mouse skeletal myoblast derived cells c2c12  (ATCC)

Journal: Cells

Article Title: Mechanisms of Energy Metabolism in Skeletal Muscle Mitochondria Following Radiation Exposure

doi: 10.3390/cells8090950

Figure Lengend Snippet: Increased mtDNA content and expression of mitochondrial biogenesis-related genes in response to radiation exposure: ( A , B ) mtDNA content in C2C12 myotubes of control and 6-Gy IR-treated mice after 24 h, analyzed by assessing the relative levels of ND2 and gDNA by conventional PCR (A) and qPCR (B). ( C ) Representative immunoblot for COXIV and α-actin in control and 6-Gy IR-treated C2C12 myotubes after 24 h. ( D ) COXIV content quantified using image J software. ( E ) Fixed C2C12 myotubes were stained with an antibody against COXIV (green): PI (red) was used to stain the nuclei. Scale bar: 10 μm. ( F ) COXIV (green) content was quantified using image J software and normalized to that in control C2C12 myotubes. ( G ) Mitochondria content in control and 6-Gy IR-treated C2C12 myotubes after 24 h, quantified using the mitochondrial marker Mito Tracker Green. ( H ) Conventional PCR analysis of ACC-1, ACC-2, Glut-1, Glut-4, PGC-1, CPT-1, and UCP-2 mRNA in control and 6-Gy IR-treated C2C12 myotubes at the indicated times. Relative expression values were quantified using image J software and normalized to those in control C2C12 myotubes. Values are expressed as means ± SD ( n = 3; *** P < 0.001).

Article Snippet: Mouse C2C12 skeletal myoblast-derived cells (ATCC; Manassas, VA, USA) were cultured in Dulbecco’s Modified Eagle Medium (DMEM) supplemented with 10% fetal bovine serum (FBS) and 1% antibiotic mixture in a humidified atmosphere of 95% air and 5% CO 2 at 37 °C.

Techniques: Expressing, Control, Western Blot, Software, Staining, Marker

Journal: Cells

Article Title: Mechanisms of Energy Metabolism in Skeletal Muscle Mitochondria Following Radiation Exposure

doi: 10.3390/cells8090950

Figure Lengend Snippet: The generation of C2C12 myotubes revealed the ability of radiation exposure to improve mitochondrial respiration. ( A ) Kinetic oxygen consumption rate (OCR) responses of control and 6-Gy IR-treated C2C12 myotubes after 16 h to 2 μM oligomycin (ATP-coupled respiration), 5 μM FCCP (maximal respiratory capacity), and 1 μM rotenone (mitochondrial respiration). ( B – D ) Calculated ATP-coupled respiration (percent of oligomycin-sensitive OCR): (B) proton leak-linked respiration (percent of oligomycin-resistant, rotenone-sensitive mitochondrial OCR) (C) and maximal mitochondrial respiratory capacity (percent of rotenone-resistant OCR) (D) in control and 6-Gy IR-treated C2C12 myotubes. Each data point represents a mean ± SD ( n = 3; ** P < 0.01).

Article Snippet: Mouse C2C12 skeletal myoblast-derived cells (ATCC; Manassas, VA, USA) were cultured in Dulbecco’s Modified Eagle Medium (DMEM) supplemented with 10% fetal bovine serum (FBS) and 1% antibiotic mixture in a humidified atmosphere of 95% air and 5% CO 2 at 37 °C.

Techniques: Control

Journal: Cells

Article Title: Mechanisms of Energy Metabolism in Skeletal Muscle Mitochondria Following Radiation Exposure

doi: 10.3390/cells8090950

Figure Lengend Snippet: Measurement of glucose uptake, glycolysis, and glycolytic capacity of control and IR-treated C2C12 myotubes. ( A ) Glucose uptake was measured at the indicated times in control and 6-Gy IR-treated C2C12 myotubes and was normalized to protein concentration (means ± SD, n = 3; ** P < 0.01). ( B ) Kinetic extracellular acidification rate (ECAR) responses of control and 6-Gy IR-treated C2C12 myotubes after 16 h to 10 mM glucose, 2 μM oligomycin, and 50 mM 2-DG. ( C , D ) Calculated glycolysis (percent of glucose-sensitive ECAR) (C) and glycolytic capacity (percent of oligomycin-resistant, 2-deoxy-D-glucose (2-DG)-sensitive mitochondrial ECAR) (D) in control and 6-Gy IR-treated C2C12 myotubes. Each data point represents a mean ± SD ( n = 3).

