Search

Background: Cancer cachexia is characterized by weight loss, especially ongoing skeletal muscle loss, and is associated with poor patient outcomes. However, the molecular mechanism of skeletal muscle wasting is not fully understood., Objectives: We aimed to investigate muscle fiber morphology and proteolysis system activity changes that may account for cancer cachexia and to relate these changes to patients' clinical phenotypes., Methods: We divided 39 patients with resectable gastric cancer into 4 groups based on the presence of cachexia (weight loss) and/or sarcopenia (low muscularity), including a noncachexia/nonsarcopenia group (N, n = 10), a cachexia/sarcopenia group (CS, n = 13), a cachexia/nonsarcopenia group (C, n = 9), and a noncachexia/sarcopenia group (S, n = 7). Rectus abdominis muscle biopsy specimens were obtained intraoperatively. Muscle fiber size, ultrastructural architecture, and the expression of autophagic-lysosomal system (ALS) and ubiquitin proteasome system (UPS) markers were assayed., Results: Mean ± SD muscle fiber cross-sectional areas were significantly decreased in the CS (460 ± 120 μm2) and S groups (480 ± 135 μm2) compared with the N (1615 ± 388 μm2, both P < 0.05) and C groups (1219 ± 302 μm2, both P < 0.05). In the C, S, and CS groups, the muscle exhibited tissue disorganization and autophagosome formation to different degrees. The levels of ALS and UPS markers were significantly increased in the CS, C, and S groups compared with the N group. Alterations in muscle fiber morphology and increased ALS and UPS activity were related to severe muscle loss, but not weight loss., Conclusions: The ALS and UPS are simultaneously activated in cancer cachexia and may play coordinated roles in cachexia-induced muscle loss., (Copyright © The Author(s) 2020.)

Cancer cachexia is a systemic wasting syndrome characterized by anorexia and loss of body weight. The xanthine oxidase (XO) inhibitor febuxostat is one of the promising candidates for cancer cachexia treatment. However, cachexic symptoms were not alleviated by oral administration of febuxostat in our cancer cachexia model. Metabolomic analysis with brains of our cachexic model showed that purine metabolism was activated and XO activity was increased, and thus suggested that febuxostat would not reach the brain. Accordingly, targeting XO in the brain, which controls appetite, may be an effective strategy for treatment of cancer cachexia., (Copyright © 2019 The Authors. Production and hosting by Elsevier B.V. All rights reserved.)

Many chronic diseases are associated with unintentional loss of body weight, which is termed "cachexia". Cachexia is a complex multifactorial syndrome associated with the underlying primary disease, and characterized by loss of skeletal muscle with or without loss of fat tissue. Patients with cachexia face dire symptoms like dyspnea, fatigue, edema, exercise intolerance, and low responsiveness to medical therapy, which worsen quality of life. Because cachexia is not a stand-alone disorder, treating primary disease - such as cancer - takes precedence for the physician, and it remains mostly a neglected illness. Existing clinical trials have demonstrated limited success mostly because of their monotherapeutic approach and late detection of the syndrome. To conquer cachexia, it is essential to identify as many molecular targets as possible using the latest technologies we have at our disposal. In this review, we have discussed different aspects of cachexia, which include various disease settings, active molecular pathways, and recent novel advances made in this field to understand consequences of this illness. We also discuss roles of the sirtuins, the NAD + -dependent lysine deacetylases, microRNAs, certain dietary options, and epigenetic drugs as potential approaches, which can be used to tackle cachexia as early as possible in its course.

Doxorubicin (DOX) is a widely used drug for cancer treatment as a chemotherapeutic agent. However, the cellular and integrative mechanism of DOX-induced immunometabolism is unclear. Two-month-old male C57BL/6J mice were divided into high- and low-dose DOX-treated groups with a maintained saline control group. The first group was injected with a high dose of DOX (H-DOX; 15 mg·kg -1 ·wk -1 ), and the second group was injected with 7.5 mg·kg -1 ·wk -1 as a latent low dose of DOX (LL-DOX). H-DOX treatment led to complete mortality in 2 wk and 70% survival in the LL-DOX group compared with the saline control group. Therefore, an additional group of mice was injected with an acute high dose of DOX (AH-DOX) and euthanized at 24 h to compare with LL-DOX and saline control groups. The LL-DOX and AH-DOX groups showed obvious apoptosis and dysfunctional and structural changes in cardiac tissue. Splenic contraction was evident in AH-DOX- and LL-DOX-treated mice, indicating the systems-wide impact of DOX on integrative organs of the spleen, which is essential for cardiac homeostasis and repair. DOX dysregulated splenic-enriched immune-sensitive lipoxygenase and cyclooxygenase in the spleen and left ventricle compared with the saline control group. As a result, lipoxygenase-dependent D- and E-series resolvin precursors, such as 16HDoHE, 4HDoHE, and 12-HEPE, as well as cyclooxygenase-mediated PG species (PGD 2 , PGE 2, and 6-keto-PG 2α ) were decreased in the left ventricle, suggestive of defective immunometabolism. Both AH-DOX and LL-DOX induced splenic contraction and expansion of red pulp with decreased CD169 + metallophilic macrophages. AH-DOX intoxicated macrophages in the spleen by depleting CD169 + cells in the acute setting and sustained the splenic macrophage loss in the chronic phase in the LL-DOX group. Thus, DOX triggers a vicious cycle of splenocardiac cachexia to facilitate defective immunometabolism and irreversible macrophage toxicity and thereby impaired the inflammation-resolution program. NEW & NOTEWORTHY Doxorubicin (DOX) triggered splenic mass loss and decreased CD169 with germinal center contraction in acute and chronic exposure. Cardiac toxicity of DOX is marked with dysregulation of immunometabolism and thereby impaired resolution of inflammation. DOX suppressed physiological levels of cytokines and chemokines with signs of splenocardiac cachexia.

The phosphodiesterase 4 (PDE4)-cAMP pathway plays a predominant role in mediating skeletal muscle proteolysis in burn injury. The present investigations to determine the PDE4 isoform(s) involved in this action revealed that burn injury increased the expression of rat skeletal muscle PDE4B mRNA by sixfold but had little or no effect on expression of other PDE4 isoforms. These observations led us to study the effects of burn in PDE4B knockout (KO) rats. As reported by us previously, burn injury significantly increased extensor digitorum longus (EDL) muscle total and myofibrillar proteolysis in wild-type (WT) rats, but there were no significant effects on either total or myofibrillar protein breakdown in EDL muscle of PDE4B KO rats with burn injury. Moreover, burn injury increased PDE4 activity in the skeletal muscle of WT rats, but this was reduced by >80% in PDE4B KO rats. Also, burn injury decreased skeletal muscle cAMP concentration in WT rats but had no significant effects in the muscles of PDE4B KO rats. Incubation of the EDL muscle of burn-PDE4B KO rats with an inhibitor of the exchange factor directly activated by cAMP, but not with a protein kinase A inhibitor, eliminated the protective effects of PDE4B KO on EDL muscle proteolysis and increased muscle proteolysis to the same extent as in the EDL of burn-WT rats. These novel findings confirm a major role for PDE4B in skeletal muscle proteolysis in burn injury and suggest that an innovative therapy based on PDE4B-selective inhibitors could be developed to treat skeletal muscle cachexia in burn injury without the fear of causing emesis, which is associated with PDE4D inhibition.

