Early changes in tests of peripheral nerve function during oxaliplatin treatment and their correlation with chemotherapy-induced polyneuropathy symptoms and signs.

BACKGROUND AND PURPOSE: Assessment of the severity of chronic peripheral neuropathy during oxaliplatin treatment is based on symptoms. Efforts to adjust the total dose of oxaliplatin to prevent severe neuropathy can be complicated by the worsening of neuropathy symptoms following treatment. Objective measures of the structure and function of peripheral nerves during early phases of treatment may aid in determining the optimal oxaliplatin dose in individual patients. Intraepidermal nerve fibre density (IENFD) has been suggested as an early marker of peripheral neuropathy. METHODS: Sixty patients were examined before treatment and following 25% and 50% of the total planned oxaliplatin dose. Fifty-five of them were also examined at completion of chemotherapy and 6 months later. IENFD in skin biopsies from the distal leg, nerve conduction studies and quantitative sensory testing at the dorsum of the foot were performed. Forty-six healthy subjects were examined at baseline and after 6 and 52 weeks for comparison. RESULTS: Intraepidermal nerve fibre density was not reduced during treatment. Sural nerve amplitude and conduction velocity, vibration detection thresholds, mechanical detection threshold and cold detection threshold were significantly reduced during treatment. Compared to reference values and spontaneous changes in healthy subjects, the largest proportions of patients with deterioration were found for vibration detection thresholds followed by nerve conduction studies, mechanical detection threshold, cold detection threshold and IENFD. CONCLUSIONS: Significant changes were most pronounced for measures of large nerve fibre function, especially vibration sensation. Skin biopsies do not seem to provide a clinically relevant objective measure of peripheral nerve deterioration during oxaliplatin treatment.

Mitochondria, glycogen, and lipid droplets in skeletal muscle during testosterone treatment and strength training: a randomized, double-blinded, placebo-controlled trial.

Low testosterone levels in aging men are associated with insulin resistance. Mitochondrial dysfunction, changes in glycogen metabolism, and lipid accumulation are linked to insulin resistance in skeletal muscle. In this randomized, double-blinded, placebo-controlled study, we investigated the effects of six-month testosterone replacement therapy (TRT) and strength training (ST) on mitochondrial, glycogen, and lipid droplet (LD) content in skeletal muscle of aging men with subnormal bioavailable testosterone (BioT) levels. Mitochondrial, glycogen, and LD volume fractions in muscle biopsies were estimated by transmission electron microscopy. Insulin sensitivity (insulin-stimulated Rd) and body composition were assessed by euglycemic-hyperinsulinemic clamp and dual X-ray absorptiometry, respectively. TRT significantly increased total testosterone levels, BioT, and lean body mass (LBM) (p < 0.05), whereas percent body fat decreased (p < 0.05), and insulin sensitivity was unchanged. Baseline mitochondrial volume fraction correlated inversely with percent body fat (ρ = -0.43; p = 0.003). Δ-mitochondrial fraction correlated positively with Δ-total testosterone (ρ = 0.70; p = 0.02), and Δ-glycogen fraction correlated inversely with Δ-LBM (ρ = -0.83; p = 0.002) during six-month TRT, but no significant changes were observed in mitochondrial, glycogen, and LD volume fractions during TRT and ST. In conclusion, in this exploratory small-scale study, the beneficial effects of six-month TRT on total testosterone, LBM, and percent body fat were not followed by significant changes in fractions of mitochondria, glycogen, or lipid in skeletal muscle of aging men with lowered testosterone levels. Six-month ST or combined three-month ST+TRT did not change intramyocellular mitochondria, glycogen, and LD fractions compared to placebo. However, further studies with a larger sample size are needed.

Influence of Resistance Training on Neuromuscular Function and Physical Capacity in ALS Patients.

