Amiloride ameliorates muscle wasting in cancer cachexia through inhibiting tumor-derived exosome release

Abstract Background Cancer cachexia (CAC) reduces patient survival and quality of life. Developments of efficient therapeutic strategies are required for the CAC treatments. This long-term process could be shortened by the drug-repositioning approach which exploits old drugs approved for non-cachexia disease. Amiloride, a diuretic drug, is clinically used for treatments of hypertension and edema due to heart failure. Here, we explored the effects of the amiloride treatment for ameliorating muscle wasting in murine models of cancer cachexia. Methods The CT26 and LLC tumor cells were subcutaneously injected into mice to induce colon cancer cachexia and lung cancer cachexia, respectively. Amiloride was intraperitoneally injected daily once tumors were formed. Cachexia features of the CT26 model and the LLC model were separately characterized by phenotypic, histopathologic and biochemical analyses. Plasma exosomes and muscle atrophy-related proteins were quantitatively analyzed. Integrative NMR-based metabolomic and transcriptomic analyses were conducted to identify significantly altered metabolic pathways and distinctly changed metabolism-related biological processes in gastrocnemius. Results The CT26 and LLC cachexia models displayed prominent cachexia features including decreases in body weight, skeletal muscle, adipose tissue, and muscle strength. The amiloride treatment in tumor-bearing mice distinctly alleviated muscle atrophy and relieved cachexia-related features without affecting tumor growth. Both the CT26 and LLC cachexia mice showed increased plasma exosome densities which were largely derived from tumors. Significantly, the amiloride treatment inhibited tumor-derived exosome release, which did not obviously affect exosome secretion from non-neoplastic tissues or induce observable systemic toxicities in normal healthy mice. Integrative-omics revealed significant metabolic impairments in cachectic gastrocnemius, including promoted muscular catabolism, inhibited muscular protein synthesis, blocked glycolysis, and impeded ketone body oxidation. The amiloride treatment evidently improved the metabolic impairments in cachectic gastrocnemius. Conclusions Amiloride ameliorates cachectic muscle wasting and alleviates cancer cachexia progression through inhibiting tumor-derived exosome release. Our results are beneficial to understanding the underlying molecular mechanisms, shedding light on the potentials of amiloride in cachexia therapy.

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Inflammation Based Regulation of Cancer Cachexia

BioMed Research International ◽

10.1155/2014/168407 ◽

2014 ◽

Vol 2014 ◽

pp. 1-7 ◽

Cited By ~ 63

Author(s):

Jill K. Onesti ◽

Denis C. Guttridge

Keyword(s):

Quality Of Life ◽

Skeletal Muscle ◽

Cancer Cachexia ◽

Molecular Mechanisms ◽

Muscle Wasting ◽

Nutritional Intake ◽

Clinical Implications ◽

Life Outcomes ◽

Skeletal Muscle Wasting

Cancer cachexia, consisting of significant skeletal muscle wasting independent of nutritional intake, is a major concern for patients with solid tumors that affects surgical, therapeutic, and quality of life outcomes. This review summarizes the clinical implications, background of inflammatory cytokines, and the origin and sources of procachectic factors including TNF-α, IL-6, IL-1, INF-γ, and PIF. Molecular mechanisms and pathways are described to elucidate the link between the immune response caused by the presence of the tumor and the final result of skeletal muscle wasting.

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Cancer-induced muscle wasting: latest findings in prevention and treatment

Therapeutic Advances in Medical Oncology ◽

10.1177/1758834017698643 ◽

2017 ◽

Vol 9 (5) ◽

pp. 369-382 ◽

Cited By ~ 69

Author(s):

Zaira Aversa ◽

Paola Costelli ◽

Maurizio Muscaritoli

Keyword(s):

Cancer Cachexia ◽

Molecular Mechanisms ◽

Muscle Wasting ◽

Early Recognition ◽

Poor Quality ◽

Future Research ◽

Pharmacological Interventions ◽

Metabolic Alterations ◽

Clinical Consequences

Cancer cachexia is a severe and disabling clinical condition that frequently accompanies the development of many types of cancer. Muscle wasting is the hallmark of cancer cachexia and is associated with serious clinical consequences such as physical impairment, poor quality of life, reduced tolerance to treatments and shorter survival. Cancer cachexia may evolve through different stages of clinical relevance, namely pre-cachexia, cachexia and refractory cachexia. Given its detrimental clinical consequences, it appears mandatory to prevent and/or delay the progression of cancer cachexia to its refractory stage by implementing the early recognition and treatment of the nutritional and metabolic alterations occurring during cancer. Research on the molecular mechanisms underlying muscle wasting during cancer cachexia has expanded in the last few years, allowing the identification of several potential therapeutic targets and the development of many promising drugs. Several of these agents have already reached the clinical evaluation, but it is becoming increasingly evident that a single therapy may not be completely successful in the treatment of cancer-related muscle wasting, given its multifactorial and complex pathogenesis. This suggests that early and structured multimodal interventions (including targeted nutritional supplementation, physical exercise and pharmacological interventions) are necessary to prevent and/or treat the devastating consequences of this cancer comorbidity, and future research should focus on this approach.

