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Marked Increased Production of Acute Phase Reactants by Skeletal Muscle during Cancer Cachexia.

ABSTRACT: Loss of skeletal muscle mass in cancer cachexia is recognized as a predictor of mortality. This study aimed to characterize the changes in the muscle secretome associated with cancer cachexia to gain a better understanding of the mechanisms involved and to identify secreted proteins which may reflect this wasting process. The changes in the muscle proteome of the C26 model were investigated by label-free proteomic analysis followed by a bioinformatic analysis in order to identify potentially secreted proteins. Multiple reaction monitoring and Western blotting were used to verify the presence of candidate proteins in the circulation. Our results revealed a marked increased muscular production of several acute phase reactants (APR: Haptoglobin, Serine protease inhibitor A3N, Complement C3, Serum amyloid A-1 protein) which are released in the circulation during C26 cancer cachexia. This was confirmed in other models of cancer cachexia as well as in cancer patients. Glucocorticoids and proinflammatory cytokines are responsible for an increased production of APR by muscle cells. Finally, their muscular expressions are strongly positively correlated with body weight loss as well as the muscular induction of atrogens. Our study demonstrates therefore a marked increased production of APR by the muscle in cancer cachexia.

SUBMITTER: Massart IS

PROVIDER: S-EPMC7693727 | biostudies-literature | 2020 Oct

REPOSITORIES: biostudies-literature

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Marked Increased Production of Acute Phase Reactants by Skeletal Muscle during Cancer Cachexia.

Massart Isabelle S IS Paulissen Geneviève G Loumaye Audrey A Lause Pascale P Pötgens Sarah A SA Thibaut Morgane M MM Balan Estelle E Deldicque Louise L Atfi Azeddine A Louis Edouard E Gruson Damien D Bindels Laure B LB Meuwis Marie-Alice MA Thissen Jean-Paul JP

Cancers 20201031 11

Loss of skeletal muscle mass in cancer cachexia is recognized as a predictor of mortality. This study aimed to characterize the changes in the muscle secretome associated with cancer cachexia to gain a better understanding of the mechanisms involved and to identify secreted proteins which may reflect this wasting process. The changes in the muscle proteome of the C26 model were investigated by label-free proteomic analysis followed by a bioinformatic analysis in order to identify potentially sec ...[more]

PMID: 33142864

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Project description:Background Loss of skeletal muscle mass in cancer cachexia is recognized as an independent predictor of mortality. Mechanisms involved in this wasting process and parameters for early diagnosis are not yet clearly defined. As skeletal muscle is considered as a secretory organ, the aim of this present experimental work was to characterize the changes in the putative muscle secretome associated with cancer-induced cachexia to gain a better understanding of cellular mechanisms involved and to identify secreted proteins which might reflect this wasting process. Methods We investigated first the changes in the muscle proteome associated with cancer-induced cachexia by using differential label-free proteomic analysis on muscle of the C26 mouse model. The differentially abundant proteins were then submitted to sequential bioinformatic secretomic analysis in order to identify potentially secreted proteins. Multiple reaction monitoring and Western blotting were used to verify the presence of candidate proteins at the circulating level. Finally, we investigated the regulation of the production of these secreted proteins by muscle in vitro and in vivo. Results Our results revealed a dramatic increased muscular production (2-to 25-fold) of several acute phase reactants (APR: haptoglobin, serpina3n, complement C3, serum amyloid A1) which are released in the circulation during C26 cancer cachexia. This observation was confirmed in two other preclinical models of cancer cachexia as well as in cancer patients. The muscular origin of these APR was demonstrated by their increased expression in skeletal muscle and myotubes. Our results showed also that IL-6 plays a major role in the muscular induction of these APR in vivo, while glucocorticoids and pro-inflammatory cytokines stimulate directly their increased expression in muscle cells in vitro. Conclusions Muscle wasting caused by cancer is associated with marked changes in muscle secretome. Our study demonstrates a marked increased production of APR by skeletal muscle in pre-clinical models of cancer cachexia and in cancer patients. Further studies are required to unravel the potential role of these proteins in muscle atrophy and their interest as biomarkers of cancer cachexia.

