Soleus muscle stability in wild hibernating black bears.

Based on studies of fast skeletal muscles, hibernating black and brown bears resist skeletal muscle atrophy during months of reduced physical activity and not feeding. The present study examined atrophy sparing in the slow soleus muscle, known to be highly prone to disuse atrophy in humans and other mammals. We demonstrated histochemically that the black bear soleus is rich in slow fibers, averaging 84.0 ± 6.6%. The percentages of slow fibers in fall (87.3 ± 4.9%) and during hibernation (87.1 ± 5.6%) did not differ ( P = 0.3152) from summer. The average fiber cross-sectional area to body mass ratio (48.6 ± 11.7 µm2/kg) in winter hibernating bears was not significantly different from that of summer (54.1 ± 11.8 µm2/kg, P = 0.4186) and fall (47.0 ± 9.7 µm2/kg, P = 0.9410) animals. The percentage of single hybrid fibers containing both slow and fast myosin heavy chains, detected biochemically, increased from 2.6 ± 3.8% in summer to 24.4 ± 24.4% ( P = 0.0244) during hibernation. The shortening velocities of individual hybrid fibers remained unchanged from that of pure slow and fast fibers, indicating low content of the minority myosins. Slow and fast fibers in winter bears exhibited elevated specific tension (kN/m2; 22%, P = 0.0161 and 11%, P = 0.0404, respectively) and maintained normalized power. The relative stability of fiber type percentage and size, fiber size-to-body mass ratio, myosin heavy chain isoform content, shortening velocity, power output, and elevated specific tension during hibernation validates the ability of the black bear to preserve the biochemical and performance characteristics of the soleus muscle during prolonged hibernation.

Protein composition of endurance trained human skeletal muscle.

Evidence suggests that myofibers from endurance trained skeletal muscle display unique contractile parameters. However, the underlying mechanisms remain unclear. To further elucidate the influence of endurance training on myofiber contractile function, we examined factors that may impact myofilament interactions (i. e., water content, concentration of specific protein fractions, actin and myosin content) or directly modulate myosin heavy chain (MHC) function (i. e., myosin light chain (MLC) composition) in muscle biopsy samples from highly-trained competitive (RUN) and recreational (REC) runners. Muscle water content was lower (P<0.05) in RUN (73±1%) compared to REC (75±1%) and total muscle and myofibrillar protein concentration was higher (P<0.05) in RUN, which may indicate differences in myofilament spacing. Content of the primary contractile proteins, myosin (0.99±0.08 and 1.01±0.07 AU) and actin (1.33±0.09 and 1.27±0.09 AU) in addition to the myosin to actin ratio (0.75±0.04 and 0.80±0.06 AU) was not different between REC and RUN, respectively, when expressed relative to the amount of myofibrillar protein. At the single-fiber level, slow-twitch MHC I myofibers from RUN contained less (P<0.05) MLC 1 and greater (P<0.05) amounts of MLC 3 than REC, while MLC composition was similar in fast-twitch MHC IIa myofibers between REC and RUN. These data suggest that the distinctive myofiber contractile profile in highly-trained runners may be partially explained by differences in the content of the primary contractile proteins and provides unique insight into the modulation of contractile function with extreme loading -patterns.

Effects of prolonged space flight on human skeletal muscle enzyme and substrate profiles.

Our primary goal was to determine the effects of 6-mo flight on the International Space Station (ISS) on selected anaerobic and aerobic enzymes, and the content of glycogen and lipids in slow and fast fibers of the soleus and gastrocnemius. Following local anesthesia, biopsies were obtained from nine ISS crew members ∼45 days preflight and on landing day (R+0) postflight. We subdivided the crew into those who ran 200 min/wk or more (high treadmill, HT) in-flight from those who ran <100 min/wk (low treadmill, LT). In the LT group, there was a loss of lipid in soleus type I fibers, and muscle glycogen significantly increased in soleus fiber types postflight. Soleus cytochrome oxidase (CO) activity was significantly depressed postflight in the type I fiber. This was attributed to the LT group where CO activity was reduced 59%. Otherwise, there was no change in the crew mean for type I or IIa fiber glycolytic or mitochondrial enzyme activities pre- vs. postflight in either muscle. However, two of the three HT subjects (Subjects E and H) showed significant increases in both ß-hydroxyacyl-CoA dehydrogenase and citrate synthase in the soleus type I fibers, and Subject E, exhibiting the largest increase in soleus oxidative enzymes, was the only subject to show a significant decrease in glycolytic enzyme activity. It is apparent that crew members performing adequate treadmill running can maintain calf muscle enzymes, which suggests that increased fatigue with weightlessness cannot be directly caused by a decline in muscle enzyme capacity.