Skeletal muscle dysfunction and mass loss is a characteristic feature in patients with chronic diseases including cancer and acute conditions such as critical illness. Maintenance of an adequate muscle mass is crucial for the patients' prognosis irrespective of the underlying condition. Moreover, aging-related sarcopenia may further aggravate the muscle wasting process associated with chronic diseases and cancer. Poly(adenosine diphosphate-ribose) polymerase (PARP) activation has been demonstrated to contribute to the pathophysiology of muscle mass loss and dysfunction in animal models of cancer-induced cachexia. Genetic inhibition of PARP activity attenuated the deleterious effects seen on depleted muscles in mouse models of oncologic cachexia. In the present minireview the mechanisms whereby PARP activity inhibition may improve muscle mass and performance in models of cancer-induced cachexia are discussed. Specifically, the beneficial effects of inhibition of PARP activity on attenuation of increased oxidative stress, protein catabolism, poor muscle anabolism and mitochondrial content and epigenetic modulation of muscle phenotype are reviewed in this article. Finally, the potential therapeutic strategies of pharmacological PARP activity inhibition for the treatment of cancer-induced cachexia are also being described in this review.

Cancer-associated cachexia is a disorder characterized by loss of body weight with specific losses of skeletal muscle and adipose tissue. Cachexia is driven by a variable combination of reduced food intake and metabolic changes, including elevated energy expenditure, excess catabolism and inflammation. Cachexia is highly associated with cancers of the pancreas, oesophagus, stomach, lung, liver and bowel; this group of malignancies is responsible for half of all cancer deaths worldwide. Cachexia involves diverse mediators derived from the cancer cells and cells within the tumour microenvironment, including inflammatory and immune cells. In addition, endocrine, metabolic and central nervous system perturbations combine with these mediators to elicit catabolic changes in skeletal and cardiac muscle and adipose tissue. At the tissue level, mechanisms include activation of inflammation, proteolysis, autophagy and lipolysis. Cachexia associates with a multitude of morbidities encompassing functional, metabolic and immune disorders as well as aggravated toxicity and complications of cancer therapy. Patients experience impaired quality of life, reduced physical, emotional and social well-being and increased use of healthcare resources. To date, no effective medical intervention completely reverses cachexia and there are no approved drug therapies. Adequate nutritional support remains a mainstay of cachexia therapy, whereas drugs that target overactivation of catabolic processes, cell injury and inflammation are currently under investigation.

Strategies to treat cachexia are still at its infancy. Enhanced muscle protein breakdown and ubiquitin-proteasome system are common features of cachexia associated with chronic conditions including lung cancer (LC). Poly(ADP-ribose) polymerases (PARP), which play a major role in chromatin structure regulation, also underlie maintenance of muscle metabolism and body composition. We hypothesized that protein catabolism, proteolytic markers, muscle fiber phenotype, and muscle anabolism may improve in respiratory and limb muscles of LC-cachectic Parp-1-deficient (Parp-1 -/- ) and Parp-2 -/- mice. In diaphragm and gastrocnemius of LC (LP07 adenocarcinoma) bearing mice (wild type, Parp-1 -/- , and Parp-2 -/- ), PARP activity (ADP-ribose polymers, pADPr), redox balance, muscle fiber phenotype, apoptotic nuclei, tyrosine release, protein ubiquitination, muscle-specific E3 ligases, NF-κB signaling pathway, markers of muscle anabolism (Akt, mTOR, p70S6K, and mitochondrial DNA) were evaluated along with body and muscle weights, and limb muscle force. Compared to wild type cachectic animals, in both respiratory and limb muscles of Parp-1 -/- and Parp-2 -/- cachectic mice: cancer induced-muscle wasting characterized by increased PARP activity, protein oxidation, tyrosine release, and ubiquitin-proteasome system (total protein ubiquitination, atrogin-1, and 20S proteasome C8 subunit) were blunted, the reduction in contractile myosin and atrophy of the fibers was attenuated, while no effects were seen in other structural features (inflammatory cells, internal or apoptotic nuclei), and markers of muscle anabolism partly improved. Activation of either PARP-1 or -2 is likely to play a role in muscle protein catabolism via oxidative stress, NF-κB signaling, and enhanced proteasomal degradation in cancer-induced cachexia. Therapeutic potential of PARP activity inhibition deserves attention., (© 2017 Wiley Periodicals, Inc.)

Cachexia is the result of complex metabolic alterations which cause morbidity and mortality in patients with advanced cancers including undifferentiated (anaplastic) thyroid carcinoma (ATC). ATC is a lethal disease with limited therapeutic options and unclear etiology for cachexia. We hypothesize that the BRAF(V600E) oncoprotein triggers microvascular endothelial cell tubule formation (in vitro angiogenesis) by means of factors which play a crucial role in angiogenic switch, inflammation/immune response and cachexia. We use human ATC cells and applied multiplex ELISA assay to screen for and measure angiogenic/cachectic and pro-inflammatory factors in the ATC-derived secretome. We find that vemurafenib anti-BRAF(V600E) therapy significantly reduces secreted VEGFA, VEGFC and IL6 protein levels compared to vehicle-treated ATC cells. As a result, the secretome from vemurafenib-treated ATC cells inhibits microvascular endothelial cell-related in vitro angiogenesis. Furthermore, ATC clinical samples express VEGFA, VEGFC and IL6 proteins. Our results suggest that angiogenic/cachectic and pro-inflammatory/immune response factors could play a crucial role in BRAF(V600E)-positive human ATC aggressiveness. Understanding the extent to which microenvironment-associated angiogenic factors participate in cachexia and cancer metabolism in advanced thyroid cancers will reveal new biomarkers and foster novel therapeutic approaches., Competing Interests: All authors declare no financial conflicts of interest, (Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.)

Respiratory dysfunction is prevalent in critically ill patients and can lead to adverse clinical outcomes, including respiratory failure and increased mortality. Respiratory muscles, which normally sustain respiration through inspiratory muscle contractions, become weakened during critical illness, and recent studies suggest that respiratory muscle weakness is related to systemic inflammation. Here, we investigate the pathophysiological role of the inflammatory JAK1/3 signaling pathway in diaphragm weakness in two distinct experimental models of critical illness. In the first experiment, mice received subcutaneous injections of PBS or C26 cancer cells and were fed chow formulated with or without the JAK1/3 inhibitor R548 for 26 days. Diaphragm specific force was significantly reduced in tumor-bearing mice receiving standard chow; however, treatment with the JAK1/3 inhibitor completely prevented diaphragm weakness. Diaphragm cross-sectional area was diminished by ∼25% in tumor-bearing mice but was similar to healthy mice in tumor-bearing animals treated with R548. In the second study, mice received sham surgery or coronary artery ligation, leading to myocardial infarction (MI), and were treated with R548 or vehicle 1 h postsurgery, and once daily for 3 days. Diaphragm specific force was comparable between sham surgery/vehicle, sham surgery/R548 and MI/R548 groups, but significantly decreased in the MI/vehicle group. Markers of oxidative damage and activated caspase-3, mechanisms previously identified to reduce muscle contractility, were not elevated in diaphragm extracts. These experiments implicate JAK1/3 signaling in cancer- and MI-mediated diaphragm weakness in mice, and provide a compelling case for further investigation., (Copyright © 2016 the American Physiological Society.)

Background: Cachexia is a metabolic disorder characterised by muscle wasting, diminished response to anti-cancer treatments and poor quality of life. Our objective was to identify blood-based biomarkers of cachexia in advanced cancer patients. Hence, we characterised the plasma cytokine and blood cell mRNA profiles of patients grouped in three cohorts: patients with cachexia, pre-cachexia (no cachexia but high CRP levels: ⩾ 5 mg l⁻¹) and no cachexia (no cachexia and CRP: < 5 mg l⁻¹)., Methods: A total of 122 newly diagnosed cancer patients with seven cancer types were studied prior to their initial therapy. Plasma levels of 22 cytokines were quantified using the bio-plex technology. mRNAs isolated from whole blood and expression profiles were determined by the chip array technology and Ingenuity Pathway Analysis (IPA) software., Results: In comparison with non-cachectic individuals, both pre-cachectic and cachectic patients showed an increase (⩾ 1.5-folds) in mRNA expression of neutrophil-derived proteases (NDPs) and significantly elevated angiotensin II (Ang II) (P = 0.005 and P = 0.02, respectively), TGFβ1 (P = 0.042 and P < 0.0001, respectively) and CRP (both P < 0.0001) in the plasma. Moreover, cachectic patients displayed a significant increase in IL-6 (P = 0.005), IL-8 (P = 0.001) and absolute neutrophil counts (P = 0.007)., Conclusions: Ang II, TGFβ1, CRP and NDP are blood biomarkers for cancer cachexia. These findings contribute to early diagnosis and prevention of cachexia.

