TY - JOUR
T1 - Gene expression and secretion of insulin-like growth factor-binding proteins during myoblast differentiation
AU - Ernst, Catherine W.
AU - McCusker, Robert H.
AU - White, Michael E.
PY - 1992/2
Y1 - 1992/2
N2 - Numerous cell types have been reported to secrete insulin-like growth factor-binding proteins (IGFBPs) in vitro. We have observed such IGFBPs in culture medium conditioned by the mouse myoblast cell line C2C12 and have identified IGFBP-2 among the secreted IGFBPs. Since C2C12 cells fuse in culture, these cells were used to examine IGFBP-2 mRNA expression and protein secretion during myogenic differentiation. Cells were harvested at approximately 80% of confluent density. Additional cultures were rinsed, fed differentiation medium, and harvested when approximately 15%, 60%, and 85% differentiated (fused). Northern and dot blot analyses were performed using total cellular RNA and a labeled cDNA specific for rat IGFBP-2. A single mRNA transcript of approximately 1.8 kilobases was observed. The level of expression of IGFBP-2 mRNA was highest in proliferating cells and decreased to 35%, 20%, and less than 10% of initial levels as differentiation progressed. Serum-free medium was conditioned for 24 h at each time point and collected from similar cultures. Three IGFBP species of 32,000, 30,000, and 24,000 mol wt (Mr) were detected in conditioned medium by probing Western blots with [125I]IGF-I (ligand blot analysis). The intensity of the 32,000 Mr band decreased with differentiation. These same blots were probed with an antibody raised against the 34,000 Mr bovine IGFBP-2. This antibody specifically bound to only the 32,000 Mr IGFBP. The level of antibody binding decreased by 50%, 90%, and nearly 100% as differentiation progressed. It, therefore, appears that IGFBP-2 is expressed and secreted in a differentiation-dependent manner by C2C12 myoblasts and may, thus, be involved in the process of myoblast differentiation.
AB - Numerous cell types have been reported to secrete insulin-like growth factor-binding proteins (IGFBPs) in vitro. We have observed such IGFBPs in culture medium conditioned by the mouse myoblast cell line C2C12 and have identified IGFBP-2 among the secreted IGFBPs. Since C2C12 cells fuse in culture, these cells were used to examine IGFBP-2 mRNA expression and protein secretion during myogenic differentiation. Cells were harvested at approximately 80% of confluent density. Additional cultures were rinsed, fed differentiation medium, and harvested when approximately 15%, 60%, and 85% differentiated (fused). Northern and dot blot analyses were performed using total cellular RNA and a labeled cDNA specific for rat IGFBP-2. A single mRNA transcript of approximately 1.8 kilobases was observed. The level of expression of IGFBP-2 mRNA was highest in proliferating cells and decreased to 35%, 20%, and less than 10% of initial levels as differentiation progressed. Serum-free medium was conditioned for 24 h at each time point and collected from similar cultures. Three IGFBP species of 32,000, 30,000, and 24,000 mol wt (Mr) were detected in conditioned medium by probing Western blots with [125I]IGF-I (ligand blot analysis). The intensity of the 32,000 Mr band decreased with differentiation. These same blots were probed with an antibody raised against the 34,000 Mr bovine IGFBP-2. This antibody specifically bound to only the 32,000 Mr IGFBP. The level of antibody binding decreased by 50%, 90%, and nearly 100% as differentiation progressed. It, therefore, appears that IGFBP-2 is expressed and secreted in a differentiation-dependent manner by C2C12 myoblasts and may, thus, be involved in the process of myoblast differentiation.
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M3 - Article
C2 - 1370791
AN - SCOPUS:0026529604
SN - 0013-7227
VL - 130
SP - 607
EP - 615
JO - Endocrinology
JF - Endocrinology
IS - 2
ER -