Steen Larsen

Biomarkers of mitochondrial content in skeletal muscle of healthy young human subjects

•  Several biochemical measures of mitochondrial components are used as biomarkers of mitochondrial content and muscle oxidative capacity. However, no studies have validated these surrogates against a morphological measure of mitochondrial content in human subjects. •  The most commonly used markers (citrate synthase activity, cardiolipin content, mitochondrial DNA content (mtDNA), complex I–V protein, and complex I–IV activity) were correlated with a measure of mitochondrial content (transmission electron microscopy) and muscle oxidative capacity (respiration in permeabilized fibres). •  Cardiolipin content followed by citrate synthase activity and complex I activity were the biomarkers showing the strongest association with mitochondrial content. •  mtDNA was found to be a poor biomarker of mitochondrial content. •  Complex IV activity was closely associated with mitochondrial oxidative phosphorylation capacity.

Simvastatin Effects on Skeletal Muscle: Relation to Decreased Mitochondrial Function and Glucose Intolerance

OBJECTIVES Glucose tolerance and skeletal muscle coenzyme Q(10) (Q(10)) content, mitochondrial density, and mitochondrial oxidative phosphorylation (OXPHOS) capacity were measured in simvastatin-treated patients (n = 10) and in well-matched control subjects (n = 9). BACKGROUND A prevalent side effect of statin therapy is muscle pain, and yet the basic mechanism behind it remains unknown. We hypothesize that a statin-induced reduction in muscle Q(10) may attenuate mitochondrial OXPHOS capacity, which may be an underlying mechanism. METHODS Plasma glucose and insulin concentrations were measured during an oral glucose tolerance test. Mitochondrial OXPHOS capacity was measured in permeabilized muscle fibers by high-resolution respirometry in a cross-sectional design. Mitochondrial content (estimated by citrate synthase [CS] activity, cardiolipin content, and voltage-dependent anion channel [VDAC] content) as well as Q(10) content was determined. RESULTS Simvastatin-treated patients had an impaired glucose tolerance and displayed a decreased insulin sensitivity index. Regarding mitochondrial studies, Q(10) content was reduced (p = 0.05), whereas mitochondrial content was similar between the groups. OXPHOS capacity was comparable between groups when complex I- and complex II-linked substrates were used alone, but when complex I + II-linked substrates were used (eliciting convergent electron input into the Q intersection [maximal ex vivo OXPHOS capacity]), a decreased (p < 0.01) capacity was observed in the patients compared with the control subjects. CONCLUSIONS These simvastatin-treated patients were glucose intolerant. A decreased Q(10) content was accompanied by a decreased maximal OXPHOS capacity in the simvastatin-treated patients. It is plausible that this finding partly explains the muscle pain and exercise intolerance that many patients experience with their statin treatment.

Glucagon-like peptide-2 increases mesenteric blood flow in humans

Objective. Mesenteric blood flow is believed to be influenced by digestion and absorption of ingested macronutrients. We hypothesized that the intestinotrophic hormone, GLP-2 (glucagons-like peptide 2), may be involved in the regulation of mesenteric blood flow. Changes in mesenteric blood flow were measured by Doppler ultrasound scanning of the superior mesenteric artery (SMA). The aim of the study was to demonstrate the influence of GLP-2 on this flow, expressed as changes in resistance index (RI). Material and methods. A homogeneous group of 10 fasting healthy volunteers completed a 2-day trial. On day 1, a standard meal was given, and RI measured in the SMA. On day 2, GLP-2 was infused intravenously (IV) at rates of 0.5, 1.0 and 2.0 pmol/kg/min over 3×45 min separated by a 15–20 min rest period. After a further 15–20 min of rest, 450 nmol synthetic GLP-2 was given subcutaneously (SC). RI in the SMA was measured before, during and after the meal and GLP-2 infusions. Results. After IV infusion of GLP-2, the following decreases in RI were observed: 0.5 pmol/kg/min: 2.7% (range 0–6.3%), 1.0 pmol/kg/min: 6.7% (range 0.4–15.9%), 2.0 pmol/kg/min: 15.3% (range 9.6–22.7%) p<0.00802. When given SC, GLP-2 elicited a maximum average change in RI of 15.6% (range 5.0–28.1%). The standard meal elicited a 14.7% (range 8.8–21.6%) change, p<0.020 There was a similar change in RI over time (0–90 min) after a standard meal and after subcutaneous GLP-2, p<0.005. Conclusions. Our study showed a significant association between IV and SC administration of synthetic GLP-2 and changes in mesenteric blood flow. An exponential dose–response relationship was observed after IV infusion. The meal-induced changes in mesenteric blood flow over time were similar to those obtained by SC GLP-2. Thus, our results support the hypothesis that GLP-2 is an important regulator of mesenteric blood flow.