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Aerobic Exercise Ameliorates Cancer Cachexia-Induced Muscle Wasting through Adiponectin Signaling

International Journal of Molecular Sciences ◽

10.3390/ijms22063110 ◽

2021 ◽

Vol 22 (6) ◽

pp. 3110

Author(s):

Makoto Morinaga ◽

Naoki Sako ◽

Mari Isobe ◽

Sachiko Lee-Hotta ◽

Hideshi Sugiura ◽

...

Keyword(s):

Aerobic Exercise ◽

Muscle Atrophy ◽

Molecular Basis ◽

Cancer Cachexia ◽

Molecular Mechanisms ◽

Muscle Wasting ◽

C2c12 Myotubes ◽

Cellular Model ◽

Muscle Loss ◽

Colon 26

Cachexia is a multifactorial syndrome characterized by muscle loss that cannot be reversed by conventional nutritional support. To uncover the molecular basis underlying the onset of cancer cachectic muscle wasting and establish an effective intervention against muscle loss, we used a cancer cachectic mouse model and examined the effects of aerobic exercise. Aerobic exercise successfully suppressed muscle atrophy and activated adiponectin signaling. Next, a cellular model for cancer cachectic muscle atrophy using C2C12 myotubes was prepared by treating myotubes with a conditioned medium from a culture of colon-26 cancer cells. Treatment of the atrophic myotubes with recombinant adiponectin was protective against the thinning of cells through the increased production of p-mTOR and suppression of LC3-II. Altogether, these findings suggest that the activation of adiponectin signaling could be part of the molecular mechanisms by which aerobic exercise ameliorates cancer cachexia-induced muscle wasting.

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Altered expression of skeletal muscle myosin isoforms in cancer cachexia

AJP Cell Physiology ◽

10.1152/ajpcell.00154.2002 ◽

2002 ◽

Vol 283 (5) ◽

pp. C1376-C1382 ◽

Cited By ~ 69

Author(s):

Gary M. Diffee ◽

Katherine Kalfas ◽

Sadeeka Al-Majid ◽

Donna O. McCarthy

Keyword(s):

Muscle Atrophy ◽

Cancer Cachexia ◽

Muscle Wasting ◽

Tumor Cell Line ◽

Protein Isoforms ◽

Type I ◽

Myosin Isoforms ◽

Skeletal Muscle Myosin ◽

Altered Expression ◽

Isoform Expression

Cachexia is commonly seen in cancer and is characterized by severe muscle wasting, but little is known about the effect of cancer cachexia on expression of contractile protein isoforms such as myosin. Other causes of muscle atrophy shift expression of myosin isoforms toward increased fast (type II) isoform expression. We injected mice with murine C-26 adenocarcinoma cells, a tumor cell line that has been shown to cause muscle wasting. Mice were killed 21 days after tumor injection, and hindlimb muscles were removed. Myosin heavy chain (MHC) and myosin light chain (MLC) content was determined in muscle homogenates by SDS-PAGE. Body weight was significantly lower in tumor-bearing (T) mice. There was a significant decrease in muscle mass in all three muscles tested compared with control, with the largest decrease occurring in the soleus. Although no type IIb MHC was detected in the soleus samples from control mice, type IIb comprised 19% of the total MHC in T soleus. Type I MHC was significantly decreased in T vs. control soleus muscle. MHC isoform content was not significantly different from control in plantaris and gastrocnemius muscles. These data are the first to show a change in myosin isoform expression accompanying muscle atrophy during cancer cachexia.

