Increased levels of apoptosis in gastrocnemius skeletal muscle in patients with peripheral arterial disease

Intermittent claudication (IC) is the major clinical manifestation of peripheral arterial disease (PAD). Apoptosis has been linked to skeletal muscle pathophysiology in other chronic diseases such as congestive heart failure. This study tested the hypothesis that there would be increased levels of apoptosis in the skeletal muscle of patients with PAD compared with control individuals. In total, 26 individuals with PAD and 28 age-appropriate controls underwent studies of peak oxygen consumption (peak VO2) and a gastrocnemius muscle biopsy in the most symptomatic leg. Muscle biopsies were analyzed for apoptosis and caspase-3 activity. Patients with PAD had a reduced peak VO2 compared with controls. Apoptosis was increased in those with PAD compared with age-appropriate controls (3.83% +/- 2.6 vs 1.53% +/- 0.96; p < 0.001). In conclusion, PAD is associated with increased levels of apoptosis in the peripheral skeletal muscle. Further study is required to ascertain whether apoptosis plays a role in decreased functional capacity.     

Increased mitochondrial DNA deletions in the skeletal muscle of myotonic dystrophy.

Mitochondrial abnormality in the skeletal muscles of 13 patients with myotonic dystrophy was analyzed by both histochemical and molecular biologic methods. Nine of 13 patients had ragged-red fibers (50 +/- 116 per 10,000 muscle fibers, mean +/- SD), and 10 patients had cytochrome c oxidase-negative fibers (41 +/- 90 per 10,000 muscle fibers). Southern blot analysis detected no mitochondrial DNA deletions, while PCR revealed multiple mitochondrial DNA deletions in all the specimens. Direct sequencing of one of the deleted mitochondrial DNAs disclosed that the junctional sequence of a 3,460-bp deletion involved a 6-bp directly repeated sequence (5'-TAGAAG-3') flanked by C-rich regions located on the CO3 gene and the ND5 gene. Quantitative analysis of PCR amplified deleted mitochondrial DNAs revealed that the amount of deleted mitochondrial DNAs had positive correlation both with the frequencies of ragged-red fibers and cytochrome c oxidase-negative fibers. Although deleted mitochondrial DNAs were observed even in controls above age 30, the mean amount of deleted mitochondrial DNAs in patients with myotonic dystrophy was significantly higher than in controls. Moreover, the increase of deleted mitochondrial DNAs with aging was more marked in myotonic dystrophy than in controls. These results suggest that increased mitochondrial DNA deletions and consequent impairment of mitochondrial function contribute to the pathophysiology of myotonic dystrophy.     

Acquired skeletal muscle metabolic myopathy in atherosclerotic peripheral arterial disease

Peripheral arterial disease (PAD) is associated with an increased risk of overall cardiovascular mortality, and substantial morbidity resulting from claudication. While the initial disease process is clearly the result of atherosclerosis in the arterial circulation of the limb, altered hemodynamics do not completely explain the pathophysiology of claudication. Work from several laboratories has demonstrated secondary changes in the skeletal muscle of patients with PAD which are consistent with the presence of an acquired metabolic myopathy in these patients. Key findings include an alteration in the expression of mitochondrial enzymes, the accumulation of metabolic intermediates, altered regulation of mitochondrial respiration, increased oxidative stress, and the presence of somatic mutations in the mitochondrial genome. Understanding the metabolic changes associated with PAD is important in understanding the pathophysiology of claudication and in the development of novel therapeutic strategies.     

High-energy phosphate metabolism in the exercising muscle of patients with peripheral arterial disease

Patients with peripheral arterial disease (PAD) suffer from impaired muscle function due to insufficient oxygen supply during exercise, mitochondrial damages, unfavourable muscle fibre type distribution and impaired exercise tolerance. These factors influence the symptoms as well as the quality of life in PAD patients and are closely connected to failures of high-energy phosphate metabolism. At onset of muscle exercise, the mitochondrial capacity cannot match the increased demand. The oxygen supply via blood flow must be increased. Meanwhile, anaerobic glycolysis and internal stores of oxygen like mixed venous blood and myoglobin as well as internal stores of high-energy phosphates like phosphocreatine (PCr) are adducted for the provision of additional adenosine-triphosphate (ATP), which is consumed by the ATPase at the myofibrils in order to fuel muscle contraction. Since the ATP production is insufficient, this phase (anaerobic phase) is characterized by a progressive decrease in PCr, which can be accurately measured by phosphorus 31 magnetic resonance spectroscopy (³¹p MRS). If the oxygen supply is improved, the mitochondrial capacity can match the increased metabolic demand. This phase is the aerobic phase, which is indicated by a steady-state of PCr hydrolysis. In PAD patients or experimental models of peripheral ischemia, the anaerobic phase is prolonged or does not pass into the aerobic phase resulting in exercise abortion. This review summarizes the results of ³¹p MRS studies investigating the high-energy phosphate metabolism during ischemic exercise in healthy humans and during ramp or incremental exercise in PAD patients.     