Prostaglandin E2 induces transcription of skeletal muscle mass regulators interleukin-6 and muscle RING finger-1 in humans.

Cyclooxygenase (COX) inhibiting drugs augment muscle mass and strength improvements during resistance exercise based treatment of sarcopenia in older individuals. Initial evidence suggests a potential mechanism of COX inhibitor blunted prostaglandin (PG) E2 stimulation of interleukin (IL)-6 and the ubiquitin ligase MuRF-1, reducing their inhibition on muscle growth. The purpose of this investigation was to determine if PGE2 stimulates IL-6 and MuRF-1 transcription in skeletal muscle. Muscle biopsies were obtained from 10 young individuals and incubated ex vivo with PGE2 or control and analyzed for IL-6 and MuRF-1 mRNA levels. PGE2 upregulated (P<0.05) expression of both IL-6 (195%) and MuRF-1 (51%). A significant relationship was found between IL-6 and MuRF-1 expression after incubation with PGE2 (r=0.77, P<0.05), suggesting regulation through a common pathway. PGE2 induces IL-6 and MuRF-1 transcription in human skeletal muscle, providing a mechanistic link between COX inhibiting drugs, PGE2, and the regulation of muscle mass.

Influence of aerobic cycle exercise training on patellar tendon cross-sectional area in older women.

Nine to 12 weeks of resistance exercise training in young individuals induces quadriceps muscle (∼6%) and region-specific patellar tendon (4-6%) hypertrophy. However, 12 weeks of resistance exercise training (∼1 h total exercise time) in older individuals (60-78 years) induces quadriceps muscle hypertrophy (9%) without impacting patellar tendon size. The current study examined if a different loading paradigm using cycle exercise would promote patellar tendon hypertrophy or alter the internal tendon properties, measured with magnetic resonance imaging signal intensity, in older individuals. Nine women (70 ± 2 years) completed 12 weeks of aerobic upright cycle exercise training (∼28 h total exercise time). Aerobic exercise training increased (P < 0.05) quadriceps muscle size (11 ± 2%) and VO2max (30 ± 9%). Mean patellar tendon cross-sectional area (CSA) (2 ± 1%) and signal intensity (-1 ± 2%) were unchanged (P > 0.05) over the 12 weeks of training. Region-specific CSA was unchanged (P > 0.05) at the proximal (-1 ± 3%) and mid regions (2 ± 2%) of the tendon but tended (P = 0.069) to increase at the distal region (5 ± 3%). Region-specific signal intensity differed along the tendon but was unchanged (P > 0.05) with training. Although more studies are needed, exercise-induced patellar tendon hypertrophy, compared with skeletal muscle, appears to be attenuated in older individuals, while the loading pattern associated with aerobic exercise seems to have more impact than resistance exercise in promoting patellar tendon hypertrophy.

Skeletal muscle plasticity with marathon training in novice runners.

The purpose of this study was to investigate leg muscle adaptation in runners preparing for their first marathon. Soleus and vastus lateralis (VL) biopsies were obtained from six recreational runners (23 ± 1 years, 61 ± 3 kg) before (T1), after 13 weeks of run training (T2), and after 3 weeks of taper and marathon (T3). Single muscle fiber size, contractile function (strength, speed, and power) and oxidative enzyme activity [citrate synthase (CS)] were measured at all three time points, and fiber type distribution was determined before and after the 16-week intervention. Training increased VO(2max) ∼9% (P<0.05). All soleus parameters were unchanged. VL MHC I fiber diameter increased (+8%; P<0.05) from T1 to T2. VL MHC I V(o) (-12%), MHC I power (-22%) and MHC IIa power (-29%) were reduced from T1 to T2 (P<0.05). No changes in VL single fiber contractile properties were observed from T2 to T3. No change was observed in soleus CS activity, whereas VL CS activity increased 66% (P<0.05). Our observations indicate that modest marathon training elicits very specific skeletal muscle adaptations that likely support the ability to perform 42.2 km of continuous running - further strengthening the existing body of evidence for skeletal muscle specificity.

