IGF Binding Protein-3 (IGFBP-3), the major IGF carrier in the blood, undergoes limited proteolysis which reduces its affinity for IGFs, thus facilitating dissociation. The functional effects of this at the cellular level were studied by comparing two serum pools, one from healthy adults, one from women during late pregnancy when IGFBP-3 proteolysis is increased. Sera were mixed to yield identical IGF-I and IGF-II concentrations in the two pools. Western ligand and immunoblotting gave the characteristic IGFBP patterns for the two types of serum. Both pools dose-dependently stimulated DNA synthesis in cultured chick embryo fibroblasts. Stimulation by pregnancy serum was twice that by normal serum at 0.05-0.2% concentrations (P < 0.001). In the presence of excess monoclonal anti-IGF-I and -II antibodies, stimulation by both (0.1-0.2%) pools was 70-80% reduced and residual stimulation was similar. Addition of recombinant human (rh)IGFBP-3 dose-dependently depressed both pools' activity, more so for normal serum at 25 and 50 ng/ml, equally for each at 100 ng/ml. At the latter concentration, slight proteolysis of the rhIGFBP-3 was detectable in the presence of 0.2% pregnancy serum, but at 25 ng/ml, proteolysis was absent. These results suggest that IGFs are released more readily from pregnancy serum, accounting for the weaker inhibitory effect of low rhIGFBP-3 concentrations. For identical IGF concentrations, pregnancy serum's greater biological activity therefore reflects greater IGF availability to the cells. This study demonstrates the functional consequences at cellular level of serum IGFBP-3 proteolysis, underlining its significance in regulating serum IGF bioavailability. (J. Clin.
Only half of colorectal‐cancer patients elicit serum antibodies in response to intratumoral p53‐gene mutations. Our study was designed to compare cellular events (p53‐protein accumulation and gene mutations) with the presence of circulating anti‐p53 antibodies (p53‐Ab). Thirty‐five colorectal‐cancer patients were studied for their intratumoral p53‐protein accumulation and circulating p53‐Ab. Tumour DNA was analyzed for genomic mutations in a sub‐set of 28 patients. In all, 18 tumours (51.4%) were positive by immunohistochemistry, and 17 tumour extracts were shown to contain “mutant” conformation p53 protein, 16 of them being were concordant by both methods. Of the 28 tumours tested by DGGE, 16 contained alterations in p53 exons 5 to 8 (57.1%). Of 12 tumours without detectable mutations, 10 were “mutant”‐conformation‐negative by immunohistochemistry and ELISA. Paradiploid tumors presented more frequently wild‐type p53 genes and were significantly less frequently immunohistochemistry‐ or p53‐Ab‐positive than polyploid tumors. Circulating p53‐Ab were detected in the serum of 11 patients (31%). In 9/11 cases, a gene mutation was found in the corresponding tumour. Three of four mutations in exon 8 and 3/3 mutations in exons 5‐6 were associated with p53‐Ab, in contrast with only 3/9 mutations in exon 7. We found good agreement in the detection of p53‐gene alterations by different methods. However, our data suggest that all gene mutations may not be equivalent in term of immunogenicity. Int. J. Cancer 81:712–718, 1999. © 1999 Wiley‐Liss, Inc.
We purified to homogeneity a growth inhibiting diffusible factor (IDF45) secreted by dense cultures of mouse 3T3 cells and which was able to inhibit 100% of DNA synthesis stimulated by serum in chick embryo fibroblasts (CEF) (Blat et al., 1989a). We then demonstrated that this factor was an IGF-binding protein (Blat et al., 1989b). Indeed, its N-terminal amino acid sequence was homologous to that of rat IGFBP-3. Our present results show that basic fibroblast growth factor (bFGF) induced, respectively, a fivefold and threefold increase in DNA synthesis in mouse embryo fibroblasts (MEF) and CEF. IDF-45 inhibited the stimulation induced by bFGF by about 65%, while stimulation induced by insulin, PDGF, or EGF was only weakly or not at all inhibited by IDF45. When bFGF stimulation was determined in the presence of a high concentration of insulin in conditions which minimize the effect of endogenous IGF-I or -II, this stimulation was decreased by about 50% in the presence of IDF45. This result suggests that addition of bFGF stimulates IGF secretion, thereby resulting in partial loss of inhibition, by IDF45, of bFGF stimulation.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.
hi@scite.ai
10624 S. Eastern Ave., Ste. A-614
Henderson, NV 89052, USA
Copyright © 2024 scite LLC. All rights reserved.
Made with 💙 for researchers
Part of the Research Solutions Family.