Muscle wasting is associated with a variety of chronic or inflammatory disorders. Evidence suggests that inflammatory cytokines play a vital role in muscle inflammatory pathology and this may result in oxidative damage and mitochondrial dysfunction in skeletal muscle. In our study, we used microwave degradation to prepare a water-soluble low molecular weight guluronate (LMG) of 3000 Da from Fucus vesiculosus obtained from Canada, the Atlantic Ocean. We demonstrated the structural characteristics, using HPLC, FTIR and NMR of LMG and investigated its effects on oxidative damage and mitochondrial dysfunction in C2C12 skeletal muscle cells induced by tumor necrosis factor alpha (TNF-α), a cell inflammatory cytokine. The results indicated that LMG could alleviate mitochondrial reactive oxygen species (ROS) production, increase the activities of antioxidant enzymes (GSH and SOD), promote mitochondrial membrane potential (MMP) and upregulate the expression of mitochondrial respiratory chain protein in TNF-α-induced C2C12 cells. LMG supplement also increased the mitochondrial DNA copy number and mitochondrial biogenesis related genes in TNF-α-induced C2C12 cells. LMG may exert these protective effects through the nuclear factor kappa B (NF-κB) signaling pathway. These suggest that LMG is capable of protecting TNF-α-induced C2C12 cells against oxidative damage and mitochondrial dysfunction.

Cancer cachexia syndrome is observed in 80% of patients with advanced-stage cancer, and it is one of the most frequent causes of death. Severe wasting accounts for more than 80% in patients with advanced pancreatic cancer. Here we wanted to define, by using an microarray approach and the Pdx1-cre;LSL-Kras(G12D) ;INK4a/arf(fl/fl) mice model, the pathways involved in muscle, liver, and white adipose tissue wasting. These mice, which develop systematically pancreatic cancer, successfully reproduced many human symptoms afflicted with this disease, and particularly cachexia. Using the profiling analysis of pancreatic cancer-dependent cachectic tissues we found that Jak2/Stat3 pathways, p53 and NFkB results activated. Thus, our interest was focused on the Jak2 pathways because it is pharmacologically targetable with low toxicity and FDA approved drugs are available. Therefore, Pdx1-cre;LSL-Kras(G12D) ;INK4a/arf(fl/fl) mice were treated with the Jak2 inhibitor AG490 compound daily starting at 7 weeks old and for a period of 3 weeks and animals were sacrificed at 10 weeks old. Body weight for control mice was 27.84 ± 2.14 g, for untreated Pdx1-cre;LSL-Kras(G12D) ;INK4a/arf(fl/fl) was 14.97 ± 1.99 g, whereas in animals treated with the AG490 compound the weight loss was significantly less to 24.53 ± 2.04 g. Treatment with AG490 compound was efficient since phosphorylation of Jak2 and circulating interleukin-6 (IL6) levels were significantly reduced in cachectic tissues and in mice respectively. In conclusion, we found that Jak2/Stat3-dependent intracellular pathway plays an essential role since its pharmacological inhibition strongly attenuates cachexia progression in a lethal transgenic pancreatic cancer model., (© 2014 Wiley Periodicals, Inc.)

Background: In cancer cachexia, muscle depletion is related to morbidity and mortality. Muscle-wasting mechanisms in cancer patients are not fully understood., Objective: We investigated the involvement of the proteolytic systems (proteasome, autophagic-lysosomal, calpain, and caspase) in muscle wasting during cancer cachexia., Design: Esophageal cancer patients [n = 14; mean ± SD age: 64.1 ± 6.6 y] and weight-stable control patients undergoing reflux surgery (n = 8; age: 57.5 ± 5.8 y) were included. Enzymatic activities were measured in the vastus lateralis and diaphragm. Protein expressions were also measured in the vastus lateralis of control (n = 7) and cancer (n = 8) patients., Results: Proteasome, calpain, and caspase 3 activities in the vastus lateralis and diaphragm muscles did not differ between the 2 groups. Cathepsin B and L activities were 90% (± SD) [2.4 ± 0.2 compared with 1.3 ± 0.2 pmol 7-amido-4-methylcoumarin (AMC) · μg protein⁻¹ · min⁻¹; P < 0.001] and 115% (5.3 ± 0.4 compared with 2.5 ± 0.3 pmol AMC · μg protein⁻¹ · min⁻¹; P < 0.001) greater, respectively, in the vastus lateralis of cancer patients than in that of control subjects. We observed (in conjunction with increased lysosomal protease activities) higher microtubule-associated protein 1 light chain 3B-II/I ratios (0.14 ± 0.08 compared with 0.04 ± 0.04) and cathepsin B and L expressions in the vastus lateralis of cancer patients than in that of control subjects (P < 0.05). Protein expression of p62 in the vastus lateralis did not differ between the 2 groups., Conclusions: The autophagic-lysosomal pathway in the skeletal muscle of cancer patients was modified, whereas other proteolytic systems were unchanged. These findings suggest involvement of the autophagic-lysosomal proteolytic system during cancer cachexia development in humans.

Background: Cachexia is a common complication of cancer and may be responsible for 22% of all cancer-related deaths. The exact cause of death in cancer cachexia patients is unknown. Recently, atrophy of the heart has been described in cancer cachexia animal models, which resulted in impaired cardiac function and is likely to contribute to mortality. In cancer patients hyperuricaemia independent of tumour lysis syndrome is often associated with a worse prognosis. Xanthine oxidase (XO) metabolizes purines to uric acid and its inhibition has been shown to improve clinical outcome in patients with chronic heart failure., Methods: The rat Yoshida AH-130 hepatoma cancer cachexia model was used in this study. Rats were treated with 4 or 40 mg/kg/d oxypurinol or placebo starting one day after tumour-inoculation for maximal 15 days. Cardiac function was analyzed by echocardiography on day 11., Results: Here we show that inhibition of XO by oxypurinol significantly reduces wasting of the heart and preserves cardiac function. LVEF was higher in tumour-bearing rats treated with 4 mg/kg/d (61±4%) or 40 mg/kg/d (64±5%) oxypurinol vs placebo (51±3%, both p<0.05). Fractional shortening was improved by 4 mg/kg/d (43±3%) oxypurinol vs placebo (30±2, p<0.05), while 40 mg/kg/d oxypurinol (41±5%) did not reach statistical significance. Cardiac output was increased in the 4 mg/kg/d dose only (71±11 mL/min vs placebo 38±4 mL/min, p<0.01)., Conclusion: Inhibition of XO with oxypurinol has beneficial effects on cardiac mass and function in a rat model of severe cancer cachexia, suggesting that XO might be a viable drug target in cancer cachexia., (Copyright © 2013 Elsevier Ireland Ltd. All rights reserved.)

