iTreg cells induce satellite cell expansion in vitro, but inhibit their differentiation.

<p>(A) Satellite cells were cultured in a myogenesis assay in the presence of iTreg, Th0, Tnaive and nTreg cells and evaluated for their ability to proliferate in the growth phase and to fuse to form myotubes in the differentiation phase (Red = BrdU, Green = MYOSIN HEAVY CHAIN (MyHC), Blue = DAPI, DNA). (B) After 3 days the number of satellite cells was significantly higher in the presence of iTregs (at the 1:50 and 1:100 ratios, n = 3 independent experiments, mean ± SEM, fold increase was calculated with reference to the satellite cell alone condition). (C) After 72 hours of co-culture, satellite cells were pulsed with BrdU for 2 hours and the percentage of BrdU<sup>+</sup> satellite cells was calculated (n = 3 independent experiments, mean ± SEM). (D). Satellite cells produced fewer elongated, multinucleated myotubes when induced to differentiate in the presence of iTregs. The fusion index (n of nuclei per cell) was calculated for satellite cells differentiated in presence of iTreg, Th0, Tnaive or nTreg. Both iTreg and Th0 cells inhibited satellite cell differentiation (1:50 ratio for all the conditions, n = 3 independent experiments, mean ± SEM; n = number of nuclei per cell).</p>