and propylthiouracyl via drinking water. Respiration was measured in muscle fibres permeabilised by saponin in order to assess the oxidative capacity of the muscles and the regulation of mitochondria in situ. Oxidative capacities were 8.9 in SOL, 8.5 in Rg SOL and 5.9 micromol O2/min/g dry weight in GS and decreased by 52, 42 and 39% respectively (P < 0.001) in hypothyroid rats. Moreover, the Km of mitochondrial respiration for the phosphate acceptor ADP exhibited a two-fold decrease in Rg SOL and intact SOL by hypothyroidism (P < 0.01), while mitochondrial creatine kinase activity and sensitivity of mitochondrial respiration to creatine were not altered. The results of this study demonstrate that hypothyroidism markedly altered the sensitivity of mitochondrial respiration to ADP but not to creatine in SOL muscles, suggesting that mitochondrial regulation could be partially controlled by thyroid hormones. On the other hand, mitochondrial function completely recovered following regeneration/degeneration, suggesting that thyroid hormones are not involved in the regeneration process per se.

Diet restriction plays an important role in the alterations of heart mitochondrial function following exposure of young rats to chronic hypoxia.

Male Wistar rats aged 4 weeks, were subjected to hypobaric hypoxia (barometric pressure 505 hPa, PI,O2 106 hPa) or to diet restriction (reproducing the effect of hypoxia-induced anorexia) for 4 weeks. Each group (control, hypoxic, pair-fed, n = 16), was divided into two sub-groups housed individually in either normal cages or cages with running wheels allowing evaluation of voluntary activity (n = 8 each). The skinned-fibre technique was used to evaluate the functional properties of myofibrillar mitochondria from right and left ventricles in situ. The oxidative fibres from the soleus and diaphragm muscles were also investigated for comparison. Analysis of variance did not detect any significant effect of voluntary running activity. With calorie restriction, the maximal respiratory rate (Vmax) in the presence of 1 mM adenosine 5'-diphosphate (ADP) in myocardial fibres fell significantly (by about 25%) but was unchanged in skeletal myocytes. Following hypoxia, Vmax in myocardial fibres increased by 25% compared with the calorie restricted group and in soleus and diaphragm muscle fibres by about 30% compared with control. In myocardial fibres of control rats, creatine (20 mM) increased the sub-maximal respiratory rate by 80% in the presence of 0.1 mM ADP. Under calorie restriction or hypoxia the stimulatory effect was significantly reduced to 34-56%. This alteration was due to a decrease in the apparent Michaelis-Menten constant (Km) of mitochondrial respiration for ADP evaluated in the absence of creatine, while the Km in presence of creatine 20 mM was unchanged. In conclusion, reduced food intake decreased the oxidative capacity (Vmax) and the apparent Km for ADP of mitochondria in both left and right ventricles. Chronic hypoxia per se was responsible for an increase in the oxidative capacity of all oxidative muscles but did not exert significant effects on the control of respiration by ADP and creatine in myocardium.

Dual influence of disease and increased load on diaphragm muscle in heart failure.

We have recently shown that mitochondrial function and energy metabolism are altered in the myocardium as well as in slow and fast locomotor muscles of rats subjected to prolonged congestive heart failure (CHF) suggesting a generalized metabolic myopathy in heart failure. Here, we investigate whether the diaphragm of CHF animals, which experiences both increased work and the general systemic influence of heart failure, will also be susceptible to altered energy metabolism. Biopsies were obtained from the costal diaphragm of failing rats 8 months after aortic banding. A marked increase in type I and type IIa myosin heavy chains at the expense of types IIx and IIb, suggests an adaptation towards a slower phenotype. Glycolytic enzymes decreased in CHF diaphragm with an increase in the H:M lactate dehydrogenase isoenzyme ratio. These results suggest a reorientation of the diaphragm muscle towards a slow, fatigue-resistant phenotype. However, maximal oxidative capacity assessed in saponin-permeabilized fibers in the presence of ADP was considerably reduced in CHF diaphragm (7.7+/-0.4 v 11.8+/-0.7 micromol O2/min/g dry weight in sham P<0.001), suggesting an alteration in oxidative phosphorylation. Furthermore, ADP sensitivity of CHF mitochondria was significantly increased (apparent Km for ADP 308+/-21 v 945+/-106 microM in sham P<0.001), whereas sensitivity to ADP in the presence of creatine was comparable (Km 79+/-12 v 90+/-11 microM in sham). In heart failure, therefore, the diaphragm muscle seems to adapt towards a more slow and economical contraction as a result of increased workload, but this adaptation is limited by the disease-induced altered mitochondrial function.

