Holger Krakowski-Roosen

Pancreatic cancerrelated cachexia: influence on metabolism and correlation to weight loss and pulmonary function

BackgroundDramatic weight loss is an often underestimated symptom in pancreatic cancer patients. Cachexia- defined as an unintended loss of stable weight exceeding 10% – is present in up to 80% of patients with cancer of the upper gastrointestinal tract, and has a significant influence on survival. The aim of the study was to show the multiple systemic effects of cachexia in pancreatic cancer patients, in terms of resection rate, effects on pulmonary function, amount of fat and muscle tissue, as well as changes in laboratory parameters.MethodsIn patients with pancreatic cancer, clinical appearance was documented, including the amount of weight loss. Laboratory parameters and lung-function tests were evaluated, and the thickness of muscle and fat tissue was measured with computed tomography scans. Statistical analysis, including multivariate analysis, was performed using SPSS software. Survival curves were calculated using Kaplan-Meier analysis and the log-rank test. To test for significant differences between the examined groups we used Students t-test and the Mann-Whitney U test. Significance was defined as p < 0.05.ResultsOf 198 patients with a ductal adenocarcinoma of the pancreas, 70% were suffering from weight loss when they presented for operation, and in 40% weight loss exceeded 10% of the stable weight. In patients with cachexia, metastases were diagnosed significantly more often (47% vs. 24%, P < 0.001), leading to a significantly reduced resection rate in these patients. Patients with cachexia had significantly reduced fat tissue amounts. Hence, dramatic weight loss in a patient with pancreatic cancer may be a hint of a more progressed or more aggressive tumour.ConclusionPancreatic cancer patients with cachexia had a higher rate of more progressed tumour stages and a worse nutritional status. Furthermore, patients with cachexia had an impaired lung function and a reduction in fat tissue. Patients with pancreatic cancer and cachexia had significantly reduced survival. If weight loss exceeded 5% there was a significantly reduced resection rate to detect, but the changes were significantly more substantial if weight loss was 10% or more. We propose that a weight loss of 10% be defined as significant in pancreatic cancer.

Morphology, metabolism, microcirculation, and strength of skeletal muscles in cancer-related cachexia

Purpose. Cancer-related cachexia is an obscure syndrome leading to muscle wasting, reduced physical fitness and quality of life. The aim of this study was to assess morphology, metabolism, and microcirculation in skeletal muscles of patients with cancer-related cachexia and to compare these data with matched healthy volunteers. Methods. In 19 patients with cancer-induced cachexia and 19 age-, gender-, and body-height-matched healthy volunteers body composition and aerobic capacity (VO2max) were analyzed. Skeletal muscle fiber size and capillarization were evaluated in biopsies of the vastus lateralis muscle. The cross-sectional area (CSA) of the quadriceps femoris muscle was measured by magnetic resonance imaging as well as its isokinetic and isometric force. The energy and lipid metabolism of the vastus lateralis muscle was quantified by 31P and 1H spectroscopy and parameters of its microcirculation by contrast-enhanced ultrasonography (CEUS). Results. Morphologic parameters were about 30% lower in cachexia than in volunteers (body mass index: 20±3 vs. 27±4 kg m−2, CSA: 45±13 vs. 67±14 cm2, total fiber size: 2854±1112 vs. 4181±1461 µm2). VO2max was reduced in cachexia (23±9 vs. 32±7 ml min−1 kg−1, p=0.03), whereas histologically determined capillary density and microcirculation in vivo were not different. Both concentrations of muscular energy metabolites, pH, and trimethyl-ammonium-containing compounds were comparable in both groups. Absolute strength of quadriceps muscle was reduced in cachexia (isometric: 107±40 vs. 160±40 Nm, isokinetic: 101±46 vs. 167±50 Nm; p=0.03), but identical when normalized on CSA (isometric: 2.4±0.5 vs. 2.4±0.4 Nm cm−2, isokinetic: 2.2±0.4 vs. 2.5±0.5 Nm cm−2). Conclusions. Cancer-related cachexia is associated with a loss of muscle volume but not of functionality, which can be a rationale for muscle training.

