Increases in insulin-like growth factor binding protein-2 accompany decreases in proliferation and differentiation when porcine muscle satellite cells undergo multiple passages

Subjecting cloned porcine myogenic satellite cells to multiple passages leads to decreased rates of cell division and myotube formation. Because IGF have been implicated in the regulation of muscle cell proliferation and differentiation, the present study was conducted to characterize secretion of IGF-I and IGF-binding proteins (IGFBP) in cultures of cloned porcine satellite cells at two stages of multiple passaging. To this end, we obtained a single porcine satellite cell clone that demonstrated relatively high capacities for cellular proliferation and differentiation into myotubes at the fifth passage but that had greatly diminished capacities for proliferation and myotube formation by the seventh passage. The predominant IGFBP secreted by this satellite cell clone was immunologically identified as IGFBP-2, and quantities of it were increased in medium from seventh-passage cultures. Quantities of IGF-I in medium were determined with a newly developed "titration" radioimmunoassay in which interference from IGFBP was minimized by adding a range of saturating quantities of IGF-II. Medium IGF-I concentrations in seventh-passage cultures were also increased relative to the fifth-passage cultures when expressed per unit of DNA. It is hypothesized that the observed increase of IGF-I in medium likely resulted from protective sequestration of IGF-I by IGFBP-2 rather than from enhanced IGF-I secretion. In summary, these data suggest that multiple passaging of cloned porcine satellite cells results in increased secretion of IGFBP-2, which is associated with depressed cell proliferation and myotube formation, perhaps because the increased IGFBP-2 sequestered IGF-I and reduced its bioactivity.