Prolonged space flight-induced alterations in the structure and function of human skeletal muscle fibres.

The primary goal of this study was to determine the effects of prolonged space flight (180 days) on the structure and function of slow and fast fibres in human skeletal muscle. Biopsies were obtained from the gastrocnemius and soleus muscles of nine International Space Station crew members 45 days pre- and on landing day (R+0) post-flight. The main findings were that prolonged weightlessness produced substantial loss of fibre mass, force and power with the hierarchy of the effects being soleus type I > soleus type II > gastrocnemius type I > gastrocnemius type II. Structurally, the quantitatively most important adaptation was fibre atrophy, which averaged 20% in the soleus type I fibres (98 to 79 µm diameter). Atrophy was the main contributor to the loss of peak force (P(0)), which for the soleus type I fibre declined 35% from 0.86 to 0.56 mN. The percentage decrease in fibre diameter was correlated with the initial pre-flight fibre size (r = 0.87), inversely with the amount of treadmill running (r = 0.68), and was associated with an increase in thin filament density (r = 0.92). The latter correlated with reduced maximal velocity (V(0)) (r = 0.51), and is likely to have contributed to the 21 and 18% decline in V(0) in the soleus and gastrocnemius type I fibres. Peak power was depressed in all fibre types with the greatest loss (55%) in the soleus. An obvious conclusion is that the exercise countermeasures employed were incapable of providing the high intensity needed to adequately protect fibre and muscle mass, and that the crew's ability to perform strenuous exercise might be seriously compromised. Our results highlight the need to study new exercise programmes on the ISS that employ high resistance and contractions over a wide range of motion to mimic the range occurring in Earth's 1 g environment.

A new method to study in vivo protein synthesis in slow- and fast-twitch muscle fibers and initial measurements in humans.

The aim of this study was to develop an approach to directly assess protein fractional synthesis rate (FSR) in isolated human muscle fibers in a fiber type-specific fashion. Individual muscle fibers were isolated from biopsies of the vastus lateralis (VL) and soleus (SOL) obtained from eight young men during a primed, continuous infusion of [5,5,5-(2)H3]leucine performed under basal conditions. To determine mixed protein FSR, a portion of each fiber was used to identify fiber type, fibers of the same type were pooled, and the [5,5,5-(2)H3]leucine enrichment was determined via GC-MS. Processing isolated slow-twitch [myosin heavy chain (MHC) I] and fast-twitch (MHC IIa) fibers for mixed protein bound [5,5,5-(2)H3]leucine enrichment yielded mass ion chromatographic peaks that were similar in shape, abundance, and measurement reliability as tissue homogenates. In the VL, MHC I fibers exhibited a 33% faster (P<0.05) mixed protein FSR compared with MHC IIa fibers (0.068+/-0.006 vs. 0.051+/-0.003%/h). MHC I fibers from the SOL (0.060+/-0.005%/h) and MHC I fibers from the VL displayed similar (P>0.05) mixed protein FSR. Feasibility of processing isolated human muscle fibers for analysis of myofibrillar protein [5,5,5-(2)H3]leucine enrichment was also confirmed in non-fiber-typed pooled fibers from the VL. These methods can be applied to the study of fiber type-specific responses in human skeletal muscle. The need for this level of investigation is underscored by the different contributions of each fiber type to whole muscle function and the numerous distinct adaptive functional and metabolic changes in MHC I and MHC II fibers originating from the same muscle.

Resistance exercise reduces muscular substrates in women.

