Differential effects of TNfα on satellite cell differentiation

[摘要] ENGLISH ABSTRACT: Tumour necrosis factor alpha (TNFα) is a pleiotropic cytokine and has a wide varietyof dose dependent cellular effects ranging from cell growth and differentiation, toinducing apoptosis. It has long been implicated in muscle and non-muscleinflammatory disorders, such as muscle wasting in chronic disease states, andrheumatoid arthritis. However, a physiological role for TNFα in muscle regenerationhas been proposed as elevated levels of the cytokine are present when muscleregeneration processes are initiated: TNFα is secreted by infiltrating inflammatorycells, and by injured muscle fibres. Adult skeletal muscle contains a population ofresident stem cell-like cells called satellite cells, which become activated, proliferateand differentiate following muscle injury to bring about repair of damaged muscle.Much research on the effects of TNFα on satellite cell differentiation has beenconducted in recent years. It is however difficult to get a complete characterisation ofthe cytokine's action as all models used slightly differ. We aimed therefore at providingcomprehensive assessment of the effects of increasing doses of chronicallysupplemented TNFα on differentiating C2C12 cells. Cells were allowed to differentiatewith or without TNFα supplementation for 7 days. Differentiation was induced at day0. The effect on differentiation was assessed at days 1, 3, 5, and 7 by western blotanalysis, and supplementary immunohistochemical analysis at days 1, 4, and 7 ofmarkers of differentiation - muscle regulatory factors: MyoD and myogenin, markers ofthe cell cycle p21, PCNA, and the integral signalling molecule, p38MAPK. TNFαsupplementation at day 1 tended to positively regulate early markers of differentiation.With continued supplementation however, markers of differentiation decreased dosedependently in treated cultures as the initial effect appeared to be reversed: A trendtowards a dose dependent decrease in MyoD, myogenin and p21 protein existed in treated cultures at days 3, 5, and 7. These findings were significant at day 5 (p21,p<0.05), and day 7 (myogenin, p<0.05). A significant dose dependent decrease in p38phosphorylation was evident at day 3 (p<0.05), while phospho-p38 was dosedependently increased at day 7 (p<0.05). Taken together, these data show that TNFαsupplementation for 24 hours following the induction of differentiation in vitro, tendsto increase levels of early markers of differentiation, and with continued TNFαsupplementation decrease markers of differentiation in a dose dependent fashion. Thisstudy provides a comprehensive characterisation of the dose and time dependent effectsof TNFα on satellite cell differentiaton in vitro. The model system used in the currentstudy, allows us to make conclusions on more chronic disease states.

[发布机构] Stellenbosch University