Cc 3244

Reagents were obtained from Sigma (Gillingham, UK) unless stated otherwise. Primary myoblasts obtained from healthy donors (CC‐2580; Lonza, Slough, UK) were maintained in-house in Skeletal Muscle Basal Medium‐2 (Lonza; CC‐3244 and CC‐3246) containing 0.1% human epidermal growth factor, 2% l‐glutamine, 10% fetal bovine serum (FBS), and 0.1% gentamicin/amphotericin‐B in the absence of GCs. Confluent myotubes were differentiated in Dulbecco’s modified Eagle’s medium (DMEM) containing 2% horse serum (HS) for 120 h. Media were replaced every 2–3 days as previously reported (16 (link)).

Reagents were obtained from Sigma (Gillingham, UK) unless stated otherwise. Primary myoblasts (CC‐2580; Lonza, Slough, UK) were maintained in house in Skeletal Muscle Basal Medium‐2 (Lonza; CC‐3244 and CC‐3246) containing 0.1% human epidermal growth factor, 2% l‐glutamine, 10% fetal bovine serum (FBS) and 0.1% gentamicin/amphotericin‐B in the absence of GCs. Confluent myotubes were differentiated in Dulbecco's modified Eagle's medium (DMEM) containing 2% horse serum (HS) for 120 h. Media were replaced every 2–3 days 25.

RMS cell lines (CW9019, RD and RH4) were kindly provided by Dr. Oscar Martínez-Tirado and Dr. Cristina Muñoz-Pinedo from the Biomedical Research Institute from Bellvitge (IDIBELL). C2C12 cells and Human skeletal muscle myoblasts (HSMM) were purchased at ATCC (ATCC^® CRL-1772™, ATCC, Manassas, Virginia, USA) and Lonza (CC-2580, Lonza, Basel, Switzerland), respectively. RMS cell lines were maintained in RPMI 1640 medium (31870, Thermo Fisher, Gibco, Paisley, Scotland) and supplemented with 1% of L-Glutamine (25030, Thermo Fisher, Gibco) and 1% of penicillin and streptomycin (15140, Thermo Fisher, Gibco) and 10% heat-inactivated fetal bovine serum (10270, Thermo Fisher, Gibco). C2C12 cells were cultured in DMEM high glucose medium (41965, Thermo Fisher, Gibco) supplemented with 10% heat-inactivated fetal bovine serum and 1% of penicillin and streptomycin. HSMM cells were maintained in SKBM-2 medium (CC-3246, Lonza) supplemented with its specific SingleQuots^TM and growth factors (CC-3244, Lonza). All cells were kept at 37 °C in a humidified atmosphere of 5% CO₂. In addition, all of them were routinely tested for mycoplasma. Drug treatments were carried out directly in the culture media. Doses and time points are indicated in every single experiment. All drugs were purchased at Selleckchem (Munich, Germany).