Murine C2C12 skeletal myoblasts (ATCC, Manassas VA) were grown as described previously (15 (link)) in high-glucose DMEM supplemented by 10% FBS, 100 U/ml penicillin, 100 μg/ml streptomycin, 100 μg/ml sodium pyruvate, 2 mmol L-glutamine, and maintained at 37°C in 5% CO2 in air. Differentiation of myotubes was induced by switching subsonfluent myoblasts to differentiation medium DMEM supplemented by 2% horse serum. The differentiation medium was replaced every other day for up to 5 days. Fully differentiated C2C12 myotubes were exposed for 48 h to 1 or 5% serum from patients enrolled in a randomized controlled trial of pamidronate. In order to determine the effect of transforming growth factor (TGF)-β pan neutralizing antibody on fiber size myotubes were exposed for 48 h to 10 μg/ml of anti-TGFβ-1/2/3, clone 1D11.16.8 (BioXcell West, Lebanon NH) in the presence or absence of burned patients' serum.
Murine c2c12 skeletal myoblasts
Manufactured by American Type Culture Collection
Sourced in United States
Murine C2C12 skeletal myoblasts are a widely used cell line derived from mouse skeletal muscle. They are capable of differentiating into myotubes, making them a valuable tool for the study of skeletal muscle development and function.
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Lab products found in correlation
Streptomycin, by Thermo Fisher Scientific (3 mentions)
Dmem ham s f12 medium, by Thermo Fisher Scientific (2 mentions)
Penicillin g, by Thermo Fisher Scientific (2 mentions)
Incb018424, by Merck Group (1 mentions)
Fetal bovine serum (fbs), by GE Healthcare (1 mentions)
Dulbecco modified eagle medium (dmem), by Thermo Fisher Scientific (1 mentions)
Fetal bovine serum (fbs), by Thermo Fisher Scientific (1 mentions)
Fetal bovine serum (fbs), by Merck Group (1 mentions)
Penicillin, by Thermo Fisher Scientific (1 mentions)
13 protocols using murine c2c12 skeletal myoblasts
1
Murine C2C12 Myoblast Differentiation
2
Bone-derived Factors Regulate Myotube Size
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Murine C2C12 skeletal myoblasts (ATCC, Manassas, VA) were grown in high glucose DMEM supplemented with 10% FBS, 100 U/ml penicillin, 100 mg/ml streptomycin, 100 mg/ml sodium pyruvate, 2 mM L-glutamine, and maintained at 37°C in 5% CO2, as shown in Pin et al. (40 (link)). Myotubes were generated by exposing the myoblasts to DMEM containing 2% horse serum (i.e., differentiation medium, DM), and replacing the medium every other day for 5 days. In order to determine the effects on myotube size dependent on bone-derived factors, myotubes were exposed to 20% bone conditioned medium (CM) for up to 48 h.
3
Differentiation of C2C12 Myoblasts into Myotubes
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Murine C2C12 skeletal myoblasts (ATCC, Manassas, VA, USA) were grown in high-glucose DMEM supplemented with 10% fetal bovine serum (FBS), 100 U/mL of penicillin, 100 mg/mL of streptomycin, 100 mg/mL of sodium pyruvate and 2 mM of L-glutamine (growth medium; GM) and maintained at 37 °C in a humidified atmosphere of 5% CO2 in air. Myoblasts were seeded on 12-well plates at a density of 30,000/cm2 in GM. To induce differentiation, after 24 h (sub-confluency), GM was replaced by DMEM containing 100 U/mL of penicillin, 100 mg/mL of streptomycin, 100 mg/mL of sodium pyruvate and 2 mM of L-glutamine, supplemented with 2% horse serum (differentiation medium; DM). The medium was changed every 48 h. After 5 days of culture in DM, mature myotubes aligned to form regular bundles.
4
Murine C2C12 Myoblast Differentiation
5
Murine C2C12 Myotube Growth Regulation
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Murine C2C12 skeletal myoblasts (ATCC, Manassas, VA, USA) were grown in high glucose DMEM supplemented with 10% FBS, 100 U/ml penicillin, 100 mg/ml streptomycin, 2 mM l -glutamine, and maintained at 37 °C in 5% CO2, as previously published [25 (link)]. Myotubes were generated by exposing the myoblasts to DMEM containing 2% horse serum (i.e., differentiation medium), and replacing the medium every other day for 5 days. In order to determine the dependence of myotube size on bone-derived factors, myotubes were exposed to 5% bone conditioned medium (CM) for 48 h. Cells were fixed and stained [26 (link), 27 ].
