The role of zinc in the anti-tumour and anti-cachectic activity of D-myo-inositol 1,2,6-triphosphate

Background: D-myo-inositol-1,2,6-triphosphate (a-trinositol, AT) is a polyanionic molecule capable of chelating divalent metal ions with anti-tumour and anti-cachectic activity in a murine model. Methods: To investigate the role of zinc in this process, mice bearing cachexia-inducing MAC16 tumour were treated with AT, with or without concomitant administration of ZnSO4. Results: At a dose of 40mgkg-1, AT effectively attenuated both weight loss and growth of the MAC16 tumour, and both effects were attenuated by co-administration of Zn2+. The concentration of zinc in gastrocnemius muscle increased with increasing weight loss, whereas administration of AT decreased the levels of zinc in plasma, skeletal muscle and tumour, which were restored back to control values after administration of ZnSO4. Conclusion: These results suggest that zinc is important in both tumour growth and cachexia in this animal model.

Role of β-adrenergic receptors in the anti-obesity and anti-diabetic effects of zinc-α2-glycoprotien (ZAG)

Objectives: The goal of the current study is to determine whether the ß-adrenoreceptor (ß-AR) plays a role in the anti-obesity and anti-diabetic effects of zinc-a2-glycoprotein (ZAG). Material and methods: This has been investigated in CHO-K1 cells transfected with the human ß1-, ß2-, ß3-AR and in ob/ob mice. Cyclic AMP assays were carried out along with binding studies. Ob/ob mice were treated with ZAG and glucose transportation and insulin were examined in the presence or absence of propranolol. Results: ZAG bound to the ß3-AR with higher affinity (Kd 46±1nM) than the ß2-AR (Kd 71±3nM) while there was no binding to the ß1-AR, and this correlated with the increases in cyclic AMP in CHO-K1 cells transfected with the various ß-AR and treated with ZAG. Treatment of ob/ob mice with ZAG increased protein expression of ß3-AR in gastrocnemius muscle, and in white and brown adipose tissues, but had no effect on expression of ß1- and ß2-AR. A reduction of body weight was seen and urinary glucose excretion, increase in body temperature, reduction in maximal plasma glucose and insulin levels in the oral glucose tolerance test, and stimulation of glucose transport into skeletal muscle and adipose tissue, were completely attenuated by the non-specific ß-AR antagonist propranolol. Conclusion: The results suggest that the effects of ZAG on body weight and insulin sensitivity in ob/ob mice are manifested through a ß-3AR, or possibly a ß2-AR.

Mechanism of induction of muscle protein loss by hyperglycaemia

Treatment of murine myotubes with high glucose concentrations (10 and 25 mM) stimulated protein degradation through the ubiquitin–proteasome pathway, and also caused activation (autophosphorylation) of PKR (double-stranded-RNA-dependent protein kinase) and eIF2a (eukaryotic initiation factor 2a). Phosphorylation of PKR and eIF2a was also seen in the gastrocnemius muscle of diabetic ob/ob mice. High glucose levels also inhibited protein synthesis. The effect of glucose on protein synthesis and degradation was not seen in myotubes transfected with a catalytically inactive variant (PKR?6). High glucose also induced an increased activity of both caspase-3 and -8, which led to activation of PKR, since this was completely attenuated by the specific caspase inhibitors. Activation of PKR also led to activation of p38MAPK (mitogen activated protein kinase), leading to ROS (reactive oxygen species) formation, since this was attenuated by the specific p38MAPK inhibitor SB203580. ROS formation was important in protein degradation, since it was completely attenuated by the antioxidant butylated hydroxytoluene. These results suggest that high glucose induces muscle atrophy through the caspase-3/-8 induced activation of PKR, leading to phosphorylation of eIF2a and depression of protein synthesis, together with PKR-mediated ROS production, through p38MAPK and increased protein degradation.