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MuRF1/TRIM63, Master Regulator of Muscle Mass

International Journal of Molecular Sciences ◽

10.3390/ijms21186663 ◽

2020 ◽

Vol 21 (18) ◽

pp. 6663 ◽

Cited By ~ 1

Author(s):

Dulce Peris-Moreno ◽

Daniel Taillandier ◽

Cécile Polge

Keyword(s):

Skeletal Muscle ◽

Muscle Mass ◽

Muscle Atrophy ◽

Molecular Mechanisms ◽

Muscle Wasting ◽

Skeletal Muscle Atrophy ◽

Cardiac Functions ◽

Important Player ◽

Cardiac Muscles ◽

Inhibitory Molecules

The E3 ubiquitin ligase MuRF1/TRIM63 was identified 20 years ago and suspected to play important roles during skeletal muscle atrophy. Since then, numerous studies have been conducted to decipher the roles, molecular mechanisms and regulation of this enzyme. This revealed that MuRF1 is an important player in the skeletal muscle atrophy process occurring during catabolic states, making MuRF1 a prime candidate for pharmacological treatments against muscle wasting. Indeed, muscle wasting is an associated event of several diseases (e.g., cancer, sepsis, diabetes, renal failure, etc.) and negatively impacts the prognosis of patients, which has stimulated the search for MuRF1 inhibitory molecules. However, studies on MuRF1 cardiac functions revealed that MuRF1 is also cardioprotective, revealing a yin and yang role of MuRF1, being detrimental in skeletal muscle and beneficial in the heart. This review discusses data obtained on MuRF1, both in skeletal and cardiac muscles, over the past 20 years, regarding the structure, the regulation, the location and the different functions identified, and the first inhibitors reported, and aim to draw the picture of what is known about MuRF1. The review also discusses important MuRF1 characteristics to consider for the design of future drugs to maintain skeletal muscle mass in patients with different pathologies.

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The ubiquitin proteasome system in atrophying skeletal muscle: roles and regulation

AJP Cell Physiology ◽

10.1152/ajpcell.00125.2016 ◽

2016 ◽

Vol 311 (3) ◽

pp. C392-C403 ◽

Cited By ~ 57

Author(s):

Philippe A. Bilodeau ◽

Erin S. Coyne ◽

Simon S. Wing

Keyword(s):

Signaling Pathways ◽

Muscle Atrophy ◽

Molecular Mechanisms ◽

Muscle Wasting ◽

Clinical Problem ◽

Ubiquitin Proteasome System ◽

Mechanisms Of Action ◽

Loss Of Function ◽

Critical Determinant ◽

Ubiquitin Proteasome

Muscle atrophy complicates many diseases as well as aging, and its presence predicts both decreased quality of life and survival. Much work has been conducted to define the molecular mechanisms involved in maintaining protein homeostasis in muscle. To date, the ubiquitin proteasome system (UPS) has been shown to play an important role in mediating muscle wasting. In this review, we have collated the enzymes in the UPS whose roles in muscle wasting have been confirmed through loss-of-function studies. We have integrated information on their mechanisms of action to create a model of how they work together to produce muscle atrophy. These enzymes are involved in promoting myofibrillar disassembly and degradation, activation of autophagy, inhibition of myogenesis as well as in modulating the signaling pathways that control these processes. Many anabolic and catabolic signaling pathways are involved in regulating these UPS genes, but none appear to coordinately regulate a large number of these genes. A number of catabolic signaling pathways appear to instead function by inhibition of the insulin/IGF-I/protein kinase B anabolic pathway. This pathway is a critical determinant of muscle mass, since it can suppress key ubiquitin ligases and autophagy, activate protein synthesis, and promote myogenesis through its downstream mediators such as forkhead box O, mammalian target of rapamycin, and GSK3β, respectively. Although much progress has been made, a more complete inventory of the UPS genes involved in mediating muscle atrophy, their mechanisms of action, and their regulation will be useful for identifying novel therapeutic approaches to this important clinical problem.

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MicroRNAs in Skeletal Muscle and Hints on Their Potential Role in Muscle Wasting During Cancer Cachexia

Frontiers in Oncology ◽

10.3389/fonc.2020.607196 ◽

2020 ◽

Vol 10 ◽

Author(s):

Gioacchino P. Marceca ◽

Giovanni Nigita ◽

Federica Calore ◽

Carlo M. Croce

Keyword(s):

Skeletal Muscle ◽

Cancer Cachexia ◽

Muscle Wasting ◽

Muscle Protein ◽

Muscle Protein Synthesis ◽

Phenotypic Alteration ◽

Specific Tumor ◽

Non Coding Rnas

Cancer-associated cachexia is a heterogeneous, multifactorial syndrome characterized by systemic inflammation, unintentional weight loss, and profound alteration in body composition. The main feature of cancer cachexia is represented by the loss of skeletal muscle tissue, which may or may not be accompanied by significant adipose tissue wasting. Such phenotypic alteration occurs as the result of concomitant increased myofibril breakdown and reduced muscle protein synthesis, actively contributing to fatigue, worsening of quality of life, and refractoriness to chemotherapy. According to the classical view, this condition is primarily triggered by interactions between specific tumor-induced pro-inflammatory cytokines and their cognate receptors expressed on the myocyte membrane. This causes a shift in gene expression of muscle cells, eventually leading to a pronounced catabolic condition and cell death. More recent studies, however, have shown the involvement of regulatory non-coding RNAs in the outbreak of cancer cachexia. In particular, the role exerted by microRNAs is being widely addressed, and several mechanistic studies are in progress. In this review, we discuss the most recent findings concerning the role of microRNAs in triggering or exacerbating muscle wasting in cancer cachexia, while mentioning about possible roles played by long non-coding RNAs and ADAR-mediated miRNA modifications.

