Development of monoclonal antibodies against the human 3β-hydroxysteroid dehydrogenase/isomerase isozymes

- Monoclonal antibody against the human HSD3B1 enzyme was developed.
- Monoclonal antibody against the human HSD3B2 enzyme was developed.
- Each antibody was specific for the corresponding protein.
- Adrenal zona glomerulosa only expressed the HSD3B2 enzyme.

The human 3β-hydroxysteroid dehydrogenase/isomerase (HSD3B) enzymes catalyze the conversion of 3β-hydroxy Δ5-6 steroids into 3-keto Δ4-5 steroids, which is required for the synthesis of the mature steroid hormones secreted by the adrenal and gonads. The human has 2 isozymes, the HSD3B1 that is traditionally located in placenta and extra-adrenal tissues and the HSD3B2 that is expressed in the adrenal and gonads. Mice with both cryptochrome 1 and 2 genes deletion were recently found to have salt-sensitive hypertension and hyperaldosteronism. These deletions were also associated with overexpression of the Hsd3b6 enzyme, the homolog of the human HSD3B1, in the zona glomerulosa which was believed to explain the hyperaldosteronism. A report using antibodies against human HSD3B1 suggested that it was expressed in the zona glomerulosa of normal human adrenals and in patients with idiopathic hyperaldosteronism and the HSD3B2 expressed in both the zona fasciculata and glomerulosa. We have developed specific monoclonal antibodies against the human HSD3B1 and HSD3B2 isozymes and found that the main enzyme expressed in the zona glomerulosa was the HSD3B2. Faint staining of the adrenal was also obtained using the anti-HSD3B1antibody only at high concentrations of antibody. This study fails to confirm that HSD3B1 expression in the human zona glomerulosa and double immunofluorescence clearly shows that the HSD3B2 is expressed in the zona glomerulosa and fasciculata and in the zona glomerulosa HSD3B2 is co-expressed with aldosterone synthase (CYP11B2).