OBJECTIVES: The present study aimed to explore the effect of resistance training in patients with amyotrophic lateral sclerosis (ALS), a disease characterized by progressive motor neuron loss and muscle weakness. MATERIALS AND METHODS: Following a 12-week "lead-in" control period, a population of ALS patients from Funen, Denmark, completed a 12-week resistance training program consisting of 2-3 sessions/week. Neuromuscular function (strength and power) and voluntary muscle activation (superimposed twitch technique) were evaluated before and after both control and training periods. Physical capacity tests (chair rise and timed up and go), the revised ALS functional rating scale (ALSFRS-R) scores, and muscle cross sectional area (histology) were also assessed. RESULTS: Of twelve ALS patients assessed for eligibility, six were included and five completed the study. Training did not significantly affect the ALSFRS-R score, and loss of neuromuscular function (strength and power) increased following the training period. However, an improved functionality (chair rise) and an increase in greatly hypertrophied type II fibres combined with an increase in atrophied fibres following the training period compared to the control period were observed. CONCLUSION: In this small study, the present form of resistance training was unable to attenuate progressive loss of neuromuscular function in ALS, despite some changes in physical capacity and morphology.

Muscular response to the first three months of deflazacort treatment in boys with Duchenne muscular dystrophy.

OBJECTIVE: Duchenne muscular dystrophy (DMD) patients are often treated with glucocorticoids; yet their precise molecular action remains unknown. METHODS: We investigated muscle biopsies from nine boys with DMD (aged: 7,6±2,8 yrs.) collected before and after three months of deflazacort treatment and compared them to eight healthy boys (aged: 5,3±2,4 yrs.). mRNA transcripts involved in activation of satellite cells, myogenesis, regeneration, adipogenesis, muscle growth and tissue inflammation were assessed. Serum creatine kinase (CK) levels and muscle protein expression by immunohistochemistry of selected targets were also analysed. RESULTS: Transcript levels for ADIPOQ, CD68, CDH15, FGF2, IGF1R, MYF5, MYF6, MYH8, MYOD, PAX7, and TNFα were significantly different in untreated patients vs. normal muscle (p⟨0.05). Linear tests for trend indicated that the expression levels of treated patients were approaching normal values (p⟨0.05) following treatment (towards an increase; CDH15, C-MET, DLK1, FGF2, IGF1R, MYF5, MYF6, MYOD, PAX7; towards a decrease: CD68, MYH8, TNFα). Treatment reduced CK levels (p⟨0.05), but we observed no effect on muscle protein expression. CONCLUSIONS: This study provides insight into the molecular actions of glucocorticoids in DMD at the mRNA level, and we show that multiple regulatory pathways are influenced. This information can be important in the development of new treatments.

Skeletal Muscle Remodelling as a Function of Disease Progression in Amyotrophic Lateral Sclerosis.

Muscle weakness is considered the pivotal sign of amyotrophic lateral sclerosis (ALS). Knowledge about the skeletal muscle degeneration/regeneration process and the myogenic potential is limited in ALS patients. Therefore, we investigate these processes in a time course perspective by analysing skeletal muscle biopsies from ALS patients collected before and after a 12-week period of normal daily activities and compare these with healthy age-matched control tissue. We do this by evaluating mRNA and protein (immunohistochemical) markers of regeneration, neurodegeneration, myogenesis, cell cycle regulation, and inflammation. Our results show morphological changes indicative of active denervation and reinnervation and an increase in small atrophic fibres. We demonstrate differences between ALS and controls in pathways controlling skeletal muscle homeostasis, cytoskeletal and regenerative markers, neurodegenerative factors, myogenic factors, cell cycle determinants, and inflammatory markers. Our results on Pax7 and MyoD protein expression suggest that proliferation and differentiation of skeletal muscle stem cells are affected in ALS patients, and the myogenic processes cannot overcome the denervation-induced wasting.

Neuronal nitric oxide synthase is dislocated in type I fibers of myalgic muscle but can recover with physical exercise training.

Trapezius myalgia is the most common type of chronic neck pain. While physical exercise reduces pain and improves muscle function, the underlying mechanisms remain unclear. Nitric oxide (NO) signaling is important in modulating cellular function, and a dysfunctional neuronal NO synthase (nNOS) may contribute to an ineffective muscle function. This study investigated nNOS expression and localization in chronically painful muscle. Forty-one women clinically diagnosed with trapezius myalgia (MYA) and 18 healthy controls (CON) were included in the case-control study. Subsequently, MYA were randomly assigned to either 10 weeks of specific strength training (SST, n = 18), general fitness training (GFT, n = 15), or health information (REF, n = 8). Distribution of fiber type, cross-sectional area, and sarcolemmal nNOS expression did not differ between MYA and CON. However, MYA showed increased sarcoplasmic nNOS localization (18.8 ± 12 versus 12.8 ± 8%, P = 0.049) compared with CON. SST resulted in a decrease of sarcoplasm-localized nNOS following training (before 18.1 ± 12 versus after 12.0 ± 12%; P = 0,027). We demonstrate that myalgic muscle displays altered nNOS localization and that 10 weeks of strength training normalize these disruptions, which supports previous findings of impaired muscle oxygenation during work tasks and reduced pain following exercise.