Are substrate use during exercise and mitochondrial respiratory capacity decreased in arm and leg muscle in type 2 diabetes

Aim/hypothesisThe aim of the study was to investigate mitochondrial function, fibre type distribution and substrate oxidation in arm and leg muscle during exercise in patients with type 2 diabetes and in obese and lean controls.MethodsIndirect calorimetry was used to calculate fat and carbohydrate oxidation during both progressive arm-cranking and leg-cycling exercises. Muscle biopsies from arm and leg were obtained. Fibre type, as well as O2 flux capacity of saponin-permeabilised muscle fibres were measured, the latter by high resolution respirometry, in patients with type 2 diabetes, age- and BMI-matched obese controls, and age-matched lean controls.ResultsFat oxidation was similar in the groups during either arm or leg exercise. During leg exercise at higher intensities, but not during arm exercise, carbohydrate oxidation was lower in patients with type 2 diabetes compared with the other groups. In patients with type 2 diabetes, ADP-stimulated state 3 respiration per mg muscle with parallel electron input from complex I+II was lower in m. vastus lateralis compared with obese and lean controls, whereas no differences between groups were present in m. deltoideus. A higher percentage of type IIX fibres was seen in m. vastus lateralis in patients with type 2 diabetes compared with obese and lean controls, whereas no difference was found in the deltoid muscle.Conclusions/interpretationThis study demonstrates similar O2 flux capacity, fibre type distribution and carbohydrate oxidation in arm muscle in the groups despite the presence of attenuated values in leg muscle in patients with type 2 diabetes compared with obese and lean controls.

Evidence of Extrapancreatic Glucagon Secretion in Man.

Glucagon is believed to be a pancreas-specific hormone, and hyperglucagonemia has been shown to contribute significantly to the hyperglycemic state of patients with diabetes. This hyperglucagonemia has been thought to arise from α-cell insensitivity to suppressive effects of glucose and insulin combined with reduced insulin secretion. We hypothesized that postabsorptive hyperglucagonemia represents a gut-dependent phenomenon and subjected 10 totally pancreatectomized patients and 10 healthy control subjects to a 75-g oral glucose tolerance test and a corresponding isoglycemic intravenous glucose infusion. We applied novel analytical methods of plasma glucagon (sandwich ELISA and mass spectrometry–based proteomics) and show that 29–amino acid glucagon circulates in patients without a pancreas and that glucose stimulation of the gastrointestinal tract elicits significant hyperglucagonemia in these patients. These findings emphasize the existence of extrapancreatic glucagon (perhaps originating from the gut) in man and suggest that it may play a role in diabetes secondary to total pancreatectomy.

Potentially avoidable perinatal deaths in Denmark and Sweden 1991

Background. Since 1950 the perinatal mortality has been significantly higher in Denmark than in Sweden. In 1991 the rate in Denmark was 8.0/1000 deliveries compared to 6.5/1000 in Sweden. An international audit was designed to investigate whether the perinatal death rates in the two countries to some extent could reflect differences in the quality of care, indicated by the numbers of perinatal deaths in categories of potentially avoidable deaths.

Regional Anatomic Differences in Skeletal Muscle Mitochondrial Respiration in Type 2 Diabetes and Obesity

CONTEXT Previous studies on leg skeletal musculature have demonstrated mitochondrial dysfunction associated with type 2 diabetes mellitus (T2DM), but it is not known whether mitochondrial dysfunction is present in the upper extremities. OBJECTIVE The aim of the study was to compare mitochondrial respiration and markers of mitochondrial content in skeletal muscle of arm and leg in patients with T2DM and obese control subjects. PATIENTS Ten patients with T2DM (age, 52.3 +/- 2.7 yr; body mass index, 30.1 +/- 1.2 kg/m(2)) (mean +/- se) were studied after a 2-wk washout period of oral antihyperglycemic agents. Ten control subjects (age, 54.3 +/- 2.8 yr; body mass index, 30.4 +/- 1.2 kg/m(2)) with normal fasting and 2-h oral glucose tolerance test blood glucose levels were also included. MAIN OUTCOME MEASURE We measured mitochondrial respiration in saponin-treated skinned muscle fibers from biopsies of m. deltoideus and m. vastus lateralis using high-resolution respirometry. RESULTS In the arm, mitochondrial respiration and citrate synthase activity did not differ between groups, but mitochondrial respiration per milligram of muscle was significantly higher in the leg muscle of the control subjects compared to T2DM. Fiber type compositions in arm and leg muscles were not different between the T2DM and control group, and maximum rate of O(2) consumption did not differ between the groups. CONCLUSION The results demonstrate that reduced mitochondrial function in T2DM is only present in the leg musculature. This novel finding suggests that mitochondrial dysfunction is not a primary defect affecting all skeletal muscle but could be related to a decreased response to locomotor muscle use in T2DM.