Oxidative-stress-mediated arterial dysfunction in patients with peripheral arterial disease.

AIMS: To investigate the existence of a relationship among flow-mediated dilation (FMD), nitric oxide (NO), and oxidative stress in patients with peripheral arterial disease (PAD), and to assess if the administration of an antioxidant was able to improve arterial dilatation. METHODS AND RESULTS: We performed a cross-sectional study comparing FMD, 8-Hydroxy-2-deoxy-2-deoxyguanosine (8-OHdG), a marker of oxidative stress, and nitrite/nitrate (NOx) serum levels in a population of 25 PAD patients and 40 controls. In the second part of the study, 21 PAD patients were randomly allocated to a treatment sequence of 7 days of i.v. infusion of placebo or 6 g/day propionyl-L-carnitine (PLC) in a cross-over design. Compared with controls, patients with PAD had enhanced 8-OHdG serum levels (2.4 +/- 1.2 vs. 4.24 +/- 3.11 ng/mL; P < 0.001), reduced NOx (17.02 +/- 6.11 vs. 11.28 +/- 6.02 microM; P < 0.001), and lowered FMD (10.34 +/- 2.14 vs. 6.69 +/- 2.95; P < 0.001). PLC infusion was associated with an increase of FMD [from 6.6 +/- 0.6 to 11.1 +/- 1.2% (mean +/- SE), P = 0.004] and NOx (from 14.5 +/- 1.4 to 17.1 +/- 1.2 microM; +18%, P = 0.012) and a decrease of 8-OHdG (from 3.62 +/- 0.37 to 2.64 +/- 0.32 ng/mL; -27%, P < 0.001). No changes were observed after placebo treatment. CONCLUSION: This study shows that in PAD patients, oxidative stress is implicated in determining reduced FMD.     

Treating peripheral arterial disease in patients with diabetes

Diabetes is associated with considerably higher risks of developing peripheral arterial disease (PAD) which, when it occurs, is more severe and progresses more rapidly than in nondiabetics. Early detection of PAD in the diabetic patient is therefore important, but may be complicated by the presence of neuropathy and calcification of the arteries such that ischaemic symptoms are not felt by the patient and ankle pressures are not reduced. Toe pressures are an alternative diagnostic tool in these patients. Good glycaemic control, while an essential part of diabetes management, does not appear to bring more than modest benefits in preventing the peripheral vascular complications of diabetes. Therefore, attention to other risk factors is needed. Treatment with the phosphodiesterase III inhibitor, cilostazol, has been shown to improve walking distances significantly in diabetes patients with intermittent claudication and also appears to improve plasma lipid profiles. Further, cilostazol has an antiplatelet action, which may prove to be of benefit in diabetes because hyperglycaemia is associated with increased platelet aggregability. Revascularization in diabetes patients with critical leg ischaemia is complex and associated with poorer outcomes than in non-diabetes patients. While surgical revascularization has better patency rates, in patients at high risk of surgical complications, percutaneous transluminal angioplasty may be a better option.     

DNA polymorphisms of the gene for apolipoprotein B in patients with peripheral arterial disease

We have determined the frequency of DNA polymorphisms of the gene for human apolipoprotein B, detected with XbaI and EcoRI, in 205 patients with documented peripheral arterial disease. Of the patients, 78 have no evidence of disease in the coronary and carotid arteries, 64 have coexisting coronary artery disease but no evidence of carotid artery disease, 26 patients have coexisting carotid artery disease but no evidence of coronary artery disease, and 37 have coexisting coronary and carotid artery disease. Levels of triglycerides, cholesterol and apolipoprotein B were measured for each patient, and RFLP frequency was determined in all the patients. Lipid, lipoprotein and apolipoprotein levels were not significantly different between the different patient groups. Compared with a sample from the clinically well London population, the frequency of the R2 allele of the polymorphism detected with EcoRI, and the frequency of the X1 allele of the XbaI polymorphism was significantly higher in the patient group. The frequency of these alleles was not significantly different in the different patient groups. In patients with only peripheral arterial disease, individuals with the XbaI genotype X1X1 have the lowest and those with the genotype X2X2 have the highest mean levels of serum cholesterol. However, in all other patient groups this trend was reversed (X1X1 highest and X2X2 lowest). Our observations suggest that variation at the apo B locus is one of the factors involved in predisposing an individual to develop arterial disease but does not determine where in the arterial system the disease develops.     