6
Investigating Tumor-Derived Effects on Myotube Size
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Human ES‐2 cells were purchased from ATCC (Manassas, VA, USA; #CRL‐1978) and were cultured in McCoy's medium supplied with 10% foetal bovine serum, 1% glutamine, 1% sodium pyruvate, 1% penicillin/streptomycin, in a 5% CO2, 37°C humidified incubator. Murine C2C12 skeletal myoblasts (ATCC) were grown in high glucose Dulbecco's modified Eagle's medium (DMEM) supplemented with 10% foetal bovine serum, 100 U/mL penicillin, 100 mg/mL streptomycin, 100 mg/mL sodium pyruvate, and 2 mM L‐glutamine and maintained at 37°C in 5% CO2. Myotubes were generated by exposing the myoblasts to DMEM containing 2% horse serum and replacing the medium every other day for 5 days. In order to determine the effects of tumor‐derived mediators on fibre size, myotubes were exposed to 25% or 50% CM, collected from confluent ES‐2 culture plates and subjected to centrifugation to remove cell debris. The JAK1/2 pharmacologic inhibitor INCB018424 (EMD Millipore, Burlington, MA, USA) was dissolved in sterile Dimethyl sulfoxide (DMSO) and administered in the culture medium at 400 nM final concentration for up to 48 h, as previously shown.15
7
Bone Factors Regulate Myotube Size
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Murine C2C12 skeletal myoblasts (ATCC, Manassas, VA, USA) were grown in high‐glucose Dulbecco's modified Eagle medium (DMEM) supplemented with 10% FBS, 100 U/mL penicillin, 100 mg/mL streptomycin, 2mM L‐glutamine, and maintained at 37°C in 5% CO2, as published.(49 ) Myotubes were generated by exposing the myoblasts to DMEM containing 2% horse serum (ie, differentiation medium [DM]), and replacing the medium every other day for 3 days. In order to determine the dependence of myotube size on bone‐derived factors, myotubes were exposed to 5% bone conditioned medium (CM) for 48 hours. Cells were fixed and stained as described.(37 )
8
Differentiation of Murine C2C12 Myoblasts
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Murine C2C12 skeletal myoblasts (CRL-1772, ATCC, Virginia, USA) were expanded in growth medium, which consisted of Dulbecco's Modified Eagle Medium (DMEM high glucose, L-glutamine) (11965092, Gibco, Thermofisher, Massachusetts, USA) supplemented with fetal bovine serum (10%, FBS) (16000044, Thermofisher, Massachusetts, USA) and penicillin/streptomycin (1%) (15140122, Thermofisher, Massachusetts, USA) at 37 ºC and 5% CO2 atmosphere. To induce differentiation into myotubes, the medium was changed to a differentiation medium, based in DMEM high glucose, supplemented with horse serum (2%) (HS) (26050088, Thermofisher, Massachusetts, USA) and 1% penicillin/streptomycin.
9
Culturing Murine C2C12 Myoblasts
10
Transdifferentiation of Healthy and DM1 Fibroblasts
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Healthy immortalized control (CNT) [33 (link)] and patient-derived (DM1) fibroblasts carrying 1300 CTG repeats were provided by Dr. Furling (Institute of Myology, Paris, France) and were transdifferentiated into myotubes by inducing MyoD expression. The fibroblasts were grown in Dulbecco’s Modified Eagle Medium (DMEM, 4.5 g/L glucose, Gibco), 1% penicillin and streptomycin (P/S, 10,000 U/mL, Thermo Fisher, Waltham, MA, USA), and 10% fetal bovine serum (FBS, Sigma-Aldrich, Saint Louis, MI, USA). Transdifferentiation was prompted by muscle differentiation medium (MDM) containing DMEM with 4.5 g/L glucose, 1% P/S, 2% horse serum, 1% apo-transferrin (10 mg/mL), 0.1% insulin (10 mg/mL), and 0.02% doxycycline (10 mg/mL). Murine C2C12 skeletal myoblasts, purchased from the American Type Culture Collection (CRL-1772, ATCC, Manassas, VA, USA), were grown in DMEM (4.5 g/L glucose, Gibco, Thermo Fisher, Waltham, MA, USA ) supplemented with 1% P/S (10,000 U/mL, Thermo Fisher, Waltham, MA, USA) and 10% FBS (Gibco, Thermo Fisher, Waltham, MA, USA). All cells were grown at 37 °C in a humidified atmosphere containing 5% CO2.
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