Article Snippet: Mouse C2C12 skeletal myoblast-derived cells (ATCC; Manassas, VA, USA) were cultured in Dulbecco’s Modified Eagle Medium (DMEM) supplemented with 10% fetal bovine serum (FBS) and 1% antibiotic mixture in a humidified atmosphere of 95% air and 5% CO 2 at 37 °C.

Techniques: Control, Protein Concentration

Journal: Cells

Article Title: Mechanisms of Energy Metabolism in Skeletal Muscle Mitochondria Following Radiation Exposure

doi: 10.3390/cells8090950

Figure Lengend Snippet: Pyruvate, fatty acid, and amino acids fueled mitochondrial metabolism following radiation exposure. ( A ) Kinetic maximal baseline-normalized OCR responses of control and 6-Gy IR-treated C2C12 myotubes after indicated times, with or without 15 μM UK5099 (blocks the mitochondrial pyruvate carrier, MPC), 30 μM etomoxir (inhibits carnitine palmitoyl-transferase 1A, CPT-1A), and 30 μM BPTES (allosteric inhibitor of glutaminase, GLS1). ( B ) Schematic illustration of cellular metabolism (glucose oxidation, long-chain fatty acid oxidation, and glutamine oxidation) pathways and assays for pyruvate, palmitate, and glutamate/glutamine. ( C ) Fuel oxidation was measured at the indicated times in control and 6-Gy IR-treated C2C12 myotubes. Each data point represents a mean ± SD ( n = 3; * P < 0.05, ** P < 0.01).

Article Snippet: Mouse C2C12 skeletal myoblast-derived cells (ATCC; Manassas, VA, USA) were cultured in Dulbecco’s Modified Eagle Medium (DMEM) supplemented with 10% fetal bovine serum (FBS) and 1% antibiotic mixture in a humidified atmosphere of 95% air and 5% CO 2 at 37 °C.

Techniques: Control

Journal: Cells

Article Title: Mechanisms of Energy Metabolism in Skeletal Muscle Mitochondria Following Radiation Exposure

doi: 10.3390/cells8090950

Figure Lengend Snippet: The activation of AMPK maintained bioenergetic homeostasis in C2C12 myotubes following radiation exposure. ( A ) Reactive oxygen species (ROS), ( B ) ATP, and ( C ) total NAD and NADH levels in C2C12 myotubes with or without 6-Gy radiation exposure were measured after 24 h and were normalized to protein concentration. Data are presented as means ± SD ( n = 3; * P < 0.05, ** P < 0.01). ( D ) Schematic illustration of cellular molecular pathways. ( E ) Representative immunoblot for p-ACC (Ser79), total ACC, p-AMPK (Thr172), and total AMPK in control and 6-Gy IR-treated C2C12 myotubes after 24 h. ( F ) Representative immunoblot for nuclear PGC-1 and U170s in control and 6-Gy IR-treated C2C12 myotubes after 24 h. ( G ) Representative immunoblot for mitochondrial CPT-1, UCP-2, and Bcl2 in control and 6-Gy IR-treated C2C12 myotubes after 24 h. ( H ) Representative immunoblot for plasma membrane Glut1, Glut4, and EGFR in control and 6-Gy IR-treated C2C12 myotubes after 24 h.

Article Snippet: Mouse C2C12 skeletal myoblast-derived cells (ATCC; Manassas, VA, USA) were cultured in Dulbecco’s Modified Eagle Medium (DMEM) supplemented with 10% fetal bovine serum (FBS) and 1% antibiotic mixture in a humidified atmosphere of 95% air and 5% CO 2 at 37 °C.

Techniques: Activation Assay, Protein Concentration, Western Blot, Control, Clinical Proteomics, Membrane