Morphological observations in skeletal muscle from patients with a mitochondrial myopathy

The regional distribution of the phenylalanine-sensitive ATP-sulphurylase in fetal calf brain coincides with demyelinated lesions observed in the central nervous systems of untreated PKU patients. This would be expected if this species of ATP-sulphurylase played a role in the pathogenesis of brain dysfunction in the untreated or poorly controlled phenylketonuria patient.     

Lower extremity ischemia, calf skeletal muscle characteristics, and functional impairment in peripheral arterial disease

OBJECTIVES: To determine whether lower ankle brachial index (ABI) levels are associated with lower calf skeletal muscle area and higher calf muscle percentage fat in persons with and without lower extremity peripheral arterial disease (PAD). DESIGN: Cross-sectional. SETTING: Three Chicago-area medical centers. PARTICIPANTS: Four hundred thirty-nine persons with PAD (ABI<0.90) and 265 without PAD (ABI 0.90-1.30). MEASUREMENTS: Calf muscle cross-sectional area and the percentage of fat in calf muscle were measured using computed tomography at 66.7% of the distance between the distal and proximal tibia. Physical activity was measured using an accelerometer. Functional measures included the 6-minute walk, 4-meter walking speed, and the Short Physical Performance Battery (SPPB). RESULTS: Adjusting for age, sex, race, comorbidities, and other potential confounders, lower ABI values were associated with lower calf muscle area (ABI<0.50, 5,193 mm(2); ABI 0.50-0.90, 5,536 mm(2); ABI 0.91-1.30, 5,941 mm(2); P for trend<.001). These significant associations remained after additional adjustment for physical activity. In participants with PAD, lower calf muscle area in the leg with higher ABI was associated with significantly poorer performance in usual- and fast-paced 4-meter walking speed and on the SPPB, adjusting for ABI, physical activity, percentage fat in calf muscle, muscle area in the leg with lower ABI, and other confounders (P<.05 for all comparisons). CONCLUSION: These data support the hypothesis that lower extremity ischemia has a direct adverse effect on calf skeletal muscle area. This association may mediate previously established relationships between PAD and functional impairment.     

Elevated osteopontin levels in patients with peripheral arterial disease

This study was carried out to compare concentrations of osteopontin (OPN) and osteoprotegerin (OPG) in peripheral arterial disease (PAD). The study population consisted of 200 consecutive subjects in whom both OPN/OPG and ankle-brachial index were measured. It was found that OPN levels, but not OPG levels, were significantly more increased in patients with PAD than those without PAD. Serum OPN levels were significantly lower in subjects with angiotensin converting enzyme inhibitors or angiotensin II receptor blockers than those without these agents. In this study, it has been demonstrated for the first time that serum OPN levels are related to PAD. Inhibition of renin- angiotensin system could decrease OPN levels and prevent the progression of PAD.     

Haemodilution in patients with peripheral arterial occlusive disease.

Haemodilution is an efficient conservative therapy of peripheral arterial occlusive disease. Already a single isovolaemic haemodilution (replacement of 500 ml blood for Haes* 0.5, 10%) increases the pain-free walking distance by 85%. These effects can be maintained by a constant therapy over six weeks and following haemodilution once or twice per month. The haematocrit values should be between 38 and 42%. The haemodilution should be done hyper- or isovolaemically. Not more than 250 ml blood and 500 ml Haes should be infused during one session in order to avoid hypovolaemia. This means an infusion of 250 ml Haes, venesection of 250 ml blood via the same access and then infusion of the remaining 250 ml. The whole procedure should not last more than one hour. Blood pressure, heart rate, lung auscultation and percussion as well as creatinine values has to be controlled during an intensive therapy. If the hydroxyethyl starch concentration exceeds 150 g per week pruritus may occur in singular cases, if the concentration exceeds 700 g per week it is observed in 50% of the cases. Provided the preventive measures are observed haemodilution is an efficient and good therapy which also increases the compliance to practice vascular exercise.     