Effect of cancer cachexia on the activity of tripeptidyl-peptidase II in skeletal muscle

The ubiquitin-proteasome proteolytic pathway plays a major role in degradation of myofibrillar proteins in skeletal muscle during cancer cachexia. The end-product of this pathway is oligopeptides and these are degraded by the extralysomal peptidase tripeptidyl-peptidase II (TPPII) together with various aminopeptidases to form tripeptides and amino acids. To investigate if a relationship exists between the activity of the proteasome and TPPII, functional activities have been measured in gastrocnemius muscle of mice bearing the MAC16 tumour, and with varying extents of weight loss. TPPII activity was quantitated using the specific substrate Ala-Ala-Phe-7-amido-4-methylcoumarin, while proteasome activity was determined as the 'chymotrypsin-like' enzyme activity. Both proteasome proteolytic activity and TPPII activity increased in parallel with increasing weight loss, reaching a maximum at 16% weight loss, after which there was a progressive decrease in activity for both proteases with increasing weight loss. In murine myotubes, proteolysis-inducing factor, which is a sulphated glycoprotein produced by cachexia-inducing tumours, induced an increase in activity of both proteasome and TPPII, with an identical dose-response curve, and both activities were inhibited by eicosapentaenoic acid. These results suggest that the activities of both the proteasome and TPPII are regulated in a parallel manner in cancer cachexia, and that both are induced by the same factor and probably have the same intracellular signalling pathways and transcription factors. © 2004 Elsevier Ireland Ltd. All rights reserved.

Induction of protein catabolism and the ubiquitin-proteasome pathway by mild oxidative stress

Muscle wasting in cancer cachexia is associated with increased levels of malondialdehyde (MDA) in gastrocnemius muscles, suggesting an increased oxidative stress. To determine whether oxidative stress contributes to muscle protein catabolism, an in vitro model system, consisting of C2C12 myotubes, was treated with either 0.2 mM FeSO4, 0.1 mM H2O2, or both, to replicate the rise in MDA content in cachexia. All treatments caused an increased protein catabolism and a decreased myosin expression. There was an increase in the proteasome chymotrypsin-like enzyme activity, while immunoblotting showed an increased expression of the 20S proteasome α-subunits, p42, and the ubiquitin-conjugating enzyme, E214k. These results show that mild oxidative stress increases protein degradation in skeletal muscle by causing an increased expression of the major components of the ubiquitin-proteasome pathway. © 2002 Elsevier Science Ireland Ltd. All rights reserved.

Effect of a tumour-derived lipid-mobilising factor on glucose and lipid metabolism in vivo

Treatment of ex-breeder male NMRI mice with lipid mobilising factor isolated from the urine of cachectic cancer patients, caused a significant increase in glucose oxidation to CO2, compared with control mice receiving phosphate buffered saline. Glucose utilisation by various tissues was determined by the 2-deoxyglucose tracer technique and shown to be elevated in brain, heart, brown adipose tissue and gastrocnemius muscle. The tissue glucose metabolic rate was increased almost three-fold in brain, accounting for the ability of lipid mobilising factor to decrease blood glucose levels. Lipid mobilising factor also increased overall lipid oxidation, as determined by the production of 14CO2 from [14C carboxy] triolein, being 67% greater than phosphate buffered saline controls over a 24 h period. There was a significant increase in [14C] lipid accumulation in plasma, liver and white and brown adipose tissue after administration of lipid mobilising factor. These results suggest that changes in carbohydrate metabolism and loss of adipose tissue, together with an increased whole body fatty acid oxidation in cachectic cancer patients, may arise from tumour production of lipid mobilising factor. © 2002 Cancer Research UK.

Leg muscles motion during whole body linear frequency sweep vibration

Whole body vibration (WBV) aims to mechanically activate muscles by eliciting stretch reflexes. Mechanical vibrations are usually transmitted to the patient body standing on a oscillating plate. WBV is now more and more utilized not only for fitness but also in physical therapy, rehabilitation and in sport medicine. Effects depend on intensity, direction and frequency of vibration; however, the training frequency is one of the most important factors involved. A preliminary vibratory session can be dedicated to find the best vibration frequency for each subject by varying, stepwise, the stimulation frequency and analyzing the resulting EMG activity. This study concentrates on the analysis of muscle motion in response to a vibration frequency sweep, while subjects held two different postures. The frequency of a vibrating platform was increased linearly from 10 to 60 Hz in 26 s, while platform and single muscles (Rectus Femoris, Biceps Femoris - long head and Gastrocnemius Lateralis) motions were monitored using tiny, lightweight three-axial MEMS accelerometers. Displacements were estimated integrating twice the acceleration data after gravity contribution removal. Mechanical frequency response (amplitude and phase) of the mechanical chains ending at the single muscles was characterized. Results revealed a mechanical resonant-like behavior at some muscles, very similar to a second-order system in the frequency interval explored; resonance frequencies and dumping factors depended on subject and its positioning onto the vibrating platform. Stimulation at the resonant frequency maximizes muscle lengthening, and in turn muscle spindle solicitation, which produce muscle activation. © 2009 Springer-Verlag.