Normal mitochondrial function and increased fat oxidation capacity in leg and arm muscles in obese humans

Aim/hypothesis:The aim of this study was to investigate mitochondrial function, fibre-type distribution and substrate oxidation during exercise in arm and leg muscles in male postobese (PO), obese (O) and age- and body mass index (BMI)-matched control (C) subjects. The hypothesis of the study was that fat oxidation during exercise might be differentially preserved in leg and arm muscles after weight loss.Methods:Indirect calorimetry was used to calculate fat and carbohydrate oxidation during both progressive arm-cranking and leg-cycling exercises. Muscle biopsy samples were obtained from musculus deltoideus (m. deltoideus) and m. vastus lateralis muscles. Fibre-type composition, enzyme activity and O2 flux capacity of saponin-permeabilized muscle fibres were measured, the latter by high-resolution respirometry.Results:During the graded exercise tests, peak fat oxidation during leg cycling and the relative workload at which it occurred (FatMax) were higher in PO and O than in C. During arm cranking, peak fat oxidation was higher in O than in C, and FatMax was higher in O than in PO and C. Similar fibre-type composition was found between groups. Plasma adiponectin was higher in PO than in C and O, and plasma leptin was higher in O than in PO and C.Conclusions:In O subjects, maximal fat oxidation during exercise and the eliciting relative exercise intensity are increased. This is associated with higher intramuscular triglyceride levels and higher resting non esterified fatty acid (NEFA) concentrations, but not with differences in fibre-type composition, mitochondrial function or muscle enzyme levels compared with Cs. In PO subjects, the changes in fat oxidation are preserved during leg, but not during arm, exercise.

Leptin receptor 170 kDa (OB-R170) protein expression is reduced in obese human skeletal muscle: a potential mechanism of leptin resistance

To examine whether obesity‐associated leptin resistance could be due to down‐regulation of leptin receptors (OB‐Rs) and/or up‐regulation of suppressor of cytokine signalling 3 (SOCS3) and protein tyrosine phosphatase 1B (PTP1B) in skeletal muscle, which blunt janus kinase 2‐dependent leptin signalling and signal transducer and activator of transcription 3 (STAT3) phosphorylation and reduce AMP‐activated protein kinase (AMPK) and acetyl‐coenzyme A carboxylase (ACC) phosphorylation. Deltoid and vastus lateralis muscle biopsies were obtained from 20 men: 10 non‐obese control subjects (mean ±s.d. age, 31 ± 5 years; height, 184 ± 9 cm; weight, 91 ± 13 kg; and percentage body fat, 24.8 ± 5.8%) and 10 obese (age, 30 ± 7 years; height, 184 ± 8 cm; weight, 115 ± 8 kg; and percentage body fat, 34.9 ± 5.1%). Skeletal muscle OB‐R170 (OB‐R long isoform) protein expression was 28 and 25% lower (both P < 0.05) in arm and leg muscles, respectively, of obese men compared with control subjects. In normal‐weight subjects, SOCS3 protein expression, and STAT3, AMPKα and ACCβ phosphorylation, were similar in the deltoid and vastus lateralis muscles. In obese subjects, the deltoid muscle had a greater amount of leptin receptors than the vastus lateralis, whilst SOCS3 protein expression was increased and basal STAT3, AMPKα and ACCβ phosphorylation levels were reduced in the vastus lateralis compared with the deltoid muscle (all P < 0.05). In summary, skeletal muscle leptin receptors and leptin signalling are reduced in obesity, particularly in the leg muscles.

Three‐dimensional reconstruction of the human skeletal muscle mitochondrial network as a tool to assess mitochondrial content and structural organization

Mitochondria undergo continuous changes in shape as result of complex fusion and fission processes. The physiological relevance of mitochondrial dynamics is still unclear. In the field of mitochondria bioenergetics, there is a need of tools to assess cell mitochondrial content. To develop a method to visualize mitochondrial networks in high resolution and assess mitochondrial volume.