Characterization and diagnostic evaluation of chronic polyneuropathies induced by oxaliplatin and docetaxel comparing skin biopsy to quantitative sensory testing and nerve conduction studies.

BACKGROUND AND PURPOSE: Chemotherapy-induced peripheral neuropathy negatively affects the quality of life for many patients treated with oxaliplatin or docetaxel for gastrointestinal cancer or breast cancer. Symptoms can persist long after treatment and often include neuropathic pain. Our objective was to characterize the neuropathies with regard to symptoms, neurological signs and objective evidence of damage to the structure and function of the peripheral nerves. Furthermore, the diagnostic values of skin biopsy, quantitative sensory testing (QST) and nerve conduction studies (NCS) were compared. METHODS: Patients complaining of neuropathy symptoms at least 3 months after completion of treatment with oxaliplatin (n = 20) or docetaxel (n = 20) were recruited from the Department of Oncology or using hospital records. Neuropathy scores were determined along with the intraepidermal nerve fibre density in skin biopsies from the proximal and distal parts of the leg, QST and NCS. RESULTS: Clinically only sensory functions were affected. In general, neuropathy scores were higher in the oxaliplatin-treated group. Both sensory and motor fibres were affected in the NCS, showing predominantly signs of axonal damage. Mechanical detection threshold was most often affected in the QST. NCS, QTS and skin biopsy were abnormal in 11, 13 and 17 and 7, 11 and 15 of the oxaliplatin-treated patients and docetaxel-treated patients, respectively. CONCLUSIONS: Chemotherapy-induced peripheral neuropathy after oxaliplatin or docetaxel treatment is a clinically sensory, axonal neuropathy affecting only small nerve fibres in some patients. NCS are often normal, whereas QST and skin biopsy have a higher diagnostic sensitivity.

Ageing is associated with diminished muscle re-growth and myogenic precursor cell expansion early after immobility-induced atrophy in human skeletal muscle.

Recovery of skeletal muscle mass from immobilisation-induced atrophy is faster in young than older individuals, yet the cellular mechanisms remain unknown. We examined the cellular and molecular regulation of muscle recovery in young and older human subjects subsequent to 2 weeks of immobility-induced muscle atrophy. Retraining consisted of 4 weeks of supervised resistive exercise in 9 older (OM: mean age) 67.3, range 61-74 yrs) and 11 young (YM: mean age 24.4, range 21-30 yrs) males. Measures of myofibre area (MFA), Pax7-positive satellite cells (SCs) associated with type I and type II muscle fibres, as well as gene expression analysis of key growth and transcription factors associated with local skeletal muscle milieu, were performed after 2 weeks immobility (Imm) and following 3 days (+3d) and 4 weeks (+4wks) of retraining. OM demonstrated no detectable gains in MFA (vastus lateralis muscle) and no increases in number of Pax7-positive SCs following 4wks retraining, whereas YM increased their MFA (P < 0.05), number of Pax7-positive cells, and had more Pax7-positive cells per type II fibre than OM at +3d and +4wks (P < 0.05). No age-related differences were observed in mRNA expression of IGF-1Ea, MGF, MyoD1 and HGF with retraining, whereas myostatin expression levels were more down-regulated in YM compared to OM at +3d (P < 0.05). In conclusion, the diminished muscle re-growth after immobilisation in elderly humans was associated with a lesser response in satellite cell proliferation in combination with an age-specific regulation of myostatin. In contrast, expression of local growth factors did not seem to explain the age-related difference in muscle mass recovery.

Use of statins and risk of glioma: a nationwide case-control study in Denmark.

