Decreased Expression of 3 Alpha-Hydroxysteroid Dehydrogenase in Primary Open Angle Glaucoma

Abstract

Abstract: : Purpose: Epidemiological data has long shown a relationship between POAG and sensitivity to the intraocular pressure raising effects of glucocorticoid therapy. We previously demonstrated altered cortisol metabolism in POAG derived cultured trabecular meshwork cells leading to increased formation of dihydrocortisol and decreased formation of the ocular hypotensive metabolite 3 alpha, 5 beta-tetrahydrocortisol. This change in metabolism was related to decreased activity of 3 alpha-hydroxysteroid dehydrogenase (3 alpha-HSD) in the POAG derived cells. Subsequently, peripheral blood lymphocytes (PBL) from POAG patients were also found to possess decreased 3 alpha-HSD activity [Exp Eye Res 62:39-45 (1996)], thereby providing an easily obtainable surrogate tissue for study. The following study was undertaken in order to determine whether the decreased 3 alpha-HSD enzyme activity in these POAG derived cells was due to decreased gene expression. Methods: PBL were isolated from POAG subjects whose PBL were previously shown to have decreased 3 alpha HSD activity and from nonPOAG controls. Homogenates from these PBL were used for Western blot analysis using 3 alpha-HSD antiserum. RNA was also isolated from these PBL and analyzed using quantitative RT-PCR with 3 alpha-HSD isoform specific primers. DNA sequencing of the 3 alpha-HSD generated cDNA was performed. Results: The POAG derived cells contained on average a 5 fold reduction of 3 alpha-HSD antigen (in relative units: 0.20, 0.17 and 0.01 for POAG cells as compared to 1.0, 0.79 and 0.45 for control cells). RT-PCR with 3 alpha-HSD isoform specific primers allowed the identification of 3 alpha-HSD type II (AKR1C3) as the predominant form expressed in these cells. There was no significant difference in the amount or base sequence of 3 alpha-HSD mRNA between four POAG and four nonPOAG derived specimens. The mechanism by which identical expression of 3 alpha-HSD mRNA in these cells gives rise to great differences in protein expression remains to be determined. Conclusions: These results strengthen the previously described relationship between decreased 3 alpha-HSD expression and the pathophysiology of POAG and may also serve as the basis of a useful blood test for those at risk for the disease.

Keywords: gene/expression • clinical (human) or epidemiologic studies: ris • enzymes/enzyme inhibitors