Project description:Cachexia, or weight loss despite adequate nutrition, significantly impairs quality of life and response to therapy in cancer patients. In cancer patients, skeletal muscle wasting, weight loss and mortality are all positively associated with increased serum cytokines, particularly Interleukin-6 (IL-6), and the presence of the acute phase response. Acute phase proteins, including fibrinogen and serum amyloid A (SAA) are synthesized by hepatocytes in response to IL-6 as part of the innate immune response. To gain insight into the relationships among these observations, we studied mice with moderate and severe Colon-26 (C26)-carcinoma cachexia.Moderate and severe C26 cachexia was associated with high serum IL-6 and IL-6 family cytokines and highly similar patterns of skeletal muscle gene expression. The top canonical pathways up-regulated in both were the complement/coagulation cascade, proteasome, MAPK signaling, and the IL-6 and STAT3 pathways. Cachexia was associated with increased muscle pY705-STAT3 and increased STAT3 localization in myonuclei. STAT3 target genes, including SOCS3 mRNA and acute phase response proteins, were highly induced in cachectic muscle. IL-6 treatment and STAT3 activation both also induced fibrinogen in cultured C2C12 myotubes. Quantitation of muscle versus liver fibrinogen and SAA protein levels indicates that muscle contributes a large fraction of serum acute phase proteins in cancer.These results suggest that the STAT3 transcriptome is a major mechanism for wasting in cancer. Through IL-6/STAT3 activation, skeletal muscle is induced to synthesize acute phase proteins, thus establishing a molecular link between the observations of high IL-6, increased acute phase response proteins and muscle wasting in cancer. These results suggest a mechanism by which STAT3 might causally influence muscle wasting by altering the profile of genes expressed and translated in muscle such that amino acids liberated by increased proteolysis in cachexia are synthesized into acute phase proteins and exported into the blood.

Project description:Evidence from cachectic cancer patients and animal models of cancer cachexia supports the involvement of Forkhead box O (FoxO) transcription factors in driving cancer-induced skeletal muscle wasting. However, the genome-wide gene networks and associated biological processes regulated by FoxO during cancer cachexia are unknown. We hypothesize that FoxO is a central upstream regulator of diverse gene networks in skeletal muscle during cancer that may act coordinately to promote the wasting phenotype.To inhibit endogenous FoxO DNA-binding, we transduced limb and diaphragm muscles of mice with AAV9 containing the cDNA for a dominant negative (d.n.) FoxO protein (or GFP control). The d.n.FoxO construct consists of only the FoxO3a DNA-binding domain that is highly homologous to that of FoxO1 and FoxO4, and which outcompetes and blocks endogenous FoxO DNA binding. Mice were subsequently inoculated with Colon-26 (C26) cells and muscles harvested 26 days later.Blocking FoxO prevented C26-induced muscle fiber atrophy of both locomotor muscles and the diaphragm and significantly spared force deficits. This sparing of muscle size and function was associated with the differential regulation of 543 transcripts (out of 2,093) which changed in response to C26. Bioinformatics analysis of upregulated gene transcripts that required FoxO revealed enrichment of the proteasome, AP-1 and IL-6 pathways, and included several atrophy-related transcription factors, including Stat3, Fos, and Cebpb. FoxO was also necessary for the cancer-induced downregulation of several gene transcripts that were enriched for extracellular matrix and sarcomere protein-encoding genes. We validated these findings in limb muscles and the diaphragm through qRT-PCR, and further demonstrate that FoxO1 and/or FoxO3a are sufficient to increase Stat3, Fos, Cebpb, and the C/EBP? target gene, Ubr2. Analysis of the Cebpb proximal promoter revealed two bona fide FoxO binding elements, which we further establish are necessary for Cebpb promoter activation in response to IL-6, a predominant cytokine in the C26 cancer model.These findings provide new evidence that FoxO-dependent transcription is a central node controlling diverse gene networks in skeletal muscle during cancer cachexia, and identifies novel candidate genes and networks for further investigation as causative factors in cancer-induced wasting.

Dataset Information

Marked Increased Production of Acute Phase Reactants by Skeletal Muscle during Cancer Cachexia.

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Marked Increased Production of Acute Phase Reactants by Skeletal Muscle during Cancer Cachexia.

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