Lack of coordinated changes in metabolic enzymes and myosin heavy chain isoforms in regenerated muscles of trained rats.

We investigated training-induced changes in biochemical properties and myosin heavy chain (MHC) composition of regenerated (cardiotoxin-injected) plantaris muscles (PLA) in rats either maintained sedentary (S, n = 9) or endurance trained on a treadmill over a 8-week period (T, n = 7). Both endurance training and regeneration altered the pattern of fast MHC expression. An analysis of the two-way interaction between training and regeneration showed that the relative content of type IIa MHC was affected (P < 0.05). The 140% increase in type IIa MHC observed in regenerated PLA from T rats compared with nontreated muscle of S rats, exceeded the 102% increase resulting from the combination of regeneration alone (26%) and training alone (61%). A similar interaction between training and regeneration was shown for the percentage of fibres expressing either type IIa or type lIb MHC (P < 0.05). In contrast, a significant increase in the citrate synthase (CS) activity was shown in PLA as a result of endurance training, without specific effect of regeneration. Furthermore, training-induced changes in CK and LDH isoenzyme distribution occurred to a similar extent in regenerated and non-treated PLA muscles, and thus did not follow the changes in MHC isoforms. An increase in the mitochondrial CK isozyme activity (mi-CK) was shown in both non-treated and previously degenerated PLA muscles (123 and 117%, P < 0.01, respectively), without specific effect of regeneration. The ratio of mi-CK to CS activity, an estimate of the mitochondrial specific activity of mi-CK was significantly increased by training (P < 0.02) and decreased by regeneration (P < 0.05). Taken together, these data suggest that while training and regeneration have cumulative effects on the pattern of fast MHC expression, the training-induced changes in the energy metabolism shown in mature non-treated myofibres are similar to those observed in regenerated fibres.

Myosin heavy chain composition of skeletal muscles in young rats growing under hypobaric hypoxia conditions.

This study investigated the effects of voluntary wheel running on the myosin heavy chain (MHC) composition of the soleus (Sol) and plantaris muscles (Pla) in rats developing under hypobaric choronic hypoxia (CH) conditions during 4 wk in comparison with those of control rats maintained under local barometric pressure conditions (C) or rats pair-fed an equivalent quantity of food to that consumed by CH animals (PF). Compared with C animals, sedentary rats subjected to CH conditions showed a significant decrease in type I MHC in Sol (-12%, P < 0.01). Although strongly decreased under hypoxia, spontaneous running activity increased the expression of type I MHC (P < 0.01) so that no difference in the MHC profile of Sol was shown between CH active and C active rats. The MHC distribution in Sol of PF rats was not significantly different from that found in C animals. CH resulted in a significant decrease in type I (P < 0.01) and type IIA (P < 0.005) MHC, concomitant with an increase in type IIB MHC in Pla (P < 0.001), compared with C and PF animals. In contrast to results in Sol muscle, this slow-to-fast shift in the MHC profile was unaffected by spontaneous running activity. These results suggest that running exercise suppresses the hypoxia-induced slow-to-fast transition in the MHC expression in Sol muscles only. The hypoxia-induced decrease in food intake has no major influence on MHC expression in developing rats.

Subcellular creatine kinase alterations. Implications in heart failure.

