Skeletal muscle atrophy occurs in a variety of clinical settings, including cachexia, disuse, and denervation. Inflammatory cytokines have been shown to be mediators of cancer cachexia; however, the role of cytokines in denervation- and immobilization-induced skeletal muscle loss remains unknown. In this study, we demonstrate that a single cytokine, TNF-like weak inducer of apoptosis (TWEAK), mediates skeletal muscle atrophy that occurs under denervation conditions. Transgenic expression of TWEAK induces atrophy, fibrosis, fiber-type switching, and the degradation of muscle proteins. Importantly, genetic ablation of TWEAK decreases the loss of muscle proteins and spared fiber cross-sectional area, muscle mass, and strength after denervation. Expression of the TWEAK receptor Fn14 (fibroblast growth factor–inducible receptor 14) and not the cytokine is significantly increased in muscle upon denervation, demonstrating an unexpected inside-out signaling pathway; the receptor up-regulation allows for TWEAK activation of nuclear factor κB, causing an increase in the expression of the E3 ubiquitin ligase MuRF1. This study reveals a novel mediator of skeletal muscle atrophy and indicates that the TWEAK–Fn14 system is an important target for preventing skeletal muscle wasting.

TWEAK cytokine has been implicated in several biological responses including inflammation, angiogenesis, and osteoclastogenesis. We have investigated the role of TWEAK in regulating skeletal muscle mass. Addition of soluble TWEAK protein to cultured myotubes reduced the mean myotube diameter and enhanced the degradation of specific muscle proteins such as CK and MyHCf. The effect of TWEAK on degradation of MyHCf was stronger than its structural homologue, TNF‐α. TWEAK increased the ubiquitination of MyHCf and the transcript levels of atrogin‐1 and MuRF1 ubiquitin ligases. TWEAK inhibited phosphor‐ylation of Akt kinase and its downstream targets GSK‐3β, FOXO1, mTOR, and p70S6K Furthermore, TWEAK increased the activation of NF‐κB transcription factor in myotubes. Adenoviral‐mediated overexpression of IκBαΔN (a degradation‐resistant mutant of NF‐κB inhibitory protein ΙκBα) in myotubes blocked the TWEAK‐induced degradation of MyHCf. Chronic administration of TWEAK in mice resulted in reduced body and skeletal muscle weight with an associated increase in the activity of ubiquitin‐proteasome system and NF‐κB. Finally, muscle‐specific transgenic overexpression of TWEAK decreased the body and skeletal muscle weight in mice. Collectively, our data suggest that TWEAK induces skeletal muscle atrophy through inhibition of the PI3K/Akt signaling pathway and activation of the ubiquitin‐proteasome and NF‐kB systems.—Dogra, C., Changotra, H., Wedhas, N., Qin, X., Wergedal, J. E., Kumar, A. TNF‐related weak inducer of apoptosis (TWEAK) is a potent skeletal muscle‐wasting cytokine. FASEB J. 21, 1857–1869 (2007)

The nuclear factor NF-κB is a pleiotropic transcription factor whose activation results in inflammation, viral replication, and growth modulation. Due to its role in pathogenesis, NF-κB is considered a key target for drug development. In the present report we show that sanguinarine (a benzophenanthridine alkaloid), a known anti-inflammatory agent, is a potent inhibitor of NF-κB activation. Treatment of human myeloid ML-1a cells with tumor necrosis factor rapidly activated NF-κB, this activation was completely suppressed by sanguinarine in a dose- and time-dependent manner. Sanguinarine did not inhibit the binding of NF-κB protein to the DNA but rather inhibited the pathway leading to NF-κB activation. The reversal of inhibitory effects of sanguinarine by reducing agents suggests a critical sulfhydryl group is involved in NF-κB activation. Sanguinarine blocked the tumor necrosis factor-induced phosphorylation and degradation of IκBα, an inhibitory subunit of NF-κB, and inhibited translocation of p65 subunit to the nucleus. As sanguinarine also inhibited NF-κB activation induced by interleukin-1, phorbol ester, and okadaic acid but not that activated by hydrogen peroxide or ceramide, the pathway leading to NF-κB activation is likely different for different inducers. Overall, our results demonstrate that sanguinarine is a potent suppressor of NF-κB activation and it acts at a step prior to IκBα phosphorylation.