The purpose of this investigation was to examine the influence of an acute bout of resistance exercise (RE) on intramuscular triglyceride (IMTG) and muscle glycogen concentrations and intracellular signaling in women with high body fat content. Six overweight women with a high percent body fat (age 29+/-3 yr; BMI 28+/-3 kg/m(2), body fat 38+/-4%) performed 6 sets of 10 repetitions of knee extension exercise at 70% 1RM. Muscle biopsies were obtained from the vastus lateralis before, 1 min after (POST1), and 2 h after (POST2) exercise. Acute RE reduced (p<0.05) IMTG content approximately 40% at POST1 and POST2 (75+/-5; 45+/-6; 50+/-10 mmol/kg/dry wt). Muscle glycogen was also reduced (p<0.05) approximately 25% at POST1 and remained lower at POST2 (317+/-14; 241+/-30; 235+/-26 mmol/kg/dry wt). ERK1/2, SAPK/JNK, and p38 phosphorylation were increased (p<0.05) approximately 2-3-fold at POST1 and ERK1/2 remained elevated and POST2 whereas SAPK/JNK and p38 returned to basal levels. AMPKalpha phosphorylation was unchanged in response to RE. These results show that both IMTG and muscle glycogen stores serve as an important energy source during RE in overweight women and the MAP kinase signaling response to RE is not compromised by high levels of body fat.

Influence of concurrent exercise or nutrition countermeasures on thigh and calf muscle size and function during 60 days of bed rest in women.

AIM: The goal of this investigation was to test specific exercise and nutrition countermeasures to lower limb skeletal muscle volume and strength losses during 60 days of simulated weightlessness (6 degrees head-down-tilt bed rest). METHODS: Twenty-four women underwent bed rest only (BR, n = 8), bed rest and a concurrent exercise training countermeasure (thigh and calf resistance training and aerobic treadmill training; BRE, n = 8), or bed rest and a nutrition countermeasure (a leucine-enriched high protein diet; BRN, n = 8). RESULTS: Thigh (quadriceps femoris) muscle volume was decreased (P < 0.05) in BR (-21 +/- 1%) and BRN (-24 +/- 2%), with BRN losing more (P < 0.05) than BR. BRE maintained (P > 0.05) thigh muscle volume. Calf (triceps surae) muscle volume was decreased (P < 0.05) to a similar extent (P > 0.05) in BR (-29 +/- 1%) and BRN (-28 +/- 1%), and this decrease was attenuated (P < 0.05) in BRE (-8 +/- 2%). BR and BRN experienced large (P < 0.05) and similar (P > 0.05) decreases in isometric and dynamic (concentric force, eccentric force, power and work) muscle strength for supine squat (-19 to -33%) and calf press (-26 to -46%). BRE maintained (P > 0.05) or increased (P < 0.05) all measures of muscle strength. CONCLUSION: The nutrition countermeasure was not effective in offsetting lower limb muscle volume or strength loss, and actually promoted thigh muscle volume loss. The concurrent aerobic and resistance exercise protocol was effective at preventing thigh muscle volume loss, and thigh and calf muscle strength loss. While the exercise protocol offset approximately 75% of the calf muscle volume loss, modification of this regimen is needed.

Resistance exercise and cyclooxygenase (COX) expression in human skeletal muscle: implications for COX-inhibiting drugs and protein synthesis.

We have shown that ibuprofen and acetaminophen block cyclooxygenase (COX) synthesis of prostaglandin PGF(2alpha) and the muscle protein synthesis increase following resistance exercise. Confusingly, these two drugs are purported to work through different mechanisms, with acetaminophen apparently unable to block COX and ibuprofen able to nonspecifically block COX-1 and COX-2. A recently discovered intron-retaining COX, now known to have three variants, has been shown to be sensitive to both drugs. We measured the expression patterns and levels of the intron 1-retaining COX-1 variants (-1b1, -1b2, and -1b3), COX-1, and COX-2 at rest and following resistance exercise to help elucidate the COX through which PGF(2alpha), ibuprofen, and acetaminophen regulate muscle protein synthesis. Skeletal muscle biopsy samples were taken from 16 individuals (8M, 8F) before, 4, and 24 h after a bout of resistance exercise and analyzed using real-time RT-PCR. Relatively few individuals expressed the intron 1-retaining COX-1b variants (COX-1b1, -1b2, and -1b3) at any time point, and when expressed, these variants were in very low abundance. COX-1 was the most abundant COX mRNA before exercise and remained unchanged (P > 0.05) following exercise. COX-2 was not expressed before exercise, but increased significantly (P < 0.05) at 4 and 24 h after exercise. The inconsistent and low levels of expression of the intron 1-retaining COX-1 variants suggest that these variants are not likely responsible for the inhibition of PGF(2alpha) production and skeletal muscle protein synthesis after resistance exercise by ibuprofen and acetaminophen. Skeletal muscle-specific inhibition of COX-1 or COX-2 by these drugs should be considered.