BACKGROUND: Laboratory studies and a single case-control study have suggested a protective effect of statins on the risk of glioma. We wished to investigate the influence of statin use on the risk of glioma in a population-based setting. METHODS: We conducted a nationwide case-control study in Denmark based on population-based medical registries. We identified all patients aged 20 to 85 years with a first diagnosis of histologically verified glioma during 2000-2009. These cases were matched on birth year and sex with population controls. Prior use of statins since 1995 was classified into short-term use (<5 years) and long-term use (5+ years). We used conditional logistic regression to compute odds ratios (ORs), with 95% confidence intervals (CIs), for glioma associated with statin use, adjusted for potential confounders. RESULTS: A total of 2656 cases and 18,480 controls were included in the study. The risk of glioma was reduced among long-term statin users (OR=0.76; 95% CI: 0.59-0.98) compared with never users of statins, and was inversely related to the intensity of statin treatment among users (OR=0.71; 95% CI: 0.44-1.15 for highest intensity). The inverse association between long-term statin treatment and glioma risk was more pronounced among men aged ≤ 60 years (OR=0.40; 95% CI: 0.17-0.91) compared with men aged 60+ years (OR=0.71; 95% CI: 0.49-1.03). An inverse association was also observed among women aged ≤ 60 years (OR=0.28; 95% CI: 0.06-1.25), but not among women over age 60 years (OR=1.23; 95% CI: 0.82-1.85). CONCLUSION: Long-term statin use may reduce the risk of glioma.

Distinct effects of subcellular glycogen localization on tetanic relaxation time and endurance in mechanically skinned rat skeletal muscle fibres.

In vitro experiments indicate a non-metabolic role of muscle glycogen in contracting skeletal muscles. Since the sequence of events in excitation\#8211;contraction (E\#8211;C) coupling is known to be located close to glycogen granules, at specific sites on the fibre, we hypothesized that the distinct compartments of glycogen have specific effects on muscle fibre contractility and fatigability. Single skeletal muscle fibres (n = 19) from fed and fasted rats were mechanically skinned and divided into two segments. In one segment glycogen localization and volume fraction were estimated by transmission electron microscopy. The other segment was mechanically skinned and, in the presence of high and constant myoplasmic ATP and PCr, electrically stimulated (10 Hz, 0.8 s every 3 s) eliciting repeated tetanic contractions until the force response was decreased by 50% (mean +/- S.E.M., 81 +/- 16, range 22-252 contractions). Initially the total myofibrillar glycogen volume percentage was 0.46 +/- 0.07%, with 72 +/- 3% in the intermyofibrillar space and 28 +/- 3% in the intramyofibrillar space. The intramyofibrillar glycogen content was positively correlated with the fatigue resistance capacity (r(2) = 0.32, P = 0.02). Intermyofibrillar glycogen was inversely correlated with the half-relaxation time in the unfatigued tetanus (r(2) = 0.25, P = 0.03). These results demonstrate for the first time that two distinct subcellular populations of glycogen have different roles in contracting single muscle fibres under conditions of high myoplasmic ATP.

The histopathology of a human mesenchymal stem cell experimental tumor model: support for an hMSC origin for Ewing's sarcoma?

Sarcomas display varied degrees of karyotypic abnormality, vascularity and mesenchymal differentiation. We have reported that a strain of telomerized adult human bone marrow mesenchymal stem cells (hMSC-TERT20) spontaneously evolved a tumorigenic phenotype after long-term continuous culture. We asked to what extent our hMSC-TERT20 derived tumors reflected events found in human sarcomas using routine histopathological procedures. Early versus late passage hMSC-TERT20 cultures persistently expressed mesenchymal lineage proteins e.g. CD105, CD44, CD99 and vimentin. However, late passage cultures, showed increased immunohistochemical staining for CyclinD1 and p21WAF1/Cip1, whereas p27Kip1 staining was reduced. Notably, spectral karyotyping showed that tumorigenic hMSC-TERT20 cells retained a normal diploid karyotype, with no detectable chromosome abnormalities. Consistent with the bone-forming potential of early passage hMSC-TERT20 cells, tumors derived from late passage cells expressed early biomarkers of osteogenesis. However, hMSC-TERT20 cells were heterogeneous for alpha smooth muscle actin (ASMA) expression and one out of six hMSC-TERT20 derived single cell clones was strongly ASMA positive. Tumors from this ASMA+ clone had distinctive vascular qualities with hot spots of high CD34+ murine endothelial cell density, together with CD34- regions with a branching periodic acid Schiff reaction pattern. Such clone-specific differences in host vascular response provide novel models to explore interactions between mesenchymal stem and endothelial cells. Despite the lack of a characteristic chromosomal translocation, the histomorphology, biomarkers and oncogenic changes were similar to those prevalent for Ewing's sarcomas. The phenotype and ontogenesis of hMSC-TERT20 tumors was consistent with the hypothesis that sarcomas may arise from hMSC, providing a unique diploid model for exploring human sarcoma biology.