Sex-related differences in Japanese patients with peripheral arterial disease

Objective

The goal of the study was to examine possible sex-related differences in the clinical characteristics and risk factors in Japanese patients with peripheral arterial disease (PAD).

Methods

Sex-related differences in clinical profiles, risk factors and treatments were examined in 730 consecutive patients with PAD (148 women (20.3%) and 582 men (79.7%)).

Results

The mean age of the women was higher than that of the men (73.6 ± 11.2 vs. 70.9 ± 9.1 years old, p = 0.002) and the proportion of subjects aged ≥75 years old was also higher in women (P = 0.005). Women more frequently had critical limb ischemia (P < 0.001) and diabetes mellitus (P = 0.026), but less frequent smoking and alcohol intake, compared to men (P < 0.001). Total cholesterol (P < 0.001) and LDL cholesterol (P = 0.014) were higher in women. Fontaine stages were correlated with age, diabetes, cerebral infarction and women (p < 0.001). The prevalence of iliac artery lesions was higher in men (p < 0.001), whereas that for below the knee lesions was higher in women (p < 0.001). The number of affected below the knee arteries was also higher in women than in men (p < 0.001). The prevalence of medical treatment was higher in women (P = 0.009) and major amputation tended to be higher in women (p = 0.056).

Conclusions

Women had more severe symptomatic states and uncontrolled risk factors. The prevalence of iliac artery lesions was lower, but below the knee lesions were more severe in women.     

Arteriolization of capillaries and FGF-2 upregulation in skeletal muscles of patients with chronic peripheral arterial disease

OBJECTIVE: Microvascular changes in ischemic skeletal muscle are described derived from patients with long-lasting peripheral arterial disease (PAD). METHODS: Skeletal muscles from the lower limb of 17 patients (obtained after amputation) with chronic PAD and 4 asymptomatic controls (obtained from biopsies after bypass surgery) were evaluated by electron microscopy and immunohistochemistry. RESULTS: The capillaries in skeletal muscles of PAD patients were surrounded by a more than 1 microm-thick coat, which was positively stained for basement membrane pericapillary coat collagen type IV. Thickness of the coat correlated with presence of PAD (p < .0001), and less strongly with diabetes mellitus (p = .023) and age of patients (p = .019). The majority of the capillaries in skeletal muscles of PAD patients (71.1 +/- 15.3%) were covered with cells positive for smooth muscle cell actin (sma) as compared to samples from asymptomatic controls (22.8% +/- 9.6%; p < .0001) suggesting advanced arteriolization. Semiquantitative analysis revealed that patients with PAD demonstrate a higher expression of FGF-2 in capillary endothelial cells (67.8 +/- 17.5%) as compared to controls (10.2 +/- 8.4%; p < .0001), whereas VEGF immunoreactivity was only occasionally present in extravascular cells. CONCLUSION: Thickened collagen type IV-positive basement membranes in combination with a significant increase in sma-coverage indicate arteriolization of capillaries characteristic for chronic PAD, what may be related to high FGF-2 expression in capillary endothelial cells.     

Altered antioxidant status in peripheral skeletal muscle of patients with COPD

Despite the growing field of interest in the role of pulmonary oxidative stress in chronic obstructive pulmonary disease (COPD), barely any data are available with respect to antioxidant capacity in the peripheral musculature of these patients. The main objective of this study was to assess in detail the antioxidant status in skeletal muscle of patients with COPD. Biopsies from the vastus lateralis of 21 patients with COPD and 12 healthy age-matched controls were analysed. Total antioxidant capacity, vitamin E, glutathione, and uric acid levels were determined and the enzyme activities of superoxide dismutase, glutathione reductase, glutathione peroxidase, and glutathione-S-transferase were measured. Malondialdehyde was measured as an index of lipid peroxidation. The total antioxidant capacity and the uric acid levels were markedly higher in COPD patients than in healthy controls (25%, P = 0.006 and 24%, P = 0.029, respectively). Glutathione-S-transferase activity was also increased (35%; P = 0.044) in patients compared to healthy subjects. Vitamin E level was lower in patients than in controls (P < 0.05). The malondialdehyde level was not different between the two groups. It can be concluded that the muscle total antioxidant capacity is increased in patients with COPD. Together with the reduced vitamin E levels, the increased glutathione-S-transferase activity and normal levels of lipid peroxidation products, these findings suggest that the antioxidant system may be exposed to and subsequently triggered by elevated levels of reactive oxygen species.