Effects of 84-days of bedrest and resistance training on single muscle fibre myosin heavy chain distribution in human vastus lateralis and soleus muscles.

AIM: This investigation determined the effects of 84 days of bedrest on the composition of myosin heavy chain (MHC) in single skeletal muscle fibres with and without a resistance-training countermeasure programme. METHODS: Muscle biopsies were obtained from the m. vastus lateralis (VL) and m. soleus (SOL) before and after 84 days of bedrest. While control (BR) subjects (VL n = 9; SOL n = 3) refrained from exercise, BRE subjects (VL n = 8; SOL n = 3) performed knee extensor and plantar flexor resistance exercise every third day. Approximately 110 fibres per sample were analysed for MHC composition using SDS-PAGE. RESULTS: BR-VL had 16 and 14% decreases (P < 0.05) in MHC I and IIa fibres, respectively. There were 10% increases (P < 0.05) in MHC I/IIa, IIa/IIx, I/IIa/IIx, and a approximately 30% increase (P < 0.05) in total hybrid fibres. BRE-VL showed a 15% reduction (P < 0.05) in MHC I fibres, no change in MHC IIa fibres, and a 13% increase (P < 0.05) in total hybrids. BR-SOL had a 19% decrease (P < 0.05) in MHC I fibres with a 22% increase in total hybrids. BRE-SOL showed no change in MHC composition across all fibre types. CONCLUSION: These data suggest that the exercise countermeasures programme prevented MHC shifts in the SOL and mitigated MHC shifts in the VL. Furthermore, in the VL it appears that the resistance training programme employed in this investigation during bedrest, emphasized the use of MHC IIa phenotype muscle fibres.

Effects of short-term concentric vs. eccentric resistance training on single muscle fiber MHC distribution in humans.

The purpose of this investigation was to determine the effects of a concentric vs. eccentric resistance training program on single muscle fiber myosin heavy chain (MHC) adaptations in humans. Fifteen sedentary, healthy males were divided into three groups: concentric training (CTG) (n = 6, 24.2 +/- 1.7 y, 181 +/- 2 cm, 82.5 +/- 4.6 kg), eccentric training (ETG) (n = 6, 23.7 +/- 1.6 y, 178 +/- 3 cm, 90.4 +/- 6.1 kg), and control (CTL) (n = 3, 23 +/- 1.5 y, 181 +/- 2 cm, 97 +/- 13.2 kg). The subjects performed 4 sets of 8 unilateral repetitions starting at 80 % of concentric 1-RM, 3 days/week for a total of 4 weeks. Subjects were tested pre- and post-training for concentric 1-RM. Muscle biopsies were obtained from the vastus lateralis pre- and post-training for determination of single fiber MHC isoform distribution using SDS-PAGE/silver staining (100 fibers analyzed/subject pre- and post-training). Fibers expressing more than one MHC isoform (i.e., hybrid fibers) were analyzed for relative MHC isoform proportions via densitometry. The training program resulted in a 19 % 1-RM strength gain for CTG (p < 0.05) with no change in ETG or CTL. MHC-IIx fibers decreased by 7 % in CTG (p < 0.05) and ETG had an 11 % increase in total hybrids (MHC-I/IIa + MHC-IIa/IIx) (p < 0.05). No other differences were noted in MHC distribution among the three groups. Densitometry analysis of hybrid fibers showed no change in relative MHC isoform proportions pre- to post-training for any group. These data suggest that the MHC distribution did not change dramatically as a result of 4 weeks of concentric vs. eccentric resistance training despite the increase in whole muscle strength from concentric muscle actions.

Single muscle fiber contractile properties during a competitive season in male runners.