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Abstract P806: Sirtuin1 Plays a Critical Role in Reversing Skeletal Muscle Atrophy in Cerebral Ischemic Stroke

Stroke ◽

10.1161/str.52.suppl_1.p806 ◽

2021 ◽

Vol 52 (Suppl_1) ◽

Author(s):

Junaith S Mohamed ◽

Peter J Ferrandi ◽

Paez G Hector ◽

Christopher R Pitzer ◽

Stephen E Alway

Keyword(s):

Skeletal Muscle ◽

Ischemic Stroke ◽

Muscle Atrophy ◽

Muscle Wasting ◽

Rna Seq ◽

Stroke Patients ◽

Post Stroke ◽

Cerebral Ischemic Stroke ◽

Cerebral Ischemic

Stroke is a leading cause of mortality and long-term disability in patients worldwide. Skeletal muscle is the primary systemic target organ of stroke that severely induces muscle wasting and weakness, which contributes more to the long-term functional disability in stroke patients than any other disease. Currently, no approved pharmacological drug is available to treat stroke-induced muscle loss. Rehabilitative therapy is the only available option to improve muscle function in stroke patients. However, higher muscle fatigability and lower muscle strength from extensive muscle wasting in post-stroke patients provide poor rehabilitative outcomes. As a result, about two-thirds of stroke survivors persist in a state of insufficient recovery and experience physical disability that drastically reduces their health and quality of life. The major challenge in the drug discovery effort for treating post-stroke muscle wasting is the lack of our understanding of the molecular and/or cellular mechanisms that underlie the muscle wasting in stroke. To understand the molecular origin of stroke-induced muscle atrophy, gene expression profiling and associated biological pathway enrichment studies were performed in a mouse model of cerebral ischemic stroke using high-throughput RNA sequencing and extensive bioinformatic analyses. RNA-seq data revealed that the elevated atrophy in skeletal muscle observed in response to stroke was primairly associated with the altered expression of genes involved in the muscle protein degradation pathway. Further analysis of RNA-seq data identified Sirtuin1 (SirT1) as a critical protein that plays a significant role in regulating post-stroke muscle mass. SirT1 gain-of-function in skeletal muscle significantly reversed stroke-induced muscle atrophy via inhibiting the activation of the ubiquitin proteasomal pathway and restoring autophagy function. Collectively, this study identified suppression of SirT1as a novel mechanism by which stroke induces muscle atrophy.

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Leucine-rich Diet Improved Muscle Function in Cachectic Walker 256 Tumor-bearing Wistar Rats

10.21203/rs.3.rs-550186/v1 ◽

2021 ◽

Author(s):

Laís Viana ◽

Gabriela Chiocchetti ◽

Lucas Oroy ◽

Willians Vieira ◽

Carla Salgado ◽

...

Keyword(s):

Protein Degradation ◽

Muscle Atrophy ◽

Muscle Function ◽

Cancer Cachexia ◽

Muscle Protein ◽

Tumor Bearing ◽

Muscle Protein Degradation ◽

Walker 256 ◽

Walker 256 Tumor

Abstract Background: Skeletal muscle atrophy occurs in several pathological conditions such as cancer, a condition termed cancer cachexia. This condition is associated with an increase in morbidity and poor treatment response, decreasing quality of life, and increased mortality in cancer patients. A leucine-rich diet could be used as a coadjutant therapy preventing muscle atrophy in cancer cachexia hosts. Besides muscle atrophy, muscle function loss is even more important to the patient’s quality of life. Therefore, this study aimed to evaluate the effects of leucine-rich diet on muscle function activity of cachectic Walker 256 tumor-bearing rats and to correlate such effects with molecular pathways of muscle atrophy. Methods: Adult Wistar rats were randomly distributed into four experimental groups. Two groups were fed with a control diet: Control (C) and Walker 256 tumor-bearing (W), and two other groups were fed with a leucine-rich diet: Leucine Control (L) and Leucine Walker 256 tumor-bearing (LW). The functional analysis (walking, behavior, and strength tests) was measured and before and after tumor inoculation. Cachexia parameters such as body weight loss, muscle and fat mass, pro-inflammatory cytokine profile, and molecular and morphological aspects of skeletal muscle were also performed. Results: Walker 256 tumor growth led to muscle function decline, cachexia manifestation symptoms, muscle fiber cross-section area reduction, associated with the altered morphological pattern and classical muscle protein degradation pathway activation, with up-regulation of FoXO1, MuRF1, and 20S proteins. On the other hand, a leucine-rich diet improved muscle strength while reducing the decline of walking and behavior, partially improving the cachexia manifestations and preventing muscle atrophy and protein degradation in Walker 256 tumor-bearing rats. Conclusions: A leucine-rich diet diminished muscle protein degradation and enhanced oxidative pathways, leading to better muscle functional performance.

