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Connective Tissue Growth Factor (CTGF) is a cysteine-rich peptide involved in human atherosclerosis and fibrotic disorders such as scleroderma. CTGF has considerable N-terminal sequence similarity with the insulin-like growth factor binding proteins (IGFBPs), including preservation of cysteines, and has been postulated to be a member of the IGFBP superfamily. Indeed, recent studies have shown that baculovirus generated CTGF, a secreted 38-kDa protein, binds IGFs in a specific manner, leading to the provisional renaming of CTGF as IGFBP-8 (or IGFBP-rP2). With immunoprecipitation and immunoblotting, using polyclonal anti-IGFBP-rP2 antibody generated against recombinant human IGFBP-rP2bac, IGFBP-rP2 can be identified in the serum-free conditioned media of Hs578T human breast cancer cells, as well as in various human biological fluids, such as normal sera, pregnancy sera, and cerebrospinal, amniotic, follicular and peritoneal fluids. Glycosylation studies with endoglycosidase F reveal that endogenous human IGFBP-rP2 is a secreted, glycosylated, approximately 32-38-kDa protein with 2-8-kDa of N-linked sugars and a 30-kDa core. There are 18- and 24-kDa proteins that appear to be IGFBP-rP2 degradation products. In Hs578T human breast cancer cells, transforming growth factor (TGF)-beta 2, a potent growth inhibitor for these cells, upregulates IGFBP-rP2 mRNA and protein levels. Expression of Hs578T IGFBP-rP2 is significantly increased by TGF-beta 2 treatment in a dose-dependent manner, with 2.5- and 6-fold increases in mRNA and protein levels, respectively, at a TGF-beta 2 concentration of 10 ng/ml. Our studies indicate that IGFBP-rP2 appears to be an important endocrine factor, and one of the critical downstream effectors of the critical downstream effectors of TGF-beta, similar to the role of IGFBP-3 in TGF-beta-induced growth inhibition in human breast cancer cells.

the journal of clinical endocrinology and metabolism/journal of clinical endocrinology and metabolism

Identification of Glycosylated 38-kDa Connective Tissue Growth Factor (IGFBP-Related Protein 2) and Proteolytic Fragments in Human Biological Fluids, and Up-Regulation of IGFBP-rP2 Expression by TGF-β in Hs578T Human Breast Cancer Cells