Melbourne Veterinary School - Research Publications
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ItemCaspase-3, myogenic transcription factors and cell cycle inhibitors are regulated by leukemia inhibitory factor to mediate inhibition of myogenic differentiation(BIOMED CENTRAL LTD, 2011)BACKGROUND: Leukemia inhibitory factor (LIF) is known to inhibit myogenic differentiation as well as to inhibit apoptosis and caspase-3 activation in non-differentiating myoblasts. In addition caspase-3 activity is required for myogenic differentiation. Therefore the aim of this study was to further investigate mechanisms of the differentiation suppressing effect of LIF in particular the possibility of a caspase-3 mediated inhibition of differentiation. RESULTS: LIF dependent inhibition of differentiation appeared to involve several mechanisms. Differentiating myoblasts that were exposed to LIF displayed increased transcripts for c-fos. Transcripts for the cell cycle inhibitor p21 as well as muscle regulatory factors myoD and myogenin were decreased with LIF exposure. However, LIF did not directly induce a proliferative effect under differentiation conditions, but did prevent the proportion of myoblasts that were proliferating from decreasing as differentiation proceeded. LIF stimulation decreased the percentage of cells positive for active caspase-3 occurring during differentiation. Both the effect of LIF inhibiting caspase-3 activation and differentiation appeared dependent on mitogen activated protein kinase and extracellular signal regulated kinase kinase (MEK) signalling. The role of LIF in myogenic differentiation was further refined to demonstrate that myoblasts are unlikely to secrete LIF endogenously. CONCLUSIONS: Altogether this study provides a more comprehensive view of the role of LIF in myogenic differentiation including LIF and receptor regulation in myoblasts and myotubes, mechanisms of inhibition of differentiation and the link between caspase-3 activation, apoptosis and myogenic differentiation.
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ItemAn anti-inflammatory role for leukemia inhibitory factor receptor signaling in regenerating skeletal muscle(SPRINGER, 2013-01)Skeletal muscle regeneration in pathology and following injury requires the coordinated actions of inflammatory cells and myogenic cells to remove damaged tissue and rebuild syncytial muscle cells, respectively. Following contusion injury to muscle, the cytokine leukemia inhibitor factor (LIF) is up-regulated and knockout of Lif negatively impacts on morphometric parameters of muscle regeneration. Although it was speculated that LIF regulates muscle regeneration through direct effects on myogenic cells, the inflammatory effects of LIF have not been examined in regenerating skeletal muscle. Therefore, the expression and function of LIF was examined using the antagonist MH35-BD during specific inflammatory and myogenic stages of notexin-induced muscle regeneration in mice. LIF protein and mRNA were up-regulated in two distinct phases following intramuscular injection of notexin into tibialis anterior muscles. The first phase of LIF up-regulation coincided with the increased expression of pro-inflammatory cytokines; the second phase coincided with myogenic differentiation and formation of new myotubes. Administration of the LIF receptor antagonist MH35-BD during the second phase of LIF up-regulation had no significant effects on transcript expression of genes required for myogenic differentiation or associated with inflammation; there were no significant differences in morphometric parameters of the regenerating muscle. Conversely, when MH35-BD was administered during the acute inflammatory phase, increased gene transcripts for the pro-inflammatory cytokines Tnf (Tumor necrosis factor), Il1b (Interleukin-1β) and Il6 (Interleukin-6) alongside an increase in the number of Ly6G positive neutrophils infiltrating the muscle were observed. This was followed by a reduction in Myog (Myogenin) mRNA, which is required for myogenic differentiation, and the subsequent number of myotubes formed was significantly decreased in MH35-BD-treated groups compared to sham. Thus, antagonism of the LIF receptor during the inflammatory phase of skeletal muscle regeneration appeared to induce an inflammatory response that inhibited subsequent myotube formation. We propose that the predominant role of LIF in skeletal muscle regeneration appears to be in regulating the inflammatory response rather than directly effecting myogenic cells.