STAT3 Activation in Skeletal Muscle Links Muscle Wasting and the Acute Phase Response in Cancer Cachexia

Abstract

BackgroundCachexia, 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.Methodology/Principal FindingsModerate 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.Conclusions/SignificanceThese 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.

Elevated SOCS3 and altered IL-6 signaling is associated with age-related human muscle stem cell dysfunction

“…In agreement with the observed elevation of muscle and systemic IL-6, muscle SOCS3 protein was 86% higher in older muscle. These alterations have been noted in previous studies and appear to be associated with normal aging (30,57), which are further elevated in disease states such as cancer cachexia (5). In animals, 14 days of IL-6 infusion increased the transcription of atrogin, SOCS3, and TNF-␣, which promoted a proinflammatory state and blunted muscle growth in young animals and induced atrophy in adults (3,25).…”

Assessment of muscle mass and strength in mice

“…16 During differentiation, the originating myotubes progressively express actin and myosin, whereas myoblasts do not normally show expression of these proteins. Here below is a brief protocol for the use of C2C12 as a model for the study of muscle atrophy in vitro 3,17 (Figure 1d). These cells require passage every 1-2 days.…”

“…We previously took advantage of this model to investigate the mechanisms that cause myotube atrophy in the presence of pro-inflammatory cytokines, such as interleukin-6 or tumor necrosis factor, previously shown to be causative of muscle wasting in both in vitro and in vivo conditions. 3,17,18 In order to do so, the cell layer is generally exposed to pro-catabolic cytokines (100 ng ml À 1 ), with or without specific inhibitors/antagonists. Typically, muscle atrophy in the myotubes can be assessed at several time points, early (30 min, 1, 2, 6 h), as well as late ones (24, 48, 72 h).…”

“…Proceed to assess the myotube size (for example, the myotubes minimum diameter) by using ImageJ (or similar software; Figure 1d). 3,17 In Vivo Assessment of Muscle Strength…”

Mechanism-Based Therapeutic Approaches to Cachexia

“…Recently, the relation between the acute phase response (systemic inflammation) and cancer cachexia was proposed from the observation that enhanced liver protein synthesis could drive muscle protein hypercatabolism contributing to the elevated resting energy expenditure (Durham et al, 2009). Not only, the skeletal muscle itself was proposed as a source of acute phase reactants, likely diverting aminoacids from the synthesis of structural proteins to that of acute phase mediators (Bonetto et al, 2011). Of interest, acute phase proteins are further increased by nutritional supplementation, suggesting that anti-inflammatory treatments should be included in the therapeutic protocol (Stephens et al, 2008).…”