Assessment of Skeletal Muscle Perfusion Using Contrast-Enhanced Ultrasonography

The purpose of this study was to develop a clinically applicable examination method to assess perfusion of the skeletal muscle using contrast‐enhanced ultrasonography (CEUS) analyzing replenishment kinetics of microbubbles.

Effect of thiol antioxidant on body fat and insulin reactivity.

Insulin signaling is enhanced by moderate concentrations of reactive oxygen species (ROS) and suppressed by persistent exposure to ROS. Diabetic patients show abnormally high ROS levels and a decrease in insulin reactivity which is ameliorated by antioxidants, such as N-acetylcysteine (NAC). A similar effect of NAC has not been reported for non-diabetic subjects. We now show that the insulin receptor (IR) kinase is inhibited in cell culture by physiologic concentrations of cysteine. In two double-blind trials involving a total of 140 non-diabetic subjects we found furthermore that NAC increased the HOMA-R index (derived from the fasting insulin and glucose concentrations) in smokers and obese patients, but not in nonobese non-smokers. In obese patients NAC also caused a decrease in glucose tolerance and body fat mass. Simultaneous treatment with creatine, a metabolite utilized by skeletal muscle and brain for the interconversion of ADP and ATP, reversed the NAC-mediated increase in HOMA-R index and the decrease in glucose tolerance without preventing the decrease in body fat. As the obese and hyperlipidemic patients had lower plasma thiol concentrations than the normolipidemic subjects, our results suggest that low thiol levels facilitate the development of obesity. Supplementation of thiols plus creatine may reduce body fat without compromising glucose tolerance.

Real-time contrast-enhanced ultrasound for the assessment of perfusion dynamics in skeletal muscle.

We developed a real-time low-MI contrast-enhanced ultrasound method (CEUS), compared it with venous occlusion plethysmography (VOP) and evaluated its robustness in the quantification of skeletal muscle perfusion during exercise. Contrast pulse sequencing (7 MHz) during continuous intravenous infusion of SonoVue (4.8 mL/300 s) was used repeatedly in eight healthy volunteers to monitor changes of the muscle perfusion before, during and after isometric exercises (10 to 50% of individual maximum strength for 20 to 30 s) of the gastrocnemius muscle in real time. CEUS was correlated with VOP at different time points, and the exactness of several CEUS parameters obtained from ultrasound-signal-intensity-time curves was evaluated. Real-time CEUS depicted a large variability of the skeletal muscle blood volume at rest (mean, 3.48; range, 0.60 to 9.92 [approximately mL]), with a significant reproducibility (r=0.72, p<0.05) and correlation with VOP (r=0.59, p<0.001). Mean blood volume during exercise was 1.58(approximately mL), increased to a mean maximum after exercise of 8.88 (approximately mL), the mean change of the local blood volume during and directly after the exercise was -0.10 and +1.57(approximately mL/s). The average CEUS signal during exercise decreased (mean area under the curve, -50.4 [approximately mL.s]) and subsequently increased post exercise (mean 118.6 [approximately mL.s]). CEUS parameters could be calculated with mean relative errors between 6 and 36%. Continuous assessment of local muscle microcirculation during exercise is possible with real-time CEUS with an acceptable robustness. Its application may be of particular interest in a better understanding of the role of perfusion during muscle training, and the monitoring of pathological vascular response, such as in diabetic microvessel diseases.

Comparison of transient arterial occlusion and muscle exercise provocation for assessment of perfusion reserve in skeletal muscle with real-time contrast-enhanced ultrasound.

