EXPRESSION OF 11-BETA-HYDROXYLASE (CYP11B1) AND ALDOSTERONE SYNTHASE (CYP11B2) IN THE HUMAN FETAL ADRENAL

OBJECTIVE: To understand better the steroidogenic capacity of the huma n fetal adrenal (HFA), we evaluated the expression of 11 beta-hydroxyl ase (CYP11B1) and aldosterone synthase (CYP11B2) in the fetal zone and neocortex of the HFA using a specific RNase protection assay. METHODS : Adrenal glands were obtained at the time of elective termination of pregnancy. Whole adrenals (n = 7) were frozen in liquid nitrogen, and subsequently total RNA extraction was performed by tissue homogenizati on followed by guanidinium/chloroform purification. In addition, RNA w as obtained from separated fetal zone (n = 4) and neocortex (n = 4) ti ssues obtained by dissection. RNase protection assays were then perfor med using radiolabeled complementary RNA probes generated by T7 RNA po lymerase directed against transcripts for CYP11B1, CYP11B2, and actin, the latter of which was used as a control for RNA integrity. Transcri pts also were examined using a reverse transcription polymerase chain reaction (RT-PCR) protocol specific for CYP11B1 or CYP11B2. RESULTS: T he RNase protection assay was designed to distinguish specific bands t hat corresponded to CYP11B1 (232 bp), CYP11B2 (262 bp), and actin (221 bp). RNA isolated from whole HFA was observed to have high levels of CYP11B1 transcript, whereas CYP11B2 was not detected. Dissected neocor tex and fetal zones were found to contain transcript for CYP11B1 using both the RNase protection assay and RT-PCR analysis. In contrast, usi ng the RNase protection assay, CYP11B2 mRNA was not observed in the RN A from the fetal zone, but after prolonged exposure there was a band c orresponding in size to CYP11B2 observed in RNA from the neocortex. Us ing the more sensitive RT-PCR method, transcript for CYP11B2 was found in both neocortex and fetal zone. CONCLUSION: The HFA expresses low l evels of CYP11B2 in accordance with its low production of mineralocort icoid. The expression of CYP11B1 in the fetal zone is intriguing becau se this enzyme is not necessary for the production of C-19 steroids. D efinition of the molecular mechanisms controlling expression of the CY P11B genes will be necessary to determine why the HFA differentially e xpresses these isoenzymes. Copyright (C) 1997 by the Society for Gynec ologic Investigation.