Inhibition of Hormone-Stimulated Steroidogenesis in Cultured Leydig Tumor Cells by a Cholesterol-Linked Phosphorothioate Oligodeoxynucleotide Antisense to Diazepam-Binding Inhibitor

• Published in: Proceedings of the National Academy of Sciences - PNAS Vol. 90; no. 12; pp. 5728 - 5731
• Main Authors: Boujrad, Noureddine, Hudson, James R., Papadopoulos, Vassilios
• Format: Journal Article
• Language: English
• Published: Washington, DC National Academy of Sciences of the United States of America 15-06-1993; National Acad Sciences
• Subjects: Animals; Base Sequence; Biological and medical sciences; Carrier Proteins - antagonists & inhibitors; Carrier Proteins - biosynthesis; Carrier Proteins - genetics; Cells; Cholesterol - pharmacology; Cholesterols; Chorionic Gonadotropin - pharmacology; Cyclic AMP - metabolism; Diazepam Binding Inhibitor; Dose-Response Relationship, Drug; Endocrinology; Fundamental and applied biological sciences. Psychology; Hormones; Hormones. Regulation; Humans; Hydroxycholesterols - pharmacology; Kinetics; Leydig Cell Tumor; Mice; Mitochondrial membranes; Molecular Sequence Data; Oligonucleotides, Antisense - pharmacology; Progesterone - biosynthesis; Proteins; Steroidogenesis; Steroids; Steroids - biosynthesis; Thionucleotides - pharmacology; Thyroid. Parathyroid. Ultimobranchial body; Tumor Cells, Cultured; Tumors; Ungulates; Vertebrates: endocrinology; Intracellular transport; Rodentia; Gonadotropin; Glycoprotein hormone; Metabolism; Testicle; Male genital system; HCG; Cholesterol; Leydig cell; Diazepam binding inhibitor; Vertebrata; Mitochondria; Mammalia; Mouse; Hormonal regulation; Steroidogenesis; Antisense DNA; Tumor cell
• ISSN: 0027-8424; 1091-6490
• DOI: 10.1073/pnas.90.12.5728

The polypeptide diazepam-binding inhibitor (DBI) has been previously shown to stimulate testicular Leydig, adrenocortical, and glial-cell mitochondrial steroidogenesis in vitro. To assess the in situ role of DBI in trophic hormone-stimulated steroidogenesis, we suppressed DBI levels in the hormone-responsive MA-10 Leydig tumor cells, using a cholesterol-linked phosphorothioate oligodeoxynucleotide (Chol-odN) antisense to DBI. Treating MA-10 cells with Chol-odN antisense to DBI resulted in a dose-dependent reduction of DBI levels (ED50= 1 μM). In contrast, Chol-odN sense to DBI did not affect its expression. Saturating amounts of human choriogonadotropin (hCG) increased MA-10 progesterone production by 150-fold. Addition of increased concentrations of Chol-odNs sense to DBI or of a nonrelated sequence did not reduce the MA-10 response to hCG. However, in the presence of Chol-odN antisense to DBI that could reduce DBI levels, MA-10 cells lost their ability to respond to hCG (ED50= 1 μM). In these studies the hCG-stimulated cAMP levels and cytochrome P450 side-chain cleavage activity, as measured by metabolism of 22(R)-hydroxycholesterol, were not affected by the Chol-odNs used. These observations provide unequivocal evidence that DBI plays a vital role in the acute stimulation of steroidogenesis by trophic hormones.