OBJECTIVE Contrast-enhanced ultrasound (CEUS) is able to quantify muscle perfusion and changes in perfusion due to muscle exercise in real-time. However, reliable measurement of standardized muscle exercise is difficult to perform in clinical examinations. We compared perfusion reserve assessed by CEUS after transient arterial occlusion and exercise to find the most suitable measurement for clinical application. METHODS Contrast pulse sequencing (7 MHz) during continuous IV infusion of SonoVue(®) (4.8 mL/300 s) was used in 8 healthy volunteers to monitor muscle perfusion of the gastrocnemius muscle during transient (1 min) arterial occlusion produced by a thigh cuff of a venous occlusion plethysmograph. Isometric muscle exercise (50% of individual maximum strength for 20s) was subsequently performed during the same examination, and several CEUS parameters obtained from ultrasound-signal-intensity-time curves and its calculation errors were compared. RESULTS The mean maximum local blood volume after occlusion was 13.9 [∼mL] (range, 4.5-28.8 [∼mL]), and similar values were measured after sub-maximum exercise 13.8 [∼mL], (range, 4.6-22.2 [∼mL]. The areas under the curve during reperfusion vs. recovery were also similar (515.2±257.5 compared to 482.2±187.5 [∼mLs]) with a strong correlation (r=0.65), as were the times to maximum (15.3s vs. 15.9s), with a significantly smaller variation for the occlusion method (±2.1s vs. ±9.0s, p=0.03). The mean errors for all calculated CEUS parameters were lower for the occlusion method than for the exercise test. CONCLUSIONS CEUS muscle perfusion measurements can be easily performed after transient arterial occlusion. It delivers data which are comparable to CEUS measurements after muscle exercise but with a higher robustness. This method can be easily applied in clinical examination of patients with e.g. PAOD or diabetic microvessel diseases to assess perfusion reserve.

Changes in the micro-circulation of skeletal muscle due to varied isometric exercise assessed by contrast-enhanced ultrasound.

PURPOSE To quantitatively assess local muscle micro-circulation with real-time contrast-enhanced ultrasound (CEUS) during different exercises and compare the results with performed muscle work and global blood flow. MATERIALS AND METHODS Sixteen low mechanical index CEUS examinations of the right lower leg flexors of healthy volunteers were performed using a continuous infusion of SonoVue(®) (4.8 mL/300 s). Several muscle perfusion parameters were extracted from derived CEUS signal intensity time curves during different isometric exercises (10-50% of maximum individual strength for 20-30s) and then correlated with the performed muscle work or force, and the whole lower leg blood flow which we measured simultaneously by venous occlusion plethysmography (VOP). RESULTS The shapes of the CEUS curve during and after exercise differed individually depending on the performed muscle work. The maximum blood volume MAX was observed only after exercise cessation and was significantly correlated with the performed muscle force (r=0.77, p<0.0001). The blood volume over exercise time was inversely correlated with the spent muscle work (r=-0.60, p=0.006). CEUS and VOP measurements correlated only at rest and after the exercise. During exercise, mean CEUS local blood volume decreased (from 3.48 to 2.19 (∼mL)), while mean VOP global blood flow increased (mean, from 3.96 to 7.71 mL/100 mg/min). CONCLUSION Real-time low-MI CEUS provides complementary information about the local muscle micro-circulation compared to established blood flow measures. CEUS may be used for a better understanding of muscle perfusion physiology and in the diagnosis of micro-circulation alterations such as in peripheral arterial occlusive disease or diabetic angiopathy.

Dynamic Contrast-enhanced Ultrasound for Assessment of Skeletal Muscle Microcirculation in Peripheral Arterial Disease

Objective:This feasibility study was performed to assess whether dynamic contrast-enhanced ultrasound (CEUS) and transient arterial occlusion are able to detect alterations in the microvascular perfusion and arterial perfusion reserve in patients suffering from peripheral arterial disease (PAD) in comparison with healthy volunteers. Materials and Methods:Twenty patients with PAD, Rutherford classification grade I, category III (mean age, 64 years; mean height, 173 cm; mean weight, 81.8 kg), and 20 volunteers (mean age, 50 years; mean height, 174 cm; mean weight, 77.8 kg) participated in the study. Low-mechanical index CEUS (7 MHz; MI, 0.28) was performed to the dominant lower leg after start of a continuous automatic intravenous injection of 4.8 mL suspension with microbubbles containing sulfur hexafluoride (SonoVue) within 5 minutes. Perfusion of the calf muscle was monitored by CEUS before, during, and after release of arterial occlusion at the thigh level lasting for 60 seconds. Several parameters, especially the time to maximum enhancement after release of occlusion (tmax), the maximum enhancement after release of occlusion (maxenh), the total vascular response after release of occlusion (AUCpost), and the resulting slope (m2) to maximum enhancement were calculated. Results:After release of the occlusion, a significantly delayed increase of the CEUS signal to maxenh was observed in the patients with PAD (32 ± 17 seconds) compared with volunteers (17 ± 8 seconds, P = 0.0009). maxenh was 66.5 ± 36.6 (∼mL) in PAD versus 135.6 ± 75.1 (∼mL) in volunteers (P = 0.0016). AUCpost was 3016.5 ± 1825.8 (∼mL·s) in PAD versus 5906.4 ± 3173.1 (∼mL·s) in volunteers (P = 0.0013), and m2 was significantly lower in PAD (3.8 ± 5.2 vs. 14.8 ± 9.7 [∼mL/s], P = 0.0001). Conclusions:Microvascular perfusion deficits and reduced arterial perfusion reserve in patients with PAD are clearly detectable with dynamic CEUS after transient arterial occlusion.