Tissue distribution and engraftment of human mesenchymal stem cells immortalized by human telomerase reverse transcriptase gene.

Engraftment of mesenchymal stem cells (MSC) in peripheral tissues for replenishing of local stem cell function has been proposed as a therapeutic approach to degenerative diseases. We have previously reported the development of an immortalized human telomerase reverse transcriptase transduced MSC line (hMSC-TERT). In the present study, we co-transduced hMSC-TERT with enhanced green fluorescent protein gene, and studied tissue distribution, engraftment, and cell survival after intracardiac and intravenous injections in immunodeficient mice. The pattern of organ distribution suggested that infused cells were efficiently arrested in microvasculature during first-pass, but only for a fraction of the infused cells was arrest followed by vascular emigration and tissue engraftment. Few engrafted cells in lungs, heart, and kidney glomeruli remained after 4 weeks. These observations are consistent with several reports on limited systemic transplantability of primary MSC. HMSC-TERT may constitute a valuable tool for mechanistic studies on how to control MSC homing and engraftment.

Improved histopathological evaluation of gliomas using tissue fragments obtained by ultrasonic aspiration.

Ultrasonic aspirators are commonly used to resect brain tumors because they allow safe, rapid and accurate removal of dissected tissue. However, the tissue fragments removed by ultrasonic aspirators are used surprisingly little in daily diagnostics and in clinical research. A comparison between diagnoses made on the tissue fragments removed by the Sonocut ultrasonic aspirator and the tissue in conventional tumor biopsies was made. The correspondence between the amount of Sonocut tissue analyzed and the probability of making the most malignant diagnosis was analyzed statistically in order to limit the amount of tissue fragments, which need to be analyzed, and in order to save tissue for other purposes like molecular genetics and in vitro drug sensitivity assays. Thirty cases were included in the present study and consisted of astrocytomas, glioblastomas, oligodendrogliomas and ependymomas. The results showed that in 8 out of 30 cases only the Sonocut tissue fragments contained the tumor components that provided the most malignant diagnosis. In further 20 cases, the Sonocut tissue fragments and conventional tumor biopsies gave the same diagnoses. In the remaining 2 cases, the most malignant foci were included in the biopsy removed for peroperative frozen section investigation. When the slides with Sonocut tissue fragments were analyzed, the probability of making the most malignant diagnosis increased from 81.3% - 99.1%, when slides from 1 - 5 paraffin blocks were analyzed, respectively. When subgroups of small, medium and big tumors were analyzed, it was found that only 2 paraffin blocks from small tumors need to be prepared to reach 98.3% probability of making the most malignant diagnosis, whereas 5 paraffin blocks from big tumors need to be prepared to reach a 96.8% probability. In conclusion, the study shows that a limited amount of Sonocut ultrasonic tissue fragments improve the diagnostic evaluation of gliomas. These tissue fragments therefore must not be discarded. Only few paraffin blocks need to be prepared to reach close to 100% probability of making the most malignant diagnosis, reducing the amount of slides, which have to be analyzed and saving Sonocut tissue fragments for future use in molecular genetics and drug sensitivity assays.

GLUT11, but not GLUT8 or GLUT12, is expressed in human skeletal muscle in a fibre type-specific pattern.