Although catabolic signaling has a well-established role in muscle wasting during cancer cachexia, the suppression of anabolic signaling also warrants further investigation. In cachectic tumor-bearing mice, circulating IL-6 levels are associated with suppressed muscle protein synthesis and mTORC1 signaling. We have found AMPK and IGF-I/insulin signaling, two well-known regulators of the mammalian target of rapamycin (mTOR), are altered with the progression of cachexia. How IL-6 can induce suppression of mTORC1 signaling remains to be established. The purpose of this study was to examine mTOR complex 1 (mTORC1) activation and regulation by IL-6 during cancer cachexia. IL-6 effects on mTOR activation were examined in Apc(Min/+) mouse skeletal muscle and C2C12 myotubes. Systemic IL-6 overexpression in Apc(Min/+) mice produced a dose-dependent suppression of mTOR signaling that corresponded to induction of STAT3 and AMPK phosphorylation. This result was also evident in IL-6-treated myotubes. Basal mTOR activation and mTOR responsiveness to glucose administration were suppressed in cachectic skeletal muscle. However, insulin induction of mTOR activity was maintained in IL-6-treated myotubes. Whereas IL-6 suppression of myotube mTOR activity was rescued by AMPK inhibition, inhibition of STAT3 signaling was not sufficient to rescue IL-6 suppression of mTOR activity. Last, treadmill exercise training was able to prevent IL-6-induced inhibition of mTOR signaling in Apc(Min/+) mice independently of activated STAT. In conclusion, we report dose-dependent suppression of mTOR activity by IL-6 and suppressed mTOR responsiveness to glucose administration in Apc(Min/+) mice. IL-6 suppression of mTOR activity was dependent on AMPK activation and independent of STAT signaling in myotubes.

Cancer associated cachexia (CAC) is a complex multiorgan syndrome frequently associated with various forms of cancer. Affected patients suffer from a dramatic loss of skeletal muscle and adipose tissue. Most cases are accompanied by anorexia, and nutritional supplements are not sufficient to stop or reverse its course. CAC impairs many forms of therapeutic interventions and accounts for 15-20% of all deaths of cancer patients. Recently, several studies have recognized the importance of lipid metabolism and triglyceride hydrolysis as a major metabolic pathway involved in the initiation and/or progression of CAC. In this review, we explore the contributions of the triglyceride lipases to CAC and discuss various factors modulating lipase activity., (Copyright © 2013 Elsevier Ltd. All rights reserved.)

Inhibitor of dsRNA-dependent protein kinase (PKRI) and medroxyprogesterone acetate (MPA) improve cancer cachexia via different mechanisms. We aimed to compare these two drugs, alone or in combination, in cancer cachexia in mice. Forty male BABL/c mice aged 6-8 weeks were randomly divided into PKRI, MPA, PKRI+MPA, placebo, and healthy control groups. The first 4 groups were injected with colon-26 adenocarcinoma and fed for 12 days and then treated with PKRI and MPA alone or in combination for 7 days. Body weight, tumor volume, wet weight of gastrocnemius muscle, serum levels of nutritional markers and cytokines were measured. The tumor growth (volume and weight) of mice treated with PKRI, MPA alone or PKRI+MPA was slower than that of placebo group. Wet weight of gastrocnemius muscle was significantly higher in PKRI and PKRI+MPA-treated than in placebo animals (P<0.01). All tumor-bearing mice had a significantly lower level of blood glucose, higher level of serum triglyceride and lower level of serum albumin compared with healthy control (P<0.001). However, PKRI, MPA and PKRI+MPA groups had a significant higher level of blood glucose and lower level of serum triglyceride compared with placebo group (P<0.001). All tumor bearing mice had a significant higher level of serum TNF-α, IL-1 and IL-6 compared with healthy control (P<0.001). Serum level of TNF-α and IL-6 was significantly lower in PKRI and PKRI+MPA-treated than in placebo animals (P<0.01). PKRI alone and combination therapy with PKRI and MPA reduce tumor growth and may alleviate cachexia.

Muscle atrophy-also known as muscle wasting-is a debilitating syndrome that slowly develops with age (sarcopenia) or rapidly appears at the late stages of deadly diseases such as cancer, AIDS, and sepsis (cachexia). Despite the prevalence and the drastic detrimental effects of these two syndromes, there are currently no widely used, effective treatment options for those suffering from muscle wasting. In an attempt to identify potential therapeutic targets, the molecular mechanisms of sarcopenia and cachexia have begun to be elucidated. Growing evidence suggests that inflammatory cytokines may play an important role in the pathology of both syndromes. As one of the key cytokines involved in both sarcopenic and cachectic muscle wasting, tumor necrosis factor α (TNFα) and its downstream effectors provide an enticing target for pharmacological intervention. However, to date, no drugs targeting the TNFα signaling pathway have been successful as a remedial option for the treatment of muscle wasting. Thus, there is a need to identify new effectors in this important pathway that might prove to be more efficacious targets. Inducible nitric oxide synthase (iNOS) has recently been shown to be an important mediator of TNFα-induced cachectic muscle loss, and studies suggest that it may also play a role in sarcopenia. In addition, investigations into the mechanism of iNOS-mediated muscle loss have begun to reveal potential therapeutic strategies. In this review, we will highlight the potential for targeting the iNOS/NO pathway in the treatment of muscle loss and discuss its functional relevance in sarcopenia and cachexia.

Cancer cachexia is characterized by progressive weight loss with the depletion of adipose tissue and skeletal muscle. Impaired fatty acid oxidation mainly resulting from the decrease of carnitine palmitoyltransferase I and II activities in the liver is an important factor that contributes to cancer cachexia . Although recent studies suggest a potential application of L-carnitine in treatment of cancer cachexia, the underlying mechanisms are unknown. In the present study, we aim to assess the effects of L-carnitine on the activity and expression of CPT I and II in the liver of cachectic cancer mice. Our results show that the inoculation of colon-26 adenocarcinoma cells into mice led to cancer cachexia characterized by notable decreases in food intake, gastrocnemius muscle and epididymus fat weight. In addition, the mRNA level and activity of liver carnitine palmitoyltransferase (CPT) I and II, and serum levels of free carnitine and acetylcarnitine were markedly decreased in cachectic mice, accompanied by marked increases in serum levels of tumor necrosis factor-alpha (TNF-α) and interleukin-6 (IL-6). A continuous oral treatment with L-carnitine at 18 mg/kg per day increased dietary uptake, gastrocnemius muscle weight and epididymus fat weight, increased blood glucose and serum albumin levels, and decreased total cholesterol level in cancer cachectic mice, but did not affect tumor growth. These effects of L-carnitine on cancer cachexia mice were accompanied by the upregulation of mRNA level of CPT I and II and increased enzyme activity of CPT I in the liver, as well as the downregulation of serum TNF-α and IL-6 levels. Moreover, free carnitine levels were negatively correlated with serum TNF-α or IL-6 level. These results indicate that L-carnitine ameliorates cancer cachexia by regulating serum TNF-α and IL-6 levels and modulating the expression and activity of CPT in the liver.

Cachexia is a multifactorial wasting syndrome most common in patients with cancer that is characterized by the uncontrolled loss of adipose and muscle mass. We show that the inhibition of lipolysis through genetic ablation of adipose triglyceride lipase (Atgl) or hormone-sensitive lipase (Hsl) ameliorates certain features of cancer-associated cachexia (CAC). In wild-type C57BL/6 mice, the injection of Lewis lung carcinoma or B16 melanoma cells causes tumor growth, loss of white adipose tissue (WAT), and a marked reduction of gastrocnemius muscle. In contrast, Atgl-deficient mice with tumors resisted increased WAT lipolysis, myocyte apoptosis, and proteasomal muscle degradation and maintained normal adipose and gastrocnemius muscle mass. Hsl-deficient mice with tumors were also protected although to a lesser degree. Thus, functional lipolysis is essential in the pathogenesis of CAC. Pharmacological inhibition of metabolic lipases may help prevent cachexia.