Concentric resistance training increases muscle strength without affecting microcirculation

PURPOSE While the evidence is conclusive regarding the positive effects of endurance training, there is still some controversy regarding the effects of resistance training on muscular capillarity. Thus, the purpose was to assess whether resistance strength training influences resting skeletal muscle microcirculation in vivo. MATERIALS AND METHODS Thirty-nine middle-aged subjects (15 female, 24 male; mean age, 54+/-9 years) were trained twice a week on an isokinetic system (altogether 16 sessions lasting 50 min, intensity 75% of maximum isokinetic and isometric force of knee flexors and extensors). To evaluate success of training, cross-sectional area (CSA) of the quadriceps femoris muscle and its isokinetic and isometric force were quantified. Muscular capillarization was measured in biopsies of the vastus lateralis muscle. In vivo, muscular energy and lipid metabolites were quantified by magnetic resonance spectroscopy and parameters of muscular microcirculation, such as local blood volume, blood flow and velocity, by contrast-enhanced ultrasound analyzing replenishment kinetics. RESULTS The significant (P<0.001) increase in CSA (60+/-16 before vs. 64+/-15 cm(2) after training) and in absolute muscle strength (isometric, 146+/-44 vs. 174+/-50 Nm; isokinetic, 151+/-53 vs. 174+/-62 Nm) demonstrated successful training. Neither capillary density ex vivo (351+/-75 vs. 326+/-62) nor ultrasonographic parameters of resting muscle perfusion were significantly different (blood flow, 1.2+/-1.2 vs. 1.1+/-1.1 ml/min/100g; blood flow velocity, 0.49+/-0.44 vs. 0.52+/-0.74 mms(-1)). Also, the intensities of high-energy phosphates phosphocreatine and beta-adenosintriphosphate were not different after training within the skeletal muscle at rest (beta-ATP/phosphocreatine, 0.29+/-0.06 vs. 0.28+/-0.04). CONCLUSION The significant increase in muscle size and strength in response to concentric isokinetic and isometric resistance training occurs without an increase in the in vivo microcirculation of the skeletal muscles at rest.

Real-imaging cDNA-AFLP transcript profiling of pancreatic cancer patients: Egr-1 as a potential key regulator of muscle cachexia

BackgroundCancer cachexia is a progressive wasting syndrome and the most prevalent characteristic of cancer in patients with advanced pancreatic adenocarcinoma. We hypothesize that genes expressed in wasted skeletal muscle of pancreatic cancer patients may determine the initiation and severity of cachexia syndrome.Experimental designWe studied gene expression in skeletal muscle biopsies from pancreatic cancer patients with and without cachexia utilizing Real-Imaging cDNA-AFLP-based transcript profiling for genome-wide expression analysis.ResultsOur approach yielded 183 cachexia-associated genes. Ontology analysis revealed characteristic changes for a number of genes involved in muscle contraction, actin cytoskeleton rearrangement, protein degradation, tissue hypoxia, immediate early response and acute-phase response.ConclusionsWe demonstrate that Real-Imaging cDNA-AFLP analysis is a robust method for high-throughput gene expression studies of cancer cachexia syndrome in patients with pancreatic cancer. According to quantitative RT-PCR validation, the expression levels of genes encoding the acute-phase proteins α-antitrypsin and fibrinogen α and the immediate early response genes Egr-1 and IER-5 were significantly elevated in the skeletal muscle of wasted patients. By immunohistochemical and Western immunoblotting analysis it was shown, that Egr-1 expression is significantly increased in patients with cachexia and cancer. This provides new evidence that chronic activation of systemic inflammatory response might be a common and unifying factor of muscle cachexia.