We have tested the hypothesis that decreased functioning of creatine kinase (CK) at sites of energy production and utilization may contribute to alterations in energy fluxes and calcium homeostasis in congestive heart failure (CHF). Heart failure was induced by aortic banding in 3-week-old rats. Myofilaments, sarcoplasmic reticulum (SR), mitochondrial functions, and CK compartmentation were studied in situ using selective membrane permeabilization of left ventricular fibers with detergents (saponin for mitochondria and SR and Triton X-100 for myofibrils). Seven months after surgery, animals were in CHF. A decrease in total CK activity could be accounted for by a 4-fold decrease in activity and content (Western blots) of mitochondrial CK and a 30% decrease in M isoform of CK (MM-CK) activity. In myofibrils, maximal force, crossbridge kinetics, and alpha-myosin heavy-chain expression decreased, whereas calcium sensitivity of tension development remained unaltered. Myofibrillar CK efficacy was unchanged. Calcium uptake capacities of SR were estimated from the surface of caffeine-induced tension transient (SCa) after loading with different substrates. In CHF, SCa decreased by 23%, and phosphocreatine was 2 times less efficient in enhancing calcium uptake. Oxidative capacities of the failing myocardium measured as oxygen consumption per gram of fiber dry weight decreased by 28%. Moreover, the control of respiration by creatine, ADP, and AMP was severely impaired. Our observations provide evidence that alterations in CK compartmentation may contribute to alterations of energy fluxes and calcium homeostasis in CHF.

Effects of hindlimb suspension and androgen treatment on testosterone receptors in rat skeletal muscles.

This study tested the specific and combined effects of testosterone treatment and hindlimb suspension (HS) on the properties of steroid receptors in skeletal muscle. Male rats were either administered weekly high doses of testosterone heptylate (10 mg x kg(-1)) or olive oil placebo, and were either tail-suspended or acted as controls. After 3 weeks of treatment, three muscles were excised from each animal, soleus (SOL), extensor digitorum longus (EDL), and plantaris. The results showed that the testosterone treatment was unable to minimise the HS-induced atrophy of skeletal muscle. As expected, HS altered the fibre-type composition of SOL muscles (-33% of type I, +188% and +161% of type IIa and intermediate fibres respectively, P < 0.01). No overall effect of treatment was detected on the fibre-type composition of either slow or fast-twitch muscles. Binding capacity determined by a radiocompetition technique was increased by HS, especially in SOL and EDL muscles (P < 0.01), while HS or steroid treatment decreased the affinity of the steroid receptors. The combination of HS and testosterone administration resulted in a decrease in binding capacity and affinity of steroid receptors in skeletal muscles. Steroid receptors in fast-twitch muscles exhibited a higher affinity than those in slow-twitch muscles, and it is suggested that it is likely that testosterone treatment is more effective in fast-twitch than in slow-twitch muscles. It was concluded that the lack of preventive effect of testosterone treatment on HS-induced SOL muscle atrophy could be explained by both a decrease in steroid sensitivity and the removal of mechanical factors.

Changes in myosin heavy chain profile of mature regenerated muscle with endurance training in rat.

The objective of the present study was to examine the response of fast-twitch muscle to endurance training long after the muscle had regenerated from toxin injury. Seventeen male Wistar rats were randomly assigned to a sedentary (S, n = 10) or a trained group (T, n = 7). Endurance training by treadmill running (5 days week(-1), 30 m min(-1), 7% grade, 2 h day(-1) for 5 weeks) was initiated 5 weeks after myofibre degeneration was induced in the right extensor digitorum longus muscle (EDL) by two injections of 0.2 mL of the unfractionated venom from Naja nigricollis snake. Gel electrophoresis analyses showed that training alone resulted in a 140% increase in type IIX myosin heavy chain (MHC) (P < 0.01) and a slight decrease in type IIB MHC (-14% P < 0.05). Regeneration alone induced an increase in both type IIA and IIX MHC expression (103%, P < 0.05, and 131%, P < 0.01, respectively), and a concomitant decrease in the percentage of type IIB MHC (P < 0.05). The shift from type IIB toward type IIA MHC composition observed in regenerated muscles of T rats resulted not only from an additive, but from a cumulative effect of training and regeneration. Immunohistochemical analysis of MHC content in individual fibres showed similar changes. These data suggest that the impact of endurance training on fast-type MHCs was more marked in mature regenerated muscles than in regenerating ones, and provide evidence of the heightened plasticity of fully regenerated muscles to repeated exercise.