Long-term cultured human myotubes decrease contractile gene expression and regulate apoptosis-related genes.
The present study examined time-dependent changes in the gene expression profile of long-term cultured human myotubes. Microarray transcriptional analysis was performed in a primary culture of differentiated myotubes from one subject over seven weeks. This analysis showed a main gradual fall in genes of the contractile apparatus, and a broad upregulation of genes involved in cell development and growth, followed by stress response and signal transduction. Glucose metabolism was also monitored, but no significant alterations in glucose uptake, oxidation or glycogen storage were observed. Mitochondrial membrane potential, or the amount of membrane lipid peroxides, remained similarly unchanged, nor was lactate dehydrogenase leakage observed. Time-dependent changes in eight genes were validated by real-time RT-PCR in primary cultured myotubes from four subjects, of similar age and isolated after equivalent replication cycles in vitro and differentiated over seven weeks. Insulin-like growth factor-binding protein 2 (IGFBP2), a modulator of the IGF signal, was upregulated. The antiapoptotic gene heat-shock 70-kd protein 2 (HSPA2) was induced, whereas the proapoptotic tumor necrosis factor receptor superfamily, member 25 (WSL-1) was suppressed. A decline in the muscle-specific gene M-cadherin and contraction genes, such as slow-twitch troponin I (TNNI1) and myosin heavy chain 2 (MYH2), myosin light chain 1 (MYL1) and myosin-binding protein H (MYBPH), which are expressed in adult fast-twitch muscle, was shown. In summary, these data demonstrate extensive downregulation of contractile genes and modulation of apoptosis-related genes, in favour of cell survival, during maintenance of cultured human myotubes.
Ferrer-Martínez
Andreu
A
Departament de Bioquímica i Biologia Molecular, Facultat de Biologia, Universitat de Barcelona, Diagonal 645, 08028-Barcelona, Spain.
Montell
Eulàlia
E
Montori-Grau
Marta
M
García-Martínez
Cèlia
C
Gómez-Foix
Anna M
AM
Roberts
Matthew-Alan
MA
Mansourian
Robert
R
Macé
Katherine
K
D000293:Adolescent; D017209:Apoptosis / Q000235:genetics*; D001706:Biopsy; D018929:Cell Culture Techniques; D002470:Cell Survival / Q000235:genetics; D002478:Cells, Cultured; D002648:Child; D015536:Down-Regulation; D020869:Gene Expression Profiling; D005947:Glucose / Q000378:metabolism; D006801:Humans; D050356:Lipid Metabolism / Q000235:genetics; D053078:Membrane Potential, Mitochondrial; D018485:Muscle Fibers, Skeletal / Q000378:metabolism*; D009124:Muscle Proteins / Q000235:genetics*; D009132:Muscles / Q000166:cytology / Q000473:pathology; D020411:Oligonucleotide Array Sequence Analysis; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013995:Time
D009124:Muscle Proteins; D005947:Glucose