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Rapid Downregulation of c-Kit Expression Is Associated with Reduced Repopulation Potential of Donor Hematopoietic Stem Cells in Recipients after Total Body Irradiation.

Blood ◽

10.1182/blood.v104.11.1197.1197 ◽

2004 ◽

Vol 104 (11) ◽

pp. 1197-1197

Author(s):

Hongmei Shen ◽

Hui Yu ◽

Youzhong Yuan ◽

Paulina Huang ◽

Tao Cheng

Keyword(s):

Bone Marrow ◽

Molecular Mechanisms ◽

Absolute Number ◽

Post Transplantation ◽

Self Renewal ◽

Hematopoietic Stem ◽

Cell Divisions ◽

Total Body

Abstract Homing, lodgment, survival and proliferation are critical early determinants for the later outcomes of hematopoietic stem cell (HSC) or bone marrow transplantation (BMT). The irradiated bone marrow microenvironment may also pose an exhausting effect to the repopulating potential of donor HSCs, but the mechanisms for the effect are largely unknown. To determine whether these early events contribute to the exhausting effect, we have examined the kinetics of transplanted HSCs in 10 Gy lethally irradiated (IR) mice in comparison with transplanted HSCs in non-irradiated (NR) mice. 18 hours after transplantation, we found that the absolute number of homed Lin-Sca-1+ cells was not significantly different between IR and NR recipients. To examine the cell proliferative rate, CFSE staining together with flow cytometry was used to track the cell divisions of transplanted cells in the recipient marrow. While there were no detectable cell divisions in NR hosts, we detected 3 cell divisions in the Lin-Sca-1+ cell population 48 hours after BMT, thereby excluding the possibility that proliferation of hematopoietic cells was constrained in IR hosts. Regarding the expression of HSC associated markers, despite the similar expression of Sca-1 expression in both NR and IR recipients, the c-Kit was significantly downregulated to a nearly absent level in IR recipients, but it was not altered in NR recipients 18 hours post transplantation. The downregulation appeared to be transient since c-Kit was readily detectable after short-term engraftment. To functionally correlate c-Kit downregulation with long-term engraftment and self-renewal potential of transplanted HSCs, we sorted the homogeneous c-Kit+ cells (CD45.2+) and injected them into NR or IR recipients (CD45.1) at 5x106 cells/mouse. As expected, c-Kit became absent in IR hosts but not in NR hosts 18 hours after transplantation. We then harvested the homed cells and performed a competitive repopulation experiment involving the use of different congenic mice as secondary recipients at the dose of 1.3x 104 CD45.2 cells mixed with 1x105 competitive cells per mouse (n=4). Relative to the competitor cells (CD45.2/CD45.1 F1) in a same recipient, engraftment of the cells from IR recipients was lower than from NR recipients at each monthly time point (6 months). Moreover, the relative engraftment to competitor cells from IR recipients gradually declined to a minimal ratio of 0.03 while the engraftment from NR recipients sustained at a ratio of 0.3 after long-term engraftment. Finally, to further assess the self-renewal of the repopulated cells in the secondary recipients, 2 x 105 sorted CD45.2+ cells together with an equal number of competitor cells were re-transplanted into tertiary recipients. None of the mice (0/3) transplanted with cells originating from IR hosts were engrafted, but all mice (3/3) transplanted with the cells originating from NR recipients were engrafted as assessed at 6 months after tertiary transplantation. Given the previous studies by others showing that c-Kit signaling is involved in HSC lodgment and mobilization, we propose here that c-Kit downregulation in IR hosts impairs the lodging process of donor HSCs in the “niches” and as a consequence, the quality of the transplanted HSCs may be compromised. Therefore, further defining the molecular mechanisms for c-Kit downmodulation may guide us to develop novel approaches aimed to enhance the efficacy of HSC transplantation.

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