Skeletal muscle interleukin-6 regulation in hyperthermia

We previously reported that IL-6 production is acutely elevated in skeletal muscles exposed to ≥41°C, but the regulatory pathways are poorly understood. The present study characterizes the heat-induced transcriptional control of IL-6 in C2C12 muscle fibers. Hyperthermia exposure (42°C for 1 h) induced transcription from an IL-6 promoter-luciferase reporter plasmid. Heat shock factor-1 (HSF-1), a principal mediator of the heat shock response, was then tested for its role in IL-6 regulation. Overexpression of a constitutively active HSF-1 construct increased basal (37°C) promoter activity, whereas overexpression of a dominant negative HSF-1 reduced IL-6 promoter activity during basal and hyperthermia conditions. Since hyperthermia also induces stress-activated protein kinase (SAPK) signaling, we tested whether mutation of a transcription site downstream of SAPK, (i.e., activator protein-1, AP-1) influences IL-6 transcription in hyperthermia. The mutation had no effect on baseline reporter activity but completely inhibited heat-induced activity. We then tested whether pharmacologically induced states of protein stress, characteristic of cellular responses to hyperthermia and known to induce SAPKs and HSF-1, would induce IL-6 production in the absence of heat. The proteasome was inhibited with MG-132 in one set of experiments, and the unfolded protein response was stimulated with dithiothreitol, thapsigargin, tunicamycin, or castanospermine in other experiments. All treatments stimulated IL-6 protein secretion in the absence of hyperthermia. These studies demonstrate that IL-6 regulation in hyperthermia is directly controlled by HSF-1 and AP-1 signaling and that the IL-6 response in C2C12 myotubes is sensitive to categories of protein stress that reflect accumulation of damaged or unfolded proteins.