The purpose of this investigation was to examine the contractile properties of individual myofibers in response to periodized training periods throughout a collegiate cross-country season in male runners. Muscle biopsies of the gastrocnemius were taken after a summer base training phase (T1), an 8-wk intense training period (T2), and a 4-wk taper phase (T3). Five runners (n = 5; age = 20 +/- 1 yr; wt = 65 +/- 4 kg; ht = 178 +/- 3 cm) completed all three time points. A total of 328 individual muscle fibers [myosin heavy chain (MHC) I = 66%; MHC IIa = 33%; hybrids = 1%] were isolated and studied at 15 degrees C for their contractile properties. Diameter of MHC I fibers was 3% smaller (P < 0.05) at T2 compared with T1 and an additional 4% smaller (P < 0.05) after the taper. Cell size was unaltered in the MHC IIa fibers. MHC I and IIa fiber strength increased 18 and 11% (P < 0.05), respectively, from T1 to T2. MHC I fibers produced 9% less force (P < 0.05) after the taper, whereas MHC IIa fibers were 9% stronger (P < 0.05). Specific tension increased 38 and 26% (P < 0.05) for MHC I and IIa fibers, respectively, from T1 to T2 and was unchanged with the taper. Maximal shortening velocity (Vo) of the MHC I fibers decreased 23% (P < 0.05) from T1 to T2 and 17% (P < 0.05) from T2 to T3, whereas MHC IIa Vo was unchanged. MHC I peak power decreased 20% (P < 0.05) from T1 to T2 and 25% (P < 0.05) from T2 to T3, whereas MHC IIa peak power was unchanged. Power corrected for cell size decreased 15% (P < 0.05) from T2 to T3 and was 24% (P < 0.05) lower at T3 compared with T1 for the MHC I fibers only. These data suggest that changes in run training alter myocellular physiology via decreases in fiber size, Vo, and power of MHC I fibers and through increases in force per cross-sectional area of slow- and fast-twitch muscle fibers.

Mitogen-activated protein kinase (MAPK) pathway activation: effects of age and acute exercise on human skeletal muscle.

The purpose of this investigation was to examine the activation (phosphorylation) and total protein content of MAPK signalling cascade proteins (ERK 1/2, p90RSK, Mnk 1, eIF4E, p38 MAPK, JNK/SAPK, MKP 1) at rest and following exercise, in sedentary young and old men. Eight young (22 +/- 1 years; YM) and eight old (79 +/- 3 years; OM) men underwent a resting muscle biopsy of the vastus lateralis; they then performed a knee extensor resistance exercise session (29 contractions at approximately 70 % of max), followed by a post-exercise biopsy. Western immunoblot analysis demonstrated that the OM had higher resting phosphorylation of ERK 1/2, p90RSK, Mnk 1, p38 MAPK and JNK/SAPK proteins versus YM (P < 0.05). The resistance exercise bout caused an increase in phosphorylation of the ERK 1/2, p90RSK and Mnk 1 proteins (P < 0.05) in the YM. Conversely, the OM had a decrease in ERK 1/2, p90RSK, Mnk 1, p38 MAPK and JNK/SAPK phosphorylation (P < 0.05) after the exercise bout. Neither group showed a change in eIF4E phosphorylation. The total amount of protein expression of the MAPK signalling proteins was not different between the YM and OM, except that there was a higher (P < 0.05) MKP 1 protein content in the OM. This investigation is the first to provide evidence that MAPK proteins are differentially activated at rest and in response to a bout of resistance exercise in skeletal muscle of young and old men. These findings may have implications for other processes (e.g. transcription and translation) involved in skeletal muscle type and growth, when examining the changes occurring with ageing muscle before and after resistance exercise/training.

Myosin heavy chain composition of single muscle fibers in male distance runners.

