The role of insulin-like growth factor-II (IGF-II) in the hypoglycemia associated with nonislet cell tumors is controversial. In this study we have addressed this question by measuring the IGF-II mRNA levels in extracts of these tumors. Hybridization of a 32P-labeled IGF-II cDNA to a Northern blot of RNA from three nonislet cell tumors associated with hypoglycemia (a hemangiopericytoma, fibrosarcoma, and malignant mesenchymal tumor) demonstrated six hybridizing bands, 6.8, 5.6, 4.7, 3.6, 2.6, and 2.1 kilobases in length. These bands were similar to those described by others in a range of tumors and normal tissues. Tissue IGF-II mRNA levels were quantitated using a solution hybridization/RNase protection assay. IGF-II mRNA levels in the tumors were similar to the level present in one line of human hepatoblastoma-derived Hep G2 cells, 5- to 6-fold higher than that in another line of Hep G2 cells, and 2- to 3-fold higher than that in term placenta. In contrast, little or no IGF-II mRNA was detected in a nonfunctioning islet cell adenoma or normal spleen. There was no evidence for amplification of the IGF-II gene in the one tumor in which it was sought. These data suggest that nonislet cell tumors associated with hypoglycemia produce large amounts of IGF-II mRNA and that this IGF-II mRNA appears to be the product of an IGF-II gene, which is apparently normal in the region encoding mature IGF-II peptide.