N-terminal truncated forms of insulin-like growth factor binding protein-3 in the peritoneal fluid of women without laparoscopic evidence of endometriosis

Abstract

To characterize and purify peritoneal mitogens able to stimulate the proliferation of human endometrial cells in vitro. Peritoneal fluids (PFs) from 50 patients were collected at laparoscopy and pooled (270 mL) for purification of mitogenic activity. University infertility clinic and endocrinology of reproduction and molecular endocrinology laboratories. Fifty subjects presenting for tubal ligation, pelvic pain, mass, or infertility but otherwise having no evidence of endometriosis inflammation, infection, or tumor. None. Assessment of mitogenic activity by 3 H-thymidine incorporation into mouse embryo fibroblasts and into primary cultures of isolated epithelial and stromal cells of human endometrium. The PF mitogens were purified successively on carboxymethyl-sepharose and heparin-sepharose columns followed by fractionation on cartridges of C 18 silica and reverse-phase high-performance liquid chromatography (HPLC). Four distinct bands were eluted from Sep-Pak, (Mississauga, Ontario, Canada) with molecular weights of 17 to 18, 20, 25, and 29 to 30 kd. The eluted fractions of Sep-Pak exerted preferential mitogenic activity on epithelial-derived human endometrial cells at an equimolar ratio with epidermal growth factor. Microsequencing of the 17 to 18, 20, 25, and 29 to 30 kd bands showed a homologous sequence with N- terminal amino acid sequences of insulin-like growth factor binding protein-3 (IGFBP-3). These data indicate that the PF of normal women without evidence of endometriosis contains AT-terminal truncated forms of IGFBP-3 that mediate an apparent preferential mitogenic action on epithelial-derived endometrial cells. Therefore, they could play a role in the ectopic growth of endometrial cells.