The purpose of this study was to characterize the myosin heavy chain (MHC) composition of single muscle fibers from the gastrocnemius of male collegiate distance (DIST; n = 7), middle-distance (MID; n = 6), and recreational runners (REC; n = 6). Additionally, mATPase histochemistry was used to serve as a comparison to previous studies and the single fiber MHC technique. SDS-PAGE of single muscle fibers revealed a higher proportion of MHC I in DIST compared to MID and REC (74.9 +/- 4.3 vs 54.4 +/- 2.8 vs 56.2 +/- 2.9 %, respectively; p < 0.05), less MHC IIa/IIx in DIST compared to MID and REC (0.0 +/- 0.0 vs 6.0 +/- 2.4 vs 15.9 +/- 4.2 %, respectively; p < 0.05), and more total hybrids (I/IIa+IIa/IIx+I/IIa/IIx) in REC than both run groups, DIST and MID (23.0 +/- 3.3 vs 6.2 +/- 1.1 vs 13.2 +/- 2.6 %, respectively; p < 0.05). ATPase histochemistry (pH 4.54) revealed a higher percentage of type I fibers in DIST compared to MID and REC (71.1 +/- 3.1 vs 56.3 +/- 2.5 vs 59.8 +/- 2.3 %, respectively; p < 0.05), a higher percentage of type IIa in MID compared to DIST and REC (43.3 +/- 2.7 vs 28.5 +/- 3.1 vs. 30.2 +/- 3.1 %, p < 0.05), and a higher distribution of type IIb in REC than both run groups (10.0 +/- 2.7 vs 0.4 +/- 0.2 vs 0.4 +/- 0.2 %, p < 0.05). These results suggest that distance running leads to an increase in MHC I expression, training for mid-distance events leads to a prevalence of MHC IIa, and run training leads to a decrease in hybrid fibers.

Alterations in single muscle fiber calcium sensitivity with resistance training in older women.

The purpose of this investigation was to determine the effects of a 12-week progressive resistance-training program (PRT) on single muscle fiber calcium sensitivity in six older women (73 +/- 2 years). Muscle biopsy samples of the vastus lateralis were obtained pre- and post-PRT. Chemically skinned single muscle fibers ( n=274) were dissected and studied. The experimental sequence for each fiber was the determination of peak maximal isometric tension ( P(o)) at pCa 4.5 (pCa=-log[Ca(2+)]), and then subsequent submaximal activations of the fiber at nine Ca(2+) concentrations (pCa 6.8 to 4.7). Myosin heavy chain (MHC) I fiber (slow-twitch) diameter increased 16% ( P<0.05) with no change in MHC IIa fibers (fast-twitch) pre- to post-PRT, respectively. P(o) in MHC I fibers increased 34% ( P<0.05) as a result of the training with no change in MHC IIa fibers. The mean MHC I Ca(2+) activation threshold (minimal amount of Ca(2+) necessary to induce tension) increased from 6.83 +/- 0.02 to 6.91 +/- 0.01 ( P<0.05), as did the mean half-maximal activation (pCa(50)), 5.51 +/- 0.02 to 5.71 +/- 0.03 ( P<0.05) with PRT. The slope of the Hill plot above ( n(1)) the pCa(50) for MHC I did not change significantly with the PRT. However, the slope of the Hill plot below ( n(2)) the pCa(50) for MHC I demonstrated an increase ( P<0.05) with training. There were no differences with MHC IIa fibers with PRT for any of the variables measured. In conclusion, the results of this investigation indicate that myofibril Ca(2+) sensitivity and activation properties are altered in MHC I, but not MHC IIa fibers with PRT in older women. The alterations in the MHC I Ca(2+) properties appear to have an effect on the mechanisms involved with skeletal muscle adaptability in older women following PRT.

Unilateral lower limb suspension does not mimic bed rest or spaceflight effects on human muscle fiber function.

We used Ca2+-activated skinned muscle fibers to test the hypothesis that unilateral lower leg suspension (ULLS) alters cross-bridge mechanisms of muscle contraction. Soleus and gastrocnemius biopsies were obtained from eight subjects before ULLS, immediately after 12 days of ULLS (post-0 h), and after 6 h of reambulation (post-6 h). Post-0 h soleus fibers expressing type I myosin heavy chain (MHC) showed significant reductions in diameter, absolute and specific peak Ca2+-activated force, unloaded shortening velocity, and absolute and normalized peak power. Fibers obtained from the gastrocnemius were less affected by ULLS, particularly fibers expressing fast MHC isoforms. Post-6 h soleus fibers produced less absolute and specific peak force than did post-0 h fibers, suggesting that reambulation after ULLS induced cell damage. Like bed rest and spaceflight, ULLS primarily affects soleus over gastrocnemius fibers. However, in contrast to these other models, slow soleus fibers obtained after ULLS showed a decrease in unloaded shortening velocity and a greater reduction in specific force.

