Preclinical models and in vitro experiments have provided valuable insight into the regulation of cancer-induced muscle wasting. Colon-26 (C26) tumor cells induce cachexia in mice, and conditioned media (CM) from these cells promotes myotube atrophy and catabolic signaling. While mechanical stimuli can prevent some effects of tumor-derived factors on myotubes, the impact of mechanical signaling on tumor-derived factor regulation of myosin heavy chain (MyHC) expression is not well understood. Therefore, we examined the effects of stretch-induced mechanical signaling on C2C12 myotube growth and MyHC expression after C26 CM exposure. C26 CM was administered to myotubes on day 5 of differentiation for 48 h. During the last 4 or 24 h of C26 CM exposure, 5% static uniaxial stretch was administered. C26 CM suppressed myotube growth and MyHC protein and mRNA expression. Stretch for 24 h increased myotube size and prevented the C26 CM suppression of MyHC-Fast protein expression. Stretch did not change suppressed MyHC mRNA expression. Stretch for 24 h reduced Atrogin-1/MAFbx, MuRF-1, and LC3B II/I ratio and increased integrin β1D protein expression and the myogenin-to-MyoD protein ratio. Stretch in the last 4 h of CM increased ERK1/2 phosphorylation but did not alter the CM induction of STAT3 or p38 phosphorylation. These results provide evidence that in myotubes pre-incubated with CM, the induction of mechanical signaling can still provide a growth stimulus and preserve MyHC-Fast protein expression independent of changes in mRNA expression.

Cancer cachexia is a chronic wasting disorder characterized by severe loss of skeletal muscle that affects approximately 80% of cancer patients, accounting for about 20 – 30% of cancer‐related deaths. Altered rest‐activity is a hallmark of circadian disruption and is often seen in cachectic patients. Additionally, loss of skeletal muscle‐specific clock gene results in muscle atrophy similar to that seen in cancer cachexia. However, it is unknown how skeletal muscle clock gene expression is altered with the progression of cachexia. Therefore, we investigated skeletal muscle clock gene expression during cachexia. ApcMin/+mice (MIN) were harvested in the morning and evening and were analyzed for clock gene expression. There was no effect of cancer on muscle Bmal1 (p = 0.55) and Clock (p = 0.59) expression. Cry1 expression showed no significant difference compared to BL – 6 mice (p = 0.24). Per 2 (p = 0.07) and Per 3 (p = 0.02) increased expression in the evening; however there was no difference between MIN and BL – 6 (Per 2 p=0.97, Per 3 p=0.87). Rev‐ERBα showed decreased expression in BL – 6 mice in the evening (p<0.001). However, there was no difference between the morning and evening MIN mice (p=0.31). RORα showed increased expression in BL – 6 mice in the evening (p = 0.03). Additionally, BL – 6 evening was significantly higher compared to MIN evening (p=0.01). However, there was no difference between the morning and evening MIN RORα expression (p>0.99). In LLC conditioned media treated myotubes, myotube diameter was decreased (p<0.001), and Bmal1 expression was decreased after 4 days in LLC conditioned media. In conclusion, the data suggest circadian clock expression is altered in a cachectic environment. Addition research is needed to understand the influence of these changes on the progression of cancer cachexia. Understanding the role of muscle‐specific circadian clock could lead to potential therapeutic interventions to attenuate cancer cachexia.