It is strongly established by numerous studies that oxidative stress-induced inflammation is one of the major causative agents in a variety of cancers. Various factors such as bacterial, viral, parasitic infections, chemical irritants, carcinogens are involved in the initiation of oxidative stress-mediated inflammation. Chronic and persistent inflammation promotes the formation of cancerous tumors. Recent investigations strongly suggest that aldose reductase [AR; AKR1B1], a member of aldo-keto reductase superfamily of proteins, is the mediator of inflammatory signals induced by growth factors, cytokines, chemokines, carcinogens etc. Further, AR reduced product(s) of lipid derived aldehydes and their metabolites such as glutathionyl 1,4-dihydroxynonanol (GS-DHN) have been shown to be involved in the activation of transcription factors such as NF-κB and AP-1 which transcribe the genes of inflammatory cytokines. The increased inflammatory cytokines and growth factors promote cell proliferation, a main feature involved in the tumorigenesis process. Inhibition of AR has been shown to prevent cancer cell growth in vitro and in vivo models. In this review, we have described the possible association between AR with oxidative stress- and inflammation- initiated carcinogenesis. A thorough understanding of the role of AR in the inflammation -associated cancers could lead to the use of AR inhibitors as novel chemotherapeutic agents against cancer.

Oxidative stress is a primary trigger of cachectic muscle wasting, but the signaling pathway(s) that links it to the muscle wasting processes remains to be defined. Here, we report that activation of p38 mitogen-activated protein kinase (MAPK) (phosphorylation) and increased oxidative stress (trans-4-hydroxy-2-nonenal protein modification) in skeletal muscle occur as early as 8 h after lipopolysaccharide (1 mg/kg) and 24 h after dexamethasone (25 mg/kg) injection (intraperitoneal) in mice, concurrent with upregulation of autophagy-related genes, Atg6, Atg7, and Atg12. Treating cultured C2C12 myotubes with oxidant hydrogen peroxide (4 h) resulted in increased p38 phosphorylation and reduced FoxO3 phosphorylation along with induced Atg7 mRNA expression without activation of NF-kappaB or FoxO3a transcriptional activities. Furthermore, inhibition of p38alpha/beta by SB202190 blocked hydrogen peroxide-induced atrophy with diminished upregulation of Atg7 and atrogenes [muscle atrophy F-box protein (MAFbx/Atrogin-1), muscle ring finger protein 1 (MuRF-1), and Nedd4]. These findings provide direct evidence for p38alpha/beta MAPK in mediating oxidative stress-induced autophagy-related genes, suggesting that p38alpha/beta MAPK regulates both the ubiquitin-proteasome and the autophagy-lysosome systems in muscle wasting.

We previously determined that ghrelin synthesis was downregulated after burn injury and that exogenous ghrelin retained its ability both to stimulate food intake and to restore plasma growth hormone levels in burned rats. These observations and the finding that anabolic hormones can attenuate skeletal muscle catabolism led us to investigate whether ghrelin could attenuate burn-induced skeletal muscle protein breakdown in rats. These studies were performed in young rats (50-60 g) 24 h after approximately 30% total body surface area burn injury. Burn injury increased total and myofibrillar protein breakdown in extensor digitorum longus (EDL) muscles assessed by in vitro tyrosine and 3-methyl-histidine release, respectively. Continuous 24-h administration of ghrelin (0.2 mg.kg(-1).h(-1)) significantly inhibited both total and myofibrillar protein breakdown in burned rats. Ghrelin significantly attenuated burn-induced changes in mRNA expression of IGFBP-1 and IGFBP-3 in liver. In EDL, ghrelin attenuated the increases in mRNA expression of the binding proteins, but had no significant effect on reduced expression of IGF-I. Ghrelin markedly reduced the elevated mRNA expression of TNF-alpha and IL-6 in EDL muscle that occurred after burn. Moreover, ghrelin normalized plasma glucocorticoid levels, which were elevated after burn. Expression of the muscle-specific ubiquitin-ligating enzyme (E3) ubiquitin ligases MuRF1 and MAFbx were markedly elevated in both EDL and gastrocnemius and were normalized by ghrelin. These results suggest that ghrelin is a powerful anticatabolic compound that reduces skeletal muscle protein breakdown through attenuating multiple burn-induced abnormalities.

Patients with sepsis in the ICU (intensive care unit) are characterized by skeletal muscle wasting. This leads to muscle dysfunction that also influences the respiratory capacity, resulting in prolonged mechanical ventilation. Catabolic conditions are associated with a general activation of the ubiquitin-proteasome pathway in skeletal muscle. The aim of the present study was to measure the proteasome proteolytic activity in both respiratory and leg muscles from ICU patients with sepsis and, in addition, to assess the variation of proteasome activity between individuals and between duplicate leg muscle biopsy specimens. When compared with a control group (n=10), patients with sepsis (n=10) had a 30% (P<0.05) and 45% (P<0.05) higher proteasome activity in the respiratory and leg muscles respectively. In a second experiment, ICU patients with sepsis (n=17) had a 55% (P<0.01) higher proteasome activity in the leg muscle compared with a control group (n=10). The inter-individual scatter of proteasome activity was larger between the patients with sepsis than the controls. We also observed a substantial intra-individual difference in activity between duplicate biopsies in several of the subjects. In conclusion, the proteolytic activity of the proteasome was higher in skeletal muscle from patients with sepsis and multiple organ failure compared with healthy controls. It was shown for the first time that respiratory and leg muscles were affected similarly. Furthermore, the variation in proteasome activity between individuals was more pronounced in the ICU patients for both muscle types, whereas the intra-individual variation between biopsies was similar for ICU patients and controls.

Chronic arthritis induces cachexia associated with an inhibition of the growth hormone (GH)-insulin-like growth factor-I (IGF-I) system and an activation of the E3 ubiquitin-ligating enzymes muscle atrophy F-box (MAFbx) and muscle Ring finger 1 (MuRF1) in the skeletal muscle. The aim of this work was to study the role of cyclooxygenase (COX)-2 in chronic arthritis-induced cachexia. Arthritis was induced in rats by Freund's adjuvant injection, and the effects of two COX inhibitors (indomethacin, a nonspecific inhibitor, and meloxicam, a selective COX-2 inhibitor on pituitary GH and on liver and serum IGF-I levels) were tested. Arthritis decreased body weight gain and GH and liver IGF-I gene expression. In the arthritic rats, both inhibitors, indomethacin and meloxicam, prevented the inhibitory effect of arthritis on body weight gain. Indomethacin and meloxicam administration to arthritic rats increased pituitary GH and liver IGF-I mRNA as well as serum levels of IGF-I. These data suggest that induction of COX-2 during chronic inflammation is involved in the inhibition of the GH-IGF-I axis and in the body weight loss. In the gastrocnemius muscle, arthritis increased the gene expression of tumor necrosis factor (TNF)-alpha, the E3 ubiquitin-ligating enzymes MAFbx and MuRF1, as well as of IGF-I and IGF-binding protein-5 (IGFBP-5). Inhibition of COX-2 by meloxicam administration increased gastrocnemius weight and decreased MAFbx, MuRF1, TNF-alpha, and IGFBP-5 gene expression. In summary, our data indicate that chronic arthritis-induced cachexia and muscle wasting are mediated by the COX-2 pathway resulting in a decreased GH-IGF-I secretion and increased expression of MAFbx and MuRF1 mRNA.

