Production of insulin-like growth factor II (IGF-II) and different forms of IGF-binding proteins by HT-29 human colon carcinoma cell line.
The serum-free medium conditioned by the human colon cancer cell line HT-29 contains insulin-like growth factors (IGF) that are entirely complexed to binding proteins (IGF-BP). Gel filtration in acid conditions of the cell-conditioned medium permits separation of IGF-BP from two molecular forms of IGF of 15,000 and 7,500 Mr. As determined by ligand blotting, IGF-BP are heterogeneous and constituted of three molecular forms of 31,000, 28,000, and 26,000 Mr. Using IGF-I and IGF-II radioreceptor assays, IGF-I radioimmunoassay (RIA), and competitive protein-binding assay specific for IGF-II, it is shown that the IGF-type eluting in 15 K and 7.5 K position from gel filtration is restricted to IGF-II. Its concentration is approximately 6 ng/10(6) HT-29 cells with 60% present as a high-molecular-weight form of IGF-II. This large 15 K IGF molecule is devoided of any IGF-binding activity and might represent incomplete processing of pro-IGF-II peptide. By contrast, the level of IGF-I detected by RIA is barely measurable and considered negligible (0.57 pg/10(6) HT-29 cells). Although these IGF-II-like peptides exhibit a growth-promoting activity on FR3T3 fibroblasts, they cannot stimulate, as recombinant IGF-I or IGF-II, 3H-thymidine incorporation into DNA of HT-29 cells, whatever the experimental conditions used. Finally, we have shown that IGF binding is restricted predominantly to the basolateral domain of the cell membrane by using HT-29-D4 clonal cells, derived from the parental HT-29 cell line, maintained in a differentiated state by culture in a medium in which glucose is replaced by galactose.
Laboratoire d'Immunologie, Faculté de Médecine, Marseille, France.
J M
JM
Culouscou
M
M
Remacle-Bonnet
F
F
Garrouste
J
J
Fantini
J
J
Marvaldi
G
G
Pommier
D000230:Adenocarcinoma / Q000378:metabolism / Q000473:pathology*; D002352:Carrier Proteins / Q000032:analysis / Q000378:metabolism*; D002471:Cell Transformation, Neoplastic / Q000187:drug effects / Q000378:metabolism / Q000473:pathology; D002850:Chromatography, Gel; D003110:Colonic Neoplasms / Q000378:metabolism / Q000473:pathology*; D003470:Culture Media / Q000032:analysis / Q000494:pharmacology; D004247:DNA / Q000096:biosynthesis; D005690:Galactose / Q000032:analysis / Q000494:pharmacology; D006801:Humans; D018969:Insulin-Like Growth Factor Binding Proteins; D007335:Insulin-Like Growth Factor II / Q000378:metabolism*; D013002:Somatomedins / Q000378:metabolism*; D014407:Tumor Cells, Cultured / Q000187:drug effects / Q000378:metabolism / Q000473:pathology
D002352:Carrier Proteins; D003470:Culture Media; D018969:Insulin-Like Growth Factor Binding Proteins; D013002:Somatomedins; D007335:Insulin-Like Growth Factor II; D004247:DNA; D005690:Galactose