Reduction in hybrid single muscle fiber proportions with resistance training in humans.

The purpose of this investigation was to examine the effects of 12 wk of progressive resistance training (PRT) on single muscle fiber myosin heavy chain (MHC; I, I/IIa, I/IIa/IIx, IIa, IIa/IIx, IIx) isoform proportions in young individuals. Young, untrained men (YM; n = 6) and women (YW; n = 6) (age = 22 +/- 1 and 25 +/- 2 yr for YW and YM, respectively) received pre- and post-PRT muscle biopsies from the right vastus lateralis for single muscle fiber MHC distribution by electrophoretic analysis (192 +/- 5 pre- and 183 +/- 6 post-fibers/subject analyzed; 4,495 fibers total). Data are presented as percentages of the total fibers analyzed per subject. The PRT protocol elicited an increase in the pure MHC IIa (Delta = + 24 and + 27; YW and YM, respectively; P < 0.05) with no change in the pure MHC I distribution. The hybrid MHC distributions decreased I/IIa/IIx (Delta = -2; YM and YW; P < 0.05), IIa/IIx (Delta = -13 and -19 for YM and YW, respectively; P < 0.05), and total hybrid fiber proportion (I/IIa + I/IIa/IIx + IIa/IIx) decreased (Delta = -19 and -30 for YM and YW, respectively; P < 0.05) with the training, as did the MHC IIx distribution (Delta = -2; YW only; P < 0.05). Alterations in the predominance of MHC isoforms within hybrid fibers (decrease in MHC I-dominant I/IIa and nondominant MHC IIa/IIx, increase in MHC IIa-dominant IIa/IIx; P < 0.05) appeared to contribute to the increase in the MHC IIa proportion. Electrophoresis of muscle cross sections revealed an approximately 7% increase (P < 0.05) in MHC IIa proportion in both groups, whereas the MHC IIx decrease by 7.5 and 11.6% post-PRT in YW and YM, respectively. MHC I proportions increase in YM by 4.8% (P < 0.05) post-PRT. These findings further support previous resistance training data in young adults with respect to the increase in the MHC IIa proportions but demonstrate that a majority of the change can be attributed to the decrease in single-fiber hybrid proportions.

Production of tissue plasminogen activator (t-PA) in Aspergillus niger.

A protease-deficient strain of Aspergillus niger has been used as a host for the production of human tissue plasminogen activator (t-PA). In defined medium, up to 0.07 mg t-PA (g biomass)(-1) was produced in batch and fed-batch cultures and production was increased two- to threefold in two-phase batch cultures in which additional glucose was provided as a single pulse at the end of the first batch growth phase. Production was increased [up to 1.9 mg t-PA (g biomass)(-1)] by the addition of soy peptone to the defined medium. The rate of t-PA production in batch cultures supplemented with soy peptone (0.2 to 0.6 mg t-PA L(-1) h(-1)) was comparable to rates observed previously in high-producing mammalian or insect cell cultures. In glucose-limited chemostat culture supplemented with soy peptone, t-PA was produced at a rate of 0.7 mg t-PA L(-1) h(-1). Expression of t-PA in A. niger resulted in increased expression of genes (bipA, pdiA, and cypB) involved in the unfolded protein response (UPR). However, when cypB was overexpressed in a t-PA-producing strain, t-PA production was not increased. The t-PA produced in A. niger was cleaved into two chains of similar molecular weight to two-chain human melanoma t-PA. The two chains appeared to be stable for at least 16 h in culture supernatant of the host strain. However, in general, <1% of the t-PA produced in A. niger was active, and active t-PA disappeared from the culture supernatant during the stationary phase of batch cultures, suggesting that the two-chain t-PA may have been incorrectly processed or that initial proteolytic cleavage occurred within the proteolytic domain of the protein. Total t-PA (detected by enzyme-linked immunoassay) also eventually disappeared from culture supernatants, confirming significant extracellular proteolytic activity, even though the host strain was protease-deficient.