In animal models of cachexia, alterations in the phosphatidylinositol 3-kinase (PI3-K)/Akt pathway have been demonstrated in atrophying skeletal muscles. Therefore, we assessed the activity of proteins in this pathway in muscle and liver biopsies from 16 patients undergoing pancreatectomy for suspect of carcinoma. Patients were divided in a non-cachectic or cachectic group according to their weight loss before operation. Extracts of skeletal muscle and liver tissue from eight cachectic patients with pancreas carcinoma and eight non-cachectic patients were analysed by Western blotting using pan- and phospho-specific antibodies directed against eight important signal transduction proteins of the PI3-K/Akt pathway. Muscle samples from cachectic patients revealed significantly decreased levels of myosin heavy chain (-45%) and actin (-18%) in comparison to non-cachectic samples. Akt protein level was decreased by -55%. The abundance and/or phosphorylation of the transcription factors Foxo1 and Foxo3a were reduced by up to fourfold in muscle biopsies from cachectic patients. Various decreases of the phosphorylated forms of the protein kinases mTOR (-82%) and p70S6K (-39%) were found. In contrast to skeletal muscle, cachexia is associated with a significant increase in phosphorylated Akt level in the liver samples with a general activation of the PI3-K/Akt cascade. Our study demonstrates a cachexia-associated loss of Akt-dependent signalling in human skeletal muscle with decreased activity of regulators of protein synthesis and a disinhibition of protein degradation.

Muscle protein degradation is thought to play a major role in muscle atrophy in cancer cachexia. To investigate the importance of the ubiquitin-proteasome pathway, which has been suggested to be the main degradative pathway mediating progressive protein loss in cachexia, the expression of mRNA for proteasome subunits C2 and C5 as well as the ubiquitin-conjugating enzyme, E2(14k), has been determined in gastrocnemius and pectoral muscles of mice bearing the MAC16 adenocarcinoma, using competitive quantitative reverse transcriptase polymerase chain reaction. Protein levels of proteasome subunits and E2(14k) were determined by immunoblotting, to ensure changes in mRNA were reflected in changes in protein expression. Muscle weights correlated linearly with weight loss during the course of the study. There was a good correlation between expression of C2 and E2(14k) mRNA and protein levels in gastrocnemius muscle with increases of 6-8-fold for C2 and two-fold for E2(14k) between 12 and 20% weight loss, followed by a decrease in expression at weight losses of 25-27%, although loss of muscle protein continued. In contrast, expression of C5 mRNA only increased two-fold and was elevated similarly at all weight losses between 7.5 and 27%. Both proteasome functional activity, and proteasome-specific tyrosine release as a measure of total protein degradation was also maximal at 18-20% weight loss and decreased at higher weight loss. Proteasome expression in pectoral muscle followed a different pattern with increases in C2 and C5 and E2(14k) mRNA only being seen at weight losses above 17%, although muscle loss increased progressively with increasing weight loss. These results suggest that activation of the ubiquitin-proteasome pathway plays a major role in protein loss in gastrocnemius muscle, up to 20% weight loss, but that other factors such as depression in protein synthesis may play a more important role at higher weight loss.

Evidence implies that nitric oxide (NO) in the central nervous systems mediates anorexia in tumor-bearing hosts. We have therefore evaluated, by immunohistochemical image analyses, net alterations of nitric oxide synthases (nNOS, eNOS, iNOS) in brain nuclei [paraventricular hypothalamic nucleus (PVN), medial habenular nucleus (MHB), lateral habenular nucleus (LHB), paraventricular thalamic nucleus (PV), lateral hypothalamic area (LHA), ventromedial hypothalamic nucleus (VMH), nucleus of the solitary tract (NTS)] of tumor-bearing mice (TB) with prostanoid-related anorexia. Pair-fed (PF) and freely fed (FF) non-tumor-bearing mice were used as controls. c-fos was analyzed as indicator of neuronal activation. nNOS was significantly increased in VMH and PVN from TB mice, while eNOS was significantly increased in LHB and LHA. iNOS was significantly increased in LHA and PVN nuclei, but decreased in MHB, LHB and VMH from tumor-bearers. However, several of these alterations were similarly observed in brain nuclei from pair-fed controls. Provision of unspecific NOS-antagonists to TB mice increased nNOS, eNOS and iNOS in several brain nuclei (PVN, LHA, VMH), but left tumor-induced anorexia unchanged. c-fos was significantly increased in all brain nuclei in PF mice except for NTS, LHA and PVN compared to controls, while tumor-bearing mice had increased c-fos in LHA and PVN only compared to controls. Our results demonstrate a complex picture of NOS expression in brain areas of relevance for appetite in tumor-bearing hosts, where most changes seemed to be secondary to stress during negative energy balance. By contrast, NOS content in PVN and LHA nuclei remains candidate behind anorexia in tumor disease. However, nitric oxide does not seem to be a primary mediator behind tumor-induced anorexia. NO may rather secondarily support energy intake in conditions with negative energy balance.

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.

The progressive depletion of skeletal muscle is a hallmark of many types of advanced cancer and frequently is associated with debility, morbidity, and mortality. Muscle wasting is primarily mediated by the activation of the ubiquitin-proteasome system, which is responsible for degrading the bulk of intracellular proteins. E3 ubiquitin ligases control polyubiquitination, a rate-limiting step in the ubiquitin-proteasome system, but their direct involvement in muscle protein catabolism in cancer remains obscure. Here, we report the full-length cloning of E3alpha-II, a novel "N-end rule" ubiquitin ligase, and its functional involvement in cancer cachexia. E3alpha-II is highly enriched in skeletal muscle, and its expression is regulated by proinflammatory cytokines. In two different animal models of cancer cachexia, E3alpha-II was significantly induced at the onset and during the progression of muscle wasting. The E3alpha-II activation in skeletal muscle was accompanied by a sharp increase in protein ubiquitination, which could be blocked by arginine methylester, an E3alpha-selective inhibitor. Treatment of myotubes with tumor necrosis factor alpha or interleukin 6 elicited marked increases in E3alpha-II but not E3alpha-I expression and ubiquitin conjugation activity in parallel. E3alpha-II transfection markedly accelerated ubiquitin conjugation to endogenous cellular proteins in muscle cultures. These findings show that E3alpha-II plays an important role in muscle protein catabolism during cancer cachexia and suggest that E3alpha-II is a potential therapeutic target for muscle wasting.

Administration of the phosphodiesterase-IV inhibitor EMD 95832/3 (Merck KGaA, Darmstadt, Germany) to rats bearing the ascites hepatoma Yoshida AH-130, a highly cachectic tumour, could not prevent either the anorexia nor the massive weight loss (affecting both adipose and skeletal muscle tissues) present in the tumour-bearing animals. This compound did not have any effects on the fractional rates of protein turnover in skeletal muscle, and did not affect circulating triacylglycerols or lipoprotein lipase activity in adipose tissue. Although the administration of EMD 95832/3 did not influence tumour growth either, it did increase the number of tumour cells undergoing apoptosis. It is concluded that the drug is unable to reverse the cachectic state in this particular experimental tumour model.

Chemotherapy has cachectic effects, but it is unknown whether cytostatic agents alter skeletal muscle proteolysis. We hypothesized that chemotherapy-induced alterations in protein synthesis should result in the increased incidence of abnormal proteins, which in turn should stimulate ubiquitin-proteasome-dependent proteolysis. The effects of the nitrosourea cystemustine were investigated in skeletal muscles from both healthy and colon 26 adenocarcinoma-bearing mice, an appropriate model for testing the impact of cytostatic agents. Muscle wasting was seen in both groups of mice 4 days after a single cystemustine injection, and the drug further increased the loss of muscle proteins already apparent in tumor-bearing animals. Cystemustine cured the tumor-bearing mice with 100% efficacy. Surprisingly, within 11 days of treatment, rates of muscle proteolysis progressively decreased below basal levels observed in healthy control mice and contributed to the cessation of muscle wasting. Proteasome-dependent proteolysis was inhibited by mechanisms that include reduced mRNA levels for 20S and 26S proteasome subunits, decreased protein levels of 20S proteasome subunits and the S14 non-ATPase subunit of the 26S proteasome, and impaired chymotrypsin- and trypsin-like activities of the enzyme. A combination of cisplatin and ifosfamide, two drugs that are widely used in the treatment of cancer patients, also depressed the expression of proteasomal subunits in muscles from rats bearing the MatB adenocarcinoma below basal levels. Thus, a down-regulation of ubiquitin-proteasome-dependent proteolysis is observed with various cytostatic agents and contributes to reverse the chemotherapy-induced muscle wasting.

