Pregnancy-associated plasma protein-A (PAPP-A) expression and insulin-like growth factor binding protein-4 protease activity in normal and malignant ovarian surface epithelial cells.
Pregnancy-Associated Plasma Protein-A (PAPP-A) proteolyses insulin-like growth factor binding protein-4 (IGFBP-4), thereby regulating local IGF availability. Reduced PAPP-A mRNA expression has been reported in ovarian cancer specimens compared to normal ovarian surface epithelial cells (OSE). To characterize PAPP-A expression and proteolytic activity in OSE, we developed a lifespan-extended human cell model using a temperature-sensitive mutant of the SV40 large T antigen (SV40LT). These OSE(tsT) cells proliferate at 34 degrees C (i.e., when SV40LT-positive), but not at 39 degrees C, a temperature at which the SV40LT is unstable (SV40LT-negative). Proteolysis of radiolabeled IGFBP-4 in conditioned media from OSE(tsT) lines was IGF-dependent and blocked by anti-PAPP-A antisera. Temperature shifts that eliminated stable SV40LT induced a 7-fold increase in PAPP-A mRNA and a 4-fold increase in protein. The converse experiment (shifting to SV40LT-positive conditions) resulted in decreased levels of PAPP-A mRNA but little change in PAPP-A protein. Nevertheless, there was a marked reduction in IGF-BP-4 proteolytic activity in medium of SV40LT-positive OSE-(tsT) cells. This decreased PAPP-A activity coincided with a nearly 20-fold increase in mRNA encoding a physiological inhibitor of PAPP-A, the precursor form of eosinophil Major Basic Protein (proMBP), and 4- to 5-fold increases in proMBP protein. Primary cultures of unmodified OSE expressed high levels of PAPP-A and undetectable proMBP, and therefore produced abundant IGFBP-4 protease activity. Short-term ovarian tumor cell cultures expressed variable levels of PAPP-A and high levels of proMBP, and consequently secreted little or no IGFBP-4 protease activity. The concurrent regulation of PAPP-A and its inhibitor, proMBP, suggests that IGFBP-4 proteolysis and local regulation of IGF availability may be altered in malignant ovarian epithelial cells.
Kalli
Kimberly R
KR
Division of Endocrinology, Metabolism and Nutrition, Endocrine Research Unit, Mayo Clinic College of Medicine, Rochester, MN 55905, USA. Kalli.kimberly@Mayo.edu
Chen
Bing-Kun
BK
Bale
Laurie K
LK
Gernand
Erica
E
Overgaard
Michael T
MT
Oxvig
Claus
C
Cliby
William A
WA
Conover
Cheryl A
CA
D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D015152:Blotting, Northern; D002455:Cell Division; D002460:Cell Line; D045744:Cell Line, Tumor; D002478:Cells, Cultured; D017077:Culture Media, Conditioned / Q000494:pharmacology; D016895:Culture Media, Serum-Free; D018076:DNA, Complementary / Q000378:metabolism; D004797:Enzyme-Linked Immunosorbent Assay; D004804:Eosinophils / Q000378:metabolism; D004847:Epithelial Cells / Q000378:metabolism*; D005260:Female; D006801:Humans; D007150:Immunohistochemistry; D018974:Insulin-Like Growth Factor Binding Protein 4 / Q000096:biosynthesis*; D008875:Middle Aged; D009154:Mutation; D010051:Ovarian Neoplasms / Q000473:pathology*; D010053:Ovary / Q000378:metabolism*; D011266:Pregnancy-Associated Plasma Protein-A / Q000096:biosynthesis* / Q000378:metabolism; D012313:RNA / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism; D013696:Temperature; D013997:Time Factors
D017077:Culture Media, Conditioned; D016895:Culture Media, Serum-Free; D018076:DNA, Complementary; D018974:Insulin-Like Growth Factor Binding Protein 4; D012333:RNA, Messenger; D012313:RNA; D011266:Pregnancy-Associated Plasma Protein-A