Nine novel sugar transporter-like proteins have been discovered in the past 5 years. The mRNA for three of these, the glucose transporters (GLUT) GLUT8, GLUT11 and GLUT12, have been detected in human skeletal muscle. In the present study, we examined the pattern of expression and localization of the GLUT isoforms 8, 11 and 12 in human skeletal muscle using an immunohistochemical approach. Biopsies of human skeletal muscle from sedentary or trained healthy adults, from fetal muscle (24 weeks of gestation), from obese type-2 diabetic subjects, and from patients suffering from polymyositis or amyotrophic lateral sclerosis (ALS) were studied. GLUT8 and 12 immunoreactivity was below detection level in both developing and adult muscle fibres. GLUT11 immunoreactivity, however, was present in slow-twitch muscle fibres, but not in fast twitch fibres. Since, in contrast, GLUT4 was expressed in all investigated muscle fibres, the pattern of expression of GLUT11 differs from that of GLUT4, suggesting a specialized function for GLUT11 with a regulation independent from that of GLUT4. Obesity, type-2 diabetes, training, conditions of de- and reinnervation (ALS) and regeneration (polymyositis) failed to induce GLUT8 or -12 expression. Likewise, the fibre type-dependent pattern of GLUT11 immunoreactivity was unaltered. However, some slow muscle fibres lose their GLUT11 immunoreactivity under regeneration. Our results indicate that GLUT11 immunoreactivity, in contrast to that of GLUT4, is expressed exclusively in slow-twitch muscle fibres and is unaffected by physiological and pathophysiological conditions except in primary myopathy. GLUT8 and GLUT12 do not appear to be of importance in human muscle under physiological and pathophysiological conditions.

Functional and immunohistochemical evaluation of porcine neonatal islet-like cell clusters.

Porcine neonatal islet-like cell clusters (NICCs) may be an attractive source of insulin-producing tissue for xenotransplantation in type I diabetic patients. We examined the functional and immunohistochemical outcome of the islet grafts in vitro during long-term culture and in vivo after transplantation to athymic nude mice. On average we obtained 29,000 NICCs from each pancreas. In a perifusion system, NICCs responded poorly to a glucose challenge alone, but 10 mmol/L arginine elicited a fourfold increase in insulin secretion and 16.7 mmol/L glucose + 10 mmol/L arginine caused a sevenfold increase in insulin section indicating some sensitivity towards glucose. Hormone content as well as the number of hormone-containing cells increased for the first 14 days of culture. When NICCs were stained for hormones, proliferation (Ki67), and duct cells (CK7), some insulin- and glucagon-positive cells co-stained for proliferation. However no co-staining was observed between insulin- and glucagon-positive cells or between hormone-and CK-positive cells. Following transplantation of 2000 NICCs under the renal capsule of diabetic nude mice, BG levels were normalized within an average of 13 weeks. Oral and IP glucose tolerance tests revealed a normal or even faster clearance of a glucose load compared with normal controls. Immunohistochemical examination of the grafts revealed primarily insulin-positive cells. In summary, in vitro, NICCs responded to a challenge including glucose and arginine. There was a potential for expansion of the beta-cell mass of NICCs in vitro as well as in vivo where NICCs eventually may normalize blood glucose of diabetic mice.

GLUT4 expression in human muscle fibres is not correlated with intracellular triglyceride (TG) content. Is TG a maker or a marker of insulin resistance?

We have recently reported a progressive decline in the expression of glucose transporter isoform 4 (GLUT4) from control subjects through obese non-diabetics to obese type 2 diabetic subjects, indicating that the reduced GLUT4 in slow twitch fibres could be secondary to obesity. In this study we investigate the association of GLUT4 expression with the intracellular triglyceride (TG) content in the same muscle fibres and with plasma lipid parameters. We used histochemistry and stereology to study the relationship between TG content and GLUT4 expression in muscle fibres from obese, obese type 2 diabetic subjects, and young lean controls. TG density was significantly higher in slow compared to fast fibres in all studied subjects (p<0.05). We found an increased TG density in slow twitch fibres of obese diabetic subjects compared to obese (p<0.05) and lean controls (p<0.008). Intracellular TG densities in slow and fast fibres did not correlate with the corresponding GLUT4 density in the same fibres in our study groups (p>0.05). Plasma TG and FFA did not correlate with GLUT4 expression in slow or fast fibres (p>0.05). In conclusion, TG content was increased in diabetic slow fibres with a reduced GLUT4 expression. The GLUT4 expression was not associated with an increased intracellular triglyceride content or with increased plasma FFA levels. Thus, intracellular TG content and circulating FFA may not influence glucose transport directly through GLUT4 expression.