Previous studies have shown that five human cancer cell lines, LS180, MKN-1, MMG-1, C32 and LX-1, induced remarkable weight loss in tumor-bearing nude mice. With the aim of identifying novel molecules involved in lipid catabolism, conditioned media of these cancer cell lines were analyzed in terms of lipoprotein lipase (LPL)-inhibiting or lipolytic activities. All conditioned media from the five cell lines significantly suppressed LPL activity in 3T3-L1 cells, while media from LX-1 and C32 promoted lipolytic activity in a dose-dependent manner. RT-PCR and ELISA demonstrated that the major factors responsible for LPL inhibition and lipolysis were not IL-1beta, IL-6, IL-11, TNF-alpha, TGF-beta1 or LIF in all the cancer cell lines except for MMG-1 cells. Preliminary biochemical analysis showed that the LPL-inhibiting factor produced by LX-1 cells was approximately 65 kD and vulnerable to heat, whereas the lipolytic factor was less than 1 kD and heat stable. These results suggested that unknown factors are partially involved in the pathogenesis of cancer cachexia in tumor-bearing nude mice and further purification will be needed.

Several lines of evidence have postulated that reduction in the activity of lipoprotein lipase (LPL) is involved in cachexia induction in cancer patients. Recently we have demonstrated that murine melanoma B16 has the ability to reduce the LPL activity and thereby induce cachexia symptoms in mice following intraperitoneal inoculation. In order to further investigate the relationship between LPL activity and cachectic syndrome, cachexia models other than melanoma B16 are required. However, there are few animal cachexia models in which LPL activity is involved in the induction of cachectic symptoms. In this study, cachectic symptoms and plasma LPL activity were investigated in mice bearing EL-4 mouse lymphoma. In EL-4 bearing mice the body weight including tumor weight in the abdominal cavity was rather higher than that of normal mice without tumor, whereas weights of carcass wet and gastrocnemius muscle were significantly decreased in EL-4 bearing mice. Elevated blood levels of triglyceride and non-esterified fatty acid were observed in mice bearing EL-4, associated with the impaired plasma LPL activity. Overall, this study indicated that EL-4 lymphoma in mice results in a severe cachexia which is possibly related to impaired LPL activity and also provided a useful cachexia model for understanding the role of LPL in the development of cancer cachexia.

The Yoshida AH-130 rat ascites hepatoma is a model system for studying the mechanisms involved in the protein hypercatabolism associated with cancer cachexia. The present study was aimed at investigating if the calpain-3 gene expression in skeletal muscle was affected by tumor growth. The results presented clearly show that calpain-3 gene expression is considerably reduced in experimental cancer cachexia, while there is a reciprocal change in the expression of the ubiquitin-dependent proteolytic system and in the ubiquitous m-calpain. The results, observed during cancer cachexia, suggest a potential counterregulatory role of calpain-3 in muscle proteolysis.

Our recent study has demonstrated that ponalrestat, an aldose reductase inhibitor, activates lipoprotein lipase (LPL) activity in the adipose tissue and alleviates the cachectic symptoms induced by B16 melanoma in mice. In this study, the effect of ponalrestat on cachexia symptoms in nude mice bearing human melanomas G361 and SEKI was investigated because it has been suggested that the suppression of LPL has an important role in cachexia induction by these two melanomas in nude mice. Mice bearing G361 subcutaneously did not gain weight and became cachectic, associated with the tumor growth. Tumor growth was not affected by ponalrestat, nevertheless treatment with ponalrestat resulted in an amelioration of the reduction in the weight of body mass, epididymal fat, gastrocnemius muscle, carcass and whole body lipid induced by the presence of G361. A severe weight loss observed in nude mice bearing SEKI was also partially attenuated by ponalrestat treatment. Overall, this study showed that ponalrestat is effective in the attenuation of the cachectic symptoms induced by human melanomas G361 and SEKI in nude mice, suggesting that ponalrestat has a potential usefulness for the treatment of cancer cachexia.

When a cachexigenic subclone (clone 20) of murine colon 26 adenocarcinoma was transplanted into female BALB/c mice, hepatic NNMT activity continued to increase until death in proportion to progressive carcass weight loss, a marker of cancer cachexia. On the other hand, noncachexigenic subclone (clone 5)-transplanted mice showed neither increase of NNMT activity nor carcass weight loss. Among cytostatic fluorinated pyrimidines, 5'-dFUrd could inhibit the increase of NNMT activity and prevent weight loss in mice bearing clone 20. On the other hand, 2'-dFUrd did not show these effects. 5-FUra and Tegafur inhibited the increase of NNMT activity at higher concentrations. These findings suggest that the levels of hepatic NNMT activity are closely associated with the degree of weight loss, and they appear to be a useful marker of cancer cachexia.

The syndrome of cancer cachexia is accompanied by several alterations of lipid metabolism, especially that in the liver. In this study we have investigated a possible mechanism whereby the presence of the Walker 256 carcinosarcoma affects hepatic fatty acid oxidative capacity in tumour-bearing rats. Hepatic mitochondrial outer membrane carnitine palmitoyltransferase I (CPT I), generally accepted as the main site of regulation of fatty acid oxidation, was unaffected by the presence of the extra-hepatic tumour. However, mitochondrial inner-membrane carnitine palmitoyltransferase II (CPT II) activity was markedly decreased in mitochondria isolated from the liver of tumour-bearing rats. Immuno-detection by Western blotting using a CPT II-specific antibody identified two bands (corresponding to M(r) 69,000 and 54,000) in tumour-bearing rats whereas only the normal-sized CPT II was present (at the expected M(r) 69,000) in mitochondria from control rats. It is suggested that the emergence of the second, smaller protein may represent a catalytically less active protein that arises in vivo, since its appearance was not affected by the inclusion of proteolysis inhibitors in the mitochondrial preparation buffers. Treatment of the tumour-bearing rats with indomethacin, a prostaglandin (including PGE2) synthesis inhibitor, increased CPT II activity to levels even higher than those found in the control animals. It is suggested that PGE2 may play a role in the control of CPT II expression in the liver of tumour-bearing rats. Indomethacin treatment did not affect either of the two CPT activities of the mitochondria isolated from tumour tissue.

Background/aims: Cancer cachexia is characterized by a variety of metabolic disorders. Alterations in fat metabolism have been reported to be associated with suppression of tissue lipoprotein lipase (LPL) activity in tumor-bearing animals. Interleukin-6 (IL- 6) has been documented to reduce tissue LPL activity and may play a role in inducing cancer cachexia. This study was conducted to clarify the changes in LPL activity and the role of IL-6 in patients with either gastrointestinal cancer or breast cancer., Methodology: Twelve patients with colorectal cancer, 7 patients with gastric cancer, 7 patients with breast cancer and 5 normal volunteers were studied. Serum concentrations of triglycerides (TG), non-esterified fatty acids (NEFA) and IL-6 were measured. LPL activity was measured in plasma post-heparin administration. The relationships of LPL activity to tumor progression, body weight loss and serum IL-6 levels were examined. The effect of tumor resection on LPL activity was also studied., Results: LPL activity was suppressed with tumor progression in patients with either gastrointestinal cancer or breast cancer. Suppression of LPL activity and the degree of weight loss were negatively correlated in patients with either gastric or colorectal cancer (r = -0.5826, p = 0.011) but not in patients with breast cancer. The decrease in LPL activity was not always reversed after resection of the tumor. Circulating IL-6 did not correlate with either plasma LPL activity or tumor progression., Conclusions: Reduced LPL activity in patients with advanced gastrointestinal or breast cancer may reflect changes in nutritional status. Serum IL-6 is less likely to be a mediator of these alterations.

