REGULATION OF ENDOTHELIAL IGFBP-3 SYNTHESIS AND SECRETION BY IGF-I AND TGF-BETA

We have examined the regulation of endothelial IGFBP-3 production by I GF-I and TGF-P, two growth factors thought to play major roles in the complications of diabetes mellitus. In addition, we developed a sensit ive method for IGFBP-3 mRNA quantitation by adapting the fluorescent m odification of the competitive PCR strategy. Our results using both No rthern analysis and the fluorescent competitive PCR method indicate th at: (1) IGFBP-3 mRNA is increased 2- to 10-fold by IGF-I and maximally reduced to 20% of control by TGF-P; (2) the changes in mRNA levels co rrelate with the levels of IGFBP-3 protein secreted into the media by these cells; (3) the induction of IGFBP-3 mRNA and protein by IGF-I an alogs was directly related to their ability to bind to the type I IGF receptor, reflecting an IGF-I receptor-mediated process; and (4) stead y state IGFBP-3 mRNA levels did not change significantly after a 6 h i ncubation with actinomycin D in the presence or absence of the growth factors suggesting that the observed IGF-I/TGF-beta effects occur at t he level of gene transcription rather than mRNA stability.