Regenerating human muscle fibres express GLUT3 protein.

The presence of the GLUT3 glucose transporter protein in human muscle cells is a matter of debate. The present study was designed to establish whether GLUT3 is expressed in mature human skeletal muscle fibres and, if so, whether its expression changes under different conditions, such as metabolic stress (obesity, obese non-insulin-dependent diabetes mellitus), hypertrophy (training), de- and reinnervation (amyotrophic lateral sclerosis) or regeneration (polymyositis). We used an immunohistochemical approach to detect and localise GLUT3. GLUT3 immunoreactivity was not detectable in adult skeletal muscle fibres, nor did metabolic stress, training or de- and re-innervation induce GLUT3 expression, while a few GLUT3 expressing fibres were seen in some cases of polymyositis. In contrast, GLUT4 was expressed in all investigated muscle fibres. GLUT3 immunoreactivity was found in perineural and endoneural cells, indicating that GLUT3 is important for glucose transport into nerves through the perineurium. Taken together, these data suggest that GLUT3 expression is restricted to regenerating muscle fibres and nerves in adult human muscle. Although the significance of GLUT3 in adult human muscle fibres appears limited, GLUT3 may be of importance for the glucose supply in fetal muscle fibres and regenerating adult muscle fibres.

GLUT4 expression at the plasma membrane is related to fibre volume in human skeletal muscle fibres.

In this study we examined the relationship between GLUT4 expression at the plasma membrane and muscle fibre size in fibre-typed human muscle fibres by immunocytochemistry and morphometry in order to gain further insight into the regulation of GLUT4 expression. At the site of the plasma membrane, GLUT4 was more abundantly expressed in slow as compared to fast fibres at the same fibre diameter (p < 0.01) and the GLUT4 expression increased with increasing fibre radius independently of fibre type (p < 0.01). The GLUT4 density at the surface of slow fibres of both diabetic and obese was reduced compared to control subjects at the same diameter (p < 0.001). Fast fibres in obese and type 2 diabetic subjects expressed a fibre-volume-dependent GLUT4 expression (p < 0.001), while this did not reach significance in slow fibres (obese p = 0.18 and diabetic p = 0.06). Our results show that increasing fibre volume is associated with increasing GLUT4 expression in both slow and fast fibres. Based on the possible dependency of GLUT4 expression on volume, we hypothesize that the reduced GLUT4 expression in obesity and type 2 diabetes may partly be compensated for by physical activity.

The basal kinetic parameters of glycogen synthase in human myotube cultures are not affected by chronic high insulin exposure.

There is no consensus regarding the results from in vivo and in vitro studies on the impact of chronic high insulin and/or high glucose exposure on acute insulin stimulation of glycogen synthase (GS) kinetic parameters in human skeletal muscle. The aim of this study was to evaluate the kinetic parameters of glycogen synthase activity in human myotube cultures at conditions of chronic high insulin combined or not with high glucose exposure, before and after a subsequent acute insulin stimulation. Acute insulin stimulation significantly increased the fractional activity (FV(0.1)) of GS, increased the sensitivity of GS to the allosteric activator glucose 6-phosphate (A(0.5)) and increased the sensitivity of GS to its substrate UDPG (K(m(0.1))) when myotubes were precultured at low insulin with/without high glucose conditions. However, this effect of acute insulin stimulation was abolished in myotubes precultured at high insulin with or without high glucose. Furthermore, we found significant correlations between the fractional velocities FV(0.1) of GS and K(m(0.1)) (rho=-0.72, P<0.0001), between FV(0.1) and A(0.5) (rho=-0.82, P<0.0001) and between K(m(0.1)) and A(0.5) values (rho=0.71, P<0.0001). Our results show that chronic exposure of human myotubes to high insulin with or without high glucose did not affect the basal kinetic parameters but abolished the reactivity of GS to acute insulin stimulation. We suggest that insulin induced insulin resistance of GS is caused by a failure of acute insulin stimulation to decrease A(0.5) and K(m(0.1)) in human skeletal muscle.