The role of insulin resistance in the tumor-induced decrease in tissue lipoprotein lipase (LPL) activity was studied in vivo and vitro in methylcholanthrene-induced sarcoma-bearing rats. Intraperitoneal (i.p.) injection of 2U of regular insulin resulted in high-adipose LPL activity in control rats (CTR) of 122.0 +/- 42.4 U/mg tissue, but it had little effect on tumor-bearing rats (TBR), which showed a value of only 9.6 +/- 5.5 U/mg tissue (P = 0.002). When adjusted for serum insulin concentrations, adipose LPL activity remained significantly different between the TBR and CTR at 0.19 +/- 0.17 and 0.78 +/- 0.29 U/mg tissue, respectively (P = 0.02). Following the in vitro incubation with either 1.44 g/l glucose of 1 x 10-8 U insulin of adipose tissue fragments obtained from the TBR and the CTR, measurable LPL activity was maintained in the tissue from the CTR for 2 h but not in that from the TBR. These results suggest that the decreased LPL activities seen in the tumor-bearing state may be mediated by insulin resistance.

Rats bearing the Yoshida AH-130 ascites hepatoma for 7 days showed an important decrease in muscle mass--over 30% in gastrocnemius and extensor digitorum longus (EDL)--in relation to non-tumour-bearing controls, which is associated with an increased proteolytic rate in in vitro incubation. In order to identify the precise biochemical process which was involved, we measured different proteolytic systems in incubated EDL muscles. The capacity for intralysosomal proteolysis, as measured by sensitivity to methylamine, was not increased in tumour-bearing rats, suggesting that the mechanism involved in the increased proteolytic rate was extralysosomal. Incubations using the Ca2+ ionophore A23187 revealed no change in the activity of calcium-dependent proteases as a consequence of tumour growth. Finally, muscle incubation in an ATP-depleted medium allowed us to conclude that energy-dependent proteases were involved in the activation of muscle proteolysis in tumour-bearing rats. In particular, the ubiquitin-dependent proteolytic system is involved, since there is an important increase in ubiquitin conjugates in the skeletal muscle of tumour-bearing rats. It may thus be suggested that extralysosomal ATP- and ubiquitin-dependent proteases underlie the biochemical mechanism of muscle wastage associated with cancer cachexia.

We investigated serum tryptophan (Trp), neopterin (NPT) and immunosuppressive acidic protein (IAP), one of tumor-associated tumor marker, concentrations in 28 patients with gastrointestinal tumors representing cancer cachexia and 10 healthy controls. NPT comes from activated macrophages presumably activated by tumor-sensitized T cells via gamma-interferon (IFN-gamma) excitation of the macrophages. We found that the NPT level was significantly higher than the control value. The negative correlation of NPT and Trp concentrations indicates activity of indoleamine 2,3-dioxygenase (IDO), a Trp degradating enzyme, in cancer-burden patients. The activity of IDO can be induced by cytokines such as IFN-gamma, and therefore low Trp levels may result from endogenous IFN-gamma production due to immune activation against tumors. We also found a positive correlation between NPT and IAP levels, suggesting that host immune activation against tumors played a role in the immunosuppression of cancer-burden states, followed by cancer-cachexia.

Physical exercise increases metabolic rate, and induces both adaptational biogenesis of mitochondria in skeletal muscle and an increase in antioxidant capacity. The onset of experimental anorexia and cachexia can be delayed by voluntary exercise. As skeletal muscle is the main target for cancer cachexia, we determined the levels of coenzymes Q9 and Q10 in skeletal muscle from tumour-bearing exercising rats, and compared them to those of sedentary tumour-bearers and controls. Both tumour-bearing groups had increased levels of coenzymes Q9 and Q10 in the anterior tibial muscle (P < 0.05 for exercised animals). In the soleus muscle, only the tumour-bearing exercising animals demonstrated an increase in the levels of both coenzymes (P < 0.05). In cardiac muscle, the presence of tumour and exercise reduced the levels of coenzymes below that of sedentary controls. Exercise counteracted the anaemia in the tumour-bearing host (P < 0.05). In conclusion, the increase in antioxidant capacity in skeletal muscle indicates a defence mechanism in the tumour-bearing hosts which is augmented by physical exercise.

We are currently using caprine arthritis encephalitis virus (CAEV) infection in goats as a model to understand changes in some clinical parameters and host response to infection with human immunodeficiency virus (HIV). The objective of this study was to measure changes in serum antioxidant activities in various age groups of goats infected with CAEV. Serum from CAEV-infected goats had significantly higher catalase activity (105.47 +/- 5.96 kU/l) than serum from healthy control goats (79.92 +/- 17.06 kU/l). Moreover, serum catalase activity increased with increase in the time after infection with CAEV. No change was observed in total superoxide dismutase (SOD) or glutathione peroxidase activity although CuZn SOD levels were elevated in infected goats. There was a positive correlation between serum catalase activity and hydrogen peroxide (H2O2) scavenging activity (r = 0.70, p < 0.05). In order to investigate cell membrane integrity, we determined lactate dehydrogenase (LDH) activity in infected goats. Although there was a transient increase in LDH no correlation was observed between increased serum catalase activity and LDH activity (r = 0.16, p > 0.05). We have earlier observed decreased oxyradical production in CAEV infected goats. This observed increase in serum catalase, a scavenger of endogenous free radicals such as H2O2 may be partly responsible for the observed decrease in oxygen radicals found in vivo.

Rates of lipogenesis and lipoprotein lipase (LPL) activity were measured in liver, adipose tissue, heart, and tumor at several stages during 10 days of palpable growth of a transplantable Leydig cell tumor in rats. This model showed the same characteristics as human cancer cachexia, including anorexia, weight loss, and muscle wasting. Comparison with pair-fed controls showed that the rate of loss of body fat was greater than could be explained by anorexia alone. The rate of lipogenesis tended to decrease during the later stages of tumor growth, particularly in the liver, where there was a statistically significant reduction on Days 5 and 10. This may be largely attributable to decreased availability of substrates caused by decreasing food intake and increasing glucose uptake by the tumor. There was a significant decrease in plasma glucose concentration by Day 10. In contrast, LPL activity in adipose tissue was depressed from the earliest stage of tumor growth, and this is likely to be a major cause of lipid depletion in cancer. There was no difference in adipose tissue LPL activity between the fed and postabsorptive states in the tumor-bearing rats, indicating that the normal response to nutrient intake was impaired. Thus, treatment of cancer cachexia should concentrate on normalizing the metabolic response to nutrient ingestion.

The relationship between serum and tumour cell surface proteolytic enzymes and the development of muscle breakdown in cancer cachexia has been studied in a murine model of the condition (MAC16). The surface of the MAC16 tumour cells carried a proteolytic enzyme referred to as guanidinobenzoatase (GB). Serum from mice also contained an enzyme (referred to as MSE) which cleaved the trypsin inhibitor 4-methylumbelliferyl-p-guanidinobenzoate as a true substrate, but there was no relationship with weight loss or the presence or absence of tumour and the level of this serum enzyme. Polyunsaturated fatty acids (PUFAs) were shown to be inhibitors of MSE at microM concentrations and one PUFA, eicosapentaenoic acid (EPA) was found to be a non-competitive inhibitor of both MSE and GB. The effect of EPA was specific since other proteolytic enzymes, trypsin, esterase and tissue plasminogen activator were unaffected by concentrations inhibiting GB and MSE. MSE and GB are two different enzymes which possess some common properties. However, GB is likely to be